Clinical trial NCT01876212
Conditions: Metastatic Melanoma
Keywords: melanoma metastatic vaccine BRAF
Sponsor: Walter J. Storkus
Start date: 2014-05-16; Completion date: 2019-07-31
Enrollment: 15 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Current therapeutic approaches available for patients with advanced-stage melanoma remain inadequate, and existing approaches including those involving immunotherapy with cytokines and/or targeted strategies have resulted in disappointingly low rates of durable and complete responses. Correcting immune dysfunction in advanced-stage melanoma patients using tyrosine-kinase inhibitor (TKI) such as dasatinib is proposed to relicense the patient's immune system to respond optimally to specific immunization. The integration of antigens expressed by tumor-associated blood vessel cells provides a means to selectively target the genetically-/antigenically-heterogeneous population of tumor cells in the advanced-stage melanoma patient.

This is a single-center, prospective randomized Phase 2 trial evaluating the activity, safety and immune effects of dasatinib given in combination with an autologous type-1 polarized Dendritic Cell (αDC1) vaccine. The current trial represents a randomized Phase 2 study to determine the activity and safety of intradermal (id) administration of αDC1s loaded with a mixture of six TBVA-derived peptides at the time of, or immediately after, an initial therapy cycle with the TKI dasatinib.

Dasatinib will be administered at the standard dose and schedule recommended by the FDA (70 mg BID). The autologous type-I DC vaccine will be administered either prior to, or concomitant with, the initiation of dasatinib administration. All patients will receive dasatinib at a starting dose of 70 mg twice daily by mouth in the outpatient setting approximately every 12 hours, at the same time each day.

The DC vaccine will be administered by a single intradermal injection of approximately 10e7 cells, with all the DCs being administered on days 1 and 15 of every cycle on an outpatient basis in the University of Pittsburgh Clinical and Translational Research Center (UPCI-CTRC).

Patients on Arm A will start dasatinib administration on cycle 2, day 1 (week 5), while those patients in Arm B will start dasatinib administration on cycle 1, day 1 (week 1).

Men and women at least 18 years of age must be HLA-A2+ and have histologically confirmed melanoma that is metastatic (Stage IV) or unresectable Stage IIIB/C and for which standard curative or palliative measures do not exist or are no longer effective.

Note: The outcome measures and time frames (previously) described in the PRS protocol record have been revised and articulated in the results section, to more accurately describe and represent the stated per-protocol investigations and endpoints, quantitatively.

Detailed description:

Arms: arm 1: Patients will start vaccine on cycle 1, day 1 and dasatinib on cycle 2, day 1 (week 5).

All patients will receive dasatinib at a starting dose of 70 mg twice daily by mouth in the outpatient setting. Dasatinib will be supplied as 50 mg and 20 mg tablets. Patients will take 1 of the 50 mg tablets and 1 of the 20 mg tablets twice daily, approximately every 12 hours, at the same time each day.

The DC vaccine will be administered by a single intradermal injection of approximately 10e7 cells, with all the DCs being administered on days 1 and 15 of each cycle. The intradermal administration will be in the vicinity of the four nodal drainage groups of the four extremities. arm 2: Patients will start vaccine on cycle 1, day 1 and dasatinib on cycle 1, day 1.

All patients will receive dasatinib at a starting dose of 70 mg twice daily by mouth in the outpatient setting. Dasatinib will be supplied as 50 mg and 20 mg tablets. Patients will take 1 of the 50 mg tablets and 1 of the 20 mg tablets twice daily, approximately every 12 hours, at the same time each day.

The DC vaccine will be administered by a single intradermal injection of approximately 10e7 cells, with all the DCs being administered on days 1 and 15 of each cycle. The intradermal administration will be in the vicinity of the four nodal drainage groups of the four extremities.
Interventions: intervention 1: DC vaccine intervention 2: Dasatinib
Intervention descriptions: intervention 1: None intervention 2: None

Protocol documents:
Prot_SAP_000.pdf: 
 
A Randomized Phase 2 Pilot Study of Type I-Polarized Autologous Dendritic Cell Vaccines 
Incorporating Tumor Blood Vessel Antigen (TBVA)-Derived Peptides in Combination with 
Dasatinib in Patients with Metastatic Melanoma 
 
UPCI Protocol #: 
12-048 
BB-IND#: 
15,224 
Phase: 
2 
Version: 
10 
Version Date: 
07-14-17 
Investigational Agents: 
Alpha-type-1 Dendritic Cell Vaccine 
Dasatinib (NSC-732517) 
 
SPONSOR: 
Walter J. Storkus, PhD 
Professor of Dermatology and Immunology 
W1041.2 Biomedical Sciences Tower 
University of Pittsburgh School of Medicine 
200 Lothrop Street 
Pittsburgh, PA 15213 
Tel: 412-648-9981 
Fax: 412-383-5857 
Email: storkuswj@upmc.edu 
 
PRINCIPAL INVESTIGATOR: 
John Kirkwood, MD 
5115 Centre Avenue 
Pittsburgh, PA  15232 
Tel: 412-623-7707 
Fax: 412-648-6650 
E-mail: kirkwoodjm@upmc.edu 
 
Biostatistician: 
Yan Lin, PhD 
Research Assistant Professor 
Department of Biostatistics, Graduate School of Public Health 
University of Pittsburgh and Biostatistics Facility 
University of Pittsburgh Cancer Institute 
Sterling Plaza, Suite 325 
201 North Craig Street 
Pittsburgh, PA 15213 
Tel: 412-383-1311 
Fax: 412-383-1535 
Email: yal14@pitt.edu 
 
Immunologic Monitoring and 
Cellular Products Laboratory: 
 
Lisa H. Butterfield, PhD, Director 
Associate Professor of Medicine, Surgery and Immunology 
Director, UPCI Immunologic Monitoring and Cellular Products 
Laboratory  
Hillman Cancer Center 
5117 Centre Avenue, Suite 1.27 
Pittsburgh, PA  15213 
 
 
 


Source(s) of Support: BMS 
 
Study Monitor: 
Education and Compliance Office for Human Subject Research 
Research Conduct and Compliance Office 
University of Pittsburgh 
3500 Fifth Avenue, Suite 205 
Pittsburgh, PA 15213 


SCHEMA 
 
 
 


TABLE OF CONTENTS 
SCHEMA ..................................................................................................................................................... 3 
LIST OF ABBREVIATIONS .................................................................................................................... 7 
1. 
OBJECTIVES .................................................................................................................................. 1 
1.1 
Primary Objectives .................................................................................................................... 1 
1.2 
Secondary Objectives ................................................................................................................ 1 
2. 
BACKGROUND ............................................................................................................................. 1 
2.1 
Melanoma .................................................................................................................................. 1 
2.2 
Dasatinib .................................................................................................................................... 2 
2.2.1 
Preclinical Anti-tumor Activity .........................................................................................................2 
2.2.1.1 
In Vitro Molecular Studies ................................................................................................ 2 
2.2.1.2 
Cellular Studies .................................................................................................................. 3 
2.2.1.3 
In Vivo Studies .................................................................................................................. 3 
2.2.1.4 
Preclinical Toxicology ....................................................................................................... 3 
2.2.2 
Clinical Pharmacokinetics .................................................................................................................3 
2.2.2.1 
Absorption ......................................................................................................................... 4 
2.2.2.2 
Distribution ........................................................................................................................ 4 
2.2.2.3 
Metabolism ........................................................................................................................ 4 
2.2.2.4 
Elimination ........................................................................................................................ 4 
2.2.3 
Clinical Experience with Dasatinib in CML and Ph+ ALL ..............................................................4 
2.2.4 
Phase 1 Experience of Dasatinib in Solid Tumors other than Melanoma .........................................6 
2.2.5 
Phase 2 Experience of Dasatinib in Melanoma Patients ...................................................................6 
2.3 
Potential for Vaccine-Induced CD8+ T cell Targeting of Tumor Antigens versus Tumor Blood 
Vessel Antigens ......................................................................................................................... 7 
2.4 
Immune Dysfunction in Patients with Advanced-Stage Solid Cancers .................................... 8 
2.5 
Dendritic Cells ......................................................................................................................... 10 
2.6 
Type-1 Polarized DC ............................................................................................................... 10 
2.7 
Improved Anti-Tumor Efficacy for Combined Dasatinib + Vaccine Therapy........................ 11 
3. 
STUDY RATIONALE .................................................................................................................. 14 
3.1 
Importance and Rationale for the Trial ................................................................................... 14 
3.2 
Rationale for Vaccine Dose, Route of Administration and Schedule ..................................... 15 
4. 
STUDY DESIGN .......................................................................................................................... 16 
4.1 
Leukapheresis .......................................................................................................................... 16 
4.2 
Vaccine .................................................................................................................................... 17 
4.2.1 
Formulation ..................................................................................................................................... 17 
4.2.2 
Storage and preparation .................................................................................................................. 17 
4.2.3 
Administration ................................................................................................................................ 17 
4.2.4 
Anticipated Adverse Events (Vaccine) ........................................................................................... 17 
4.3 
Dasatinib .................................................................................................................................. 18 
4.3.1 
Packaging and Labeling .................................................................................................................. 18 
4.3.2 
Storage ............................................................................................................................................ 18 
4.3.3 
Handling and Dispensing ................................................................................................................ 18 
4.3.4 
Drug Ordering and Accountability .................................................................................................. 19 
4.3.5 
Safety of Dasatinib in Clinical Studies in CML and Ph+ ALL ....................................................... 19 
4.3.6 
Laboratory Abnormalities ............................................................................................................... 21 
4.3.7 
Anticipated Adverse Events (Dasatinib) ......................................................................................... 21 
4.3.7.1 
Myelosuppression ............................................................................................................ 21 
4.3.7.2 
Bleeding Related Events .................................................................................................. 21 
4.3.7.3 
Fluid Retention ................................................................................................................ 22 
4.3.7.4 
QT Prolongation .............................................................................................................. 22 


5. 
STUDY TREATMENT PLAN ...................................................................................................... 22 
5.1 
Dasatinib Administration ........................................................................................................ 22 
5.2 
Vaccine administration ............................................................................................................ 23 
5.3 
Duration of Study Treatment ................................................................................................... 23 
5.4 
Duration of Follow Up ............................................................................................................ 24 
6. 
STUDY CALENDAR ................................................................................................................... 24 
7. 
DOSE-LIMITING TOXICITIES ................................................................................................... 25 
7.1 
Definition of Dose-Limiting Toxicity ..................................................................................... 25 
7.2 
Dasatinib Dosing delays/modifications ................................................................................... 26 
7.2.1 
Grades 1 or 2 hematologic or non-hematologic toxicities ............................................................... 26 
7.2.2 
Grades 3 or 4 hematologic or non-hematologic toxicities ............................................................... 26 
7.3 
Vaccine Dosing delays/modifications ..................................................................................... 26 
8. 
PATIENT SELECTION ................................................................................................................ 26 
8.1 
Inclusion Criteria ..................................................................................................................... 26 
8.2 
Exclusion Criteria .................................................................................................................... 27 
8.3 
Inclusion of Women and Minorities ........................................................................................ 28 
8.4 
General Concomitant Medication and Supportive Care Guidelines........................................ 28 
8.4.1 
Bisphosphonates ............................................................................................................................. 29 
8.4.2 
Antacids .......................................................................................................................................... 29 
8.4.3 
Medications that prolong the QT interval ....................................................................................... 29 
8.4.4 
Medications that affect CYP3A4 .................................................................................................... 29 
9. 
RESEARCH SAMPLES ................................................................................................................ 30 
9.1 
Biopsy tissue ........................................................................................................................... 30 
9.2 
Blood samples ......................................................................................................................... 31 
10. 
CORRELATIVE STUDIES .......................................................................................................... 31 
10.1 
Immune Monitoring Analysis of TBVA-specific CD8+ T cell responses (Primary Endpoint).
 ................................................................................................................................................. 31 
10.2 
Quantitation of CD8+ T cells, Treg, MDSC and blood vessels in melanoma biopsy tissue .... 32 
10.3 
Treg analysis in PBMC ............................................................................................................. 32 
10.4 
MDSC Analysis in PBMC ...................................................................................................... 33 
10.5 
EphA2 Protein Levels in tumor biopsies ................................................................................. 33 
10.6 
CXCL10 Levels in Patient Serum ........................................................................................... 33 
11. 
MEASUREMENT OF EFFECT .................................................................................................... 33 
11.1 
Antitumor Effect...................................................................................................................... 34 
11.1.1 
Definitions....................................................................................................................................... 34 
11.1.2 
Disease Parameters ......................................................................................................................... 34 
11.1.3 
Methods for Evaluation of Measurable Disease.............................................................................. 35 
11.1.4 
Response Criteria ............................................................................................................................ 38 
11.1.4.1 
Evaluation of Target Lesions ......................................................................................... 38 
11.1.4.2 
Evaluation of Non-Target Lesions ................................................................................. 38 
11.1.4.3 
Evaluation of Best Overall Response ............................................................................ 38 
11.1.5 
Duration of Response ...................................................................................................................... 39 
11.1.6 
Progression-Free Survival ............................................................................................................... 40 
12. 
STATISTICAL CONSIDERATIONS ........................................................................................... 40 
12.1 
Study Design ........................................................................................................................... 40 
12.2 
Safety Monitoring.................................................................................................................... 40 
12.3 
Sample Size/Accrual Rate ....................................................................................................... 41 
12.4 
Data Analysis .......................................................................................................................... 41 
12.4.1 
Analysis Sets ................................................................................................................................... 41 
12.4.2 
Baseline Characteristics .................................................................................................................. 41 
12.4.3 
Safety Profile .................................................................................................................................. 41 


12.4.4 
Efficacy Analysis ............................................................................................................................ 42 
12.5 
Reporting and Exclusions ........................................................................................................ 42 
12.5.1 
Evaluation of toxicity ...................................................................................................................... 42 
12.5.2 
Evaluation of response .................................................................................................................... 42 
13. 
DATA SAFETY MONITORING PLAN ...................................................................................... 43 
14. 
ADVERSE EVENT REPORTING ................................................................................................ 44 
14.1 
Adverse event definitions ........................................................................................................ 44 
14.2 
Recording/Reporting requirements ......................................................................................... 45 
14.2.1 
Eliciting AE Information ................................................................................................................ 45 
14.2.2 
Recording Requirements ................................................................................................................. 45 
14.2.2.1 
Abnormal Test Findings ................................................................................................ 46 
14.2.2.2 
Causality and severity assessment ................................................................................. 46 
14.3 
Reporting of Suspected Adverse Reactions ............................................................................ 46 
14.4 
Review of Safety Information: Sponsor Responsibilities[1] ................................................... 47 
14.5 
Reporting of Adverse Reactions to the FDA ........................................................................... 47 
14.5.1 
Written IND safety reports .............................................................................................................. 47 
14.5.2 
Telephoned IND safety reports – fatal or life-threatening suspected adverse reactions.................. 48 
14.6 
Reporting adverse events to the responsible IRB .................................................................... 48 
15. 
WITHDRAWAL OF SUBJECTS DUE TO ADVERSE EVENTS .............................................. 48 
16. 
QUALITY CONTROL AND QUALITY ASSURANCE ............................................................. 49 
17. 
DATA HANDLING AND RECORD-KEEPING ......................................................................... 49 
18. 
ETHICS ......................................................................................................................................... 49 
18.1 
Institutional Review Board (IRB) Approval ........................................................................... 49 
18.2 
Ethical and Scientific Conduct of the Clinical Study .............................................................. 50 
18.3 
Subject Informed Consent ....................................................................................................... 50 
19. 
REFERENCES .............................................................................................................................. 51 
 


LIST OF ABBREVIATIONS 
AE 
Adverse event 
ANC 
Absolute Neutrophil Count 
BID 
Twice a Day 
BMS 
Bristol-Myers Squibb Company 
CBC 
Complete Blood Count 
CTCAE 
Common Terminology Criteria for Adverse Events 
DLT 
Dose Limiting Toxicity 
DSMB 
Data Safety Monitoring Board 
ECOG 
Eastern Cooperative Oncology Group 
EKG 
Electrocardiogram 
ESR 
Expedited Safety Report 
FDA 
Food and Drug Administration 
FSH 
Follicle-Stimulating Hormone 
GCP 
Good Clinical Practice 
HCG 
Human Chorionic Gonadotropin 
HIPAA 
Health Insurance Portability and Accountability Act 
HRT 
Hormone Replacement Therapy 
IB 
Investigators’ Brochure 
ICH 
International Conference on Harmonisation 
IEC 
Independent Ethics Committee 
IND 
Investigational New Drug (Application) 
IRB 
Institutional Review Board 
NCI 
National Cancer Institute 
NSAE 
Non-Serious Adverse Event 
PFS 
Progression Free Survival 
PO 
By Mouth 
PR 
Partial Response 
QD 
Once Daily 
QoL 
Quality Of Life 
SAE 
Serious Adverse Event 
SUSAR 
Suspected Unexpected Serious Adverse Reaction 
ULN 
Upper Limit of Normal 
WBC 
White Blood Count 
WOCBP 
Women of Child-Bearing Potential 


 
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1. 
OBJECTIVES 
1.1 
Primary Objectives 
To determine the effect of the addition of dasatinib on the proportion of HLA-A2+ melanoma 
patients that exhibit improved peripheral blood CD8+ T cell responses against 3 or more peptide 
epitopes after active vaccination with Type I-polarized autologous dendritic cell (DC1) vaccine 
incorporating 6 tumor blood vessel-associated antigen (TBVA)-derived peptides. 
1.2 
Secondary Objectives 
 
To determine the safety and feasibility of a regimen consisting of the combination of 
dasatinib and vaccine. 
 
To determine the objective response rate by RECIST 1.1 of melanoma patients treated 
with the combination of dasatinib and vaccine. 
 
To determine the progression-free and overall survival of patients treated with the 
combination of dasatinib and vaccine. 
 
To determine the quantitative effect of treatment on the following immunological 
endpoints; a) number of CD8+ T cells infiltrating into melanoma lesions; b) number of 
suppressor cell populations and blood vessels in melanoma tumor biopsies; c) number of 
suppressor cell populations in patients peripheral blood, d) level of EphA2 protein 
expression in tumor biopsies, and e) serum concentration of the T cell-recruiting 
chemokine CXCL10/IP-10. 
2. 
BACKGROUND 
2.1 
Melanoma 
The incidence of melanoma continues to rise at an alarming pace. The American Cancer Society 
estimates that over 70,000 patients will be diagnosed with melanoma in 2011 with an estimated 
death toll of over 8,700 deaths (1). Early stages of the disease are surgically curable and adjuvant 
therapy of high risk disease is modestly effective in preventing recurrent metastatic disease. 
Once metastatic, melanoma is incurable with very high disease-fatality rates (2). Few agents 
have shown clinical benefit in this setting and none has shown a survival advantage, until 
ipilimumab, most recently with the first ever metastatic melanoma trial to show improvement of 
overall survival leading to FDA approval in March 2011. Hydroxyurea was the first agent to get 
FDA approval in 1967 followed by dacarbazine in 1971 which quickly became the “standard of 
care” as no other agent, singly or in combination, achieved a survival advantage over 
dacarbazine (3-5). Immunotherapy has achieved limited success with high-dose bolus IL-2 
leading to durable responses in a small subset of patients (5-15%; ref. 6). The approval of 
ipilimumab is likely to alter the therapeutic landscape of metastatic melanoma but it should be 
noted that only 10-15% of patients respond and not all achieve durable responses (7). Even more 
promise is associated with the advent of highly selective BRAF-targeted agents that have led to 


 
UPCI 12-048 
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02-15-2016 
the highest response rates seen in melanoma in patients that harbor the BRAF mutation (8). 
While this is clearly encouraging, to date no durable responses have been described with BRAF 
inhibitors and those are only available for the 40-50% of patients that have the mutation. 
Therefore there continues to be an urgent need to develop therapeutic strategies that offer long 
term control of the disease. 
Mutation-directed therapy has expanded the therapeutic opportunities for c-KIT-mutated 
melanomas. Unlike melanomas that occur in chronic sun-damaged areas, melanomas arising in 
non-sun exposed areas such as the mucosal surfaces, palms, soles, and nail beds are thought to 
result from different risk factors and perhaps different oncogenes. Indeed, melanomas with 
oncogenic mutations in c-KIT have been reported in several studies. Recently, c-KIT-activating 
mutations were reported in 21% of mucosal melanomas, 11% of acral melanomas, and 16.7% of 
melanomas arising in chronically sun-damaged skin (9-13). 
2.2 
Dasatinib 
Dasatinib (BMS-354825; NSC-732517) is a potent, broad spectrum ATP-competitive inhibitor 
of 5 critical oncogenic tyrosine kinases/kinase families: BCR-ABL, SRC, c-KIT, PDGFR, and 
ephrin (Eph) receptor kinases. Overexpression or activation of these kinases plays a critical role 
in the etiology of various cancer types, as well as the malignant behavior associated with these 
diseases, such as unregulated proliferation and metastasis. The Philadelphia chromosome and 
resultant constitutively expressed BCR-ABL protein tyrosine kinase (PTK) is present in > 90-
95% of patients with CML and 20% to 30% of adult patients with ALL. BCR-ABL activity is 
required for the cancer-causing ability of this protein. Dasatinib is ~500-fold more potent than 
imatinib in inhibiting BCR-ABL by binding to both active and inactive conformations of c-ABL, 
whereas imatinib only binds to the inactive state. This difference in binding may be responsible 
for the increased potency of dasatinib over imatinib (14). 
Dasatinib is indicated for the treatment of newly diagnosed adults with CML in chronic phase, 
the treatment of adults with chronic, accelerated, myeloid or lymphoid blast phase CML with 
resistance or intolerance to prior therapy including imatinib, and the treatment of adults with Ph+ 
ALL with resistance or intolerance to prior therapy (14). 
2.2.1 
Preclinical Anti-tumor Activity 
2.2.1.1 In Vitro Molecular Studies 
Dasatinib competes with ATP for the ATP-binding site in the kinase domain of selected protein 
tyrosine kinases (PTKs) and has been shown to inhibit at least five protein tyrosine 
kinases/kinase families: SRC family kinases, BCR-ABL, c-KIT, EphA2 and the PDGFR 
receptor. Dasatinib is much more potent than imatinib mesylate. Dasatinib was 260-, 8-, 60-, and 
>1000-fold more potent than imatinib versus BCR-ABL, c-KIT, PDGFR, and SRC kinases, 
respectively (14). 


 
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2.2.1.2 Cellular Studies 
Dasatinib inhibits the BCR-ABL kinase with an in vitro IC50 of 3 nM, a potency that was 
260-fold greater than that of imatinib mesylate (IC50 = 790 nM). In cellular assays, dasatinib 
killed or inhibited the proliferation of all BCR-ABL dependent leukemic cell lines tested to date. 
Dasatinib also demonstrated undiminished antitumor activity against several preclinically- and 
clinically-derived models of imatinib mesylate resistance. These results demonstrate that 
dasatinib is effective in reducing the proliferation or survival of both imatinib mesylate-sensitive 
and resistant cells, and its inhibitory activity is not solely dependent on BCR-ABL. 
2.2.1.3 In Vivo Studies 
The activity of dasatinib against CML cells in vitro was reproduced in vivo against several 
human CML xenograft models grown subcutaneously in SCID mice. Against the K562/imatinib 
mesylate/R CML model, dasatinib was curative in 100% of the treated animals. In contrast, at its 
optimal dose and schedule, imatinib mesylate was inactive. 
Dasatinib exhibited in vivo antitumor activity in a broad spectrum of solid tumor types including 
the rhabdomyosarcoma line RD1 implanted in nude mice. The activity appeared to be from a 
cytostatic rather than cytotoxic effect. 
2.2.1.4 Preclinical Toxicology 
Single or repeated oral administration of dasatinib principally affected the gastrointestinal (GI) 
tract, including the liver, the hematopoietic and lymphoid systems in rats and monkeys. Other 
prominent effects after single oral administration of dasatinib included renal and cardiac toxicity 
in rats at lethal doses, and cutaneous hemorrhage in monkeys. Dasatinib can also affect the 
immune system and bone turnover. 
Dasatinib in vitro activity in the HERG/IKr and Purkinje-fiber assays indicated a moderate 
liability for prolongation of cardiac ventricular repolarization (QT interval) in the clinic. 
However, there were no dasatinib-related changes observed in electrocardiograms, nervous 
system function, respirations and heart rate, blood pressure, or arterial oxygen saturation in 
single-dose, 10-day, or 1-month oral toxicity studies in monkeys. 
Dasatinib was found to exhibit a profile of broad-spectrum platelet inhibition best typified by 
anti-platelet agents such as the GPIIb/IIIa antagonists, integrelin and abciximab. 
Finally, modulation of SRC kinase activity could also affect osteoclast morphology and function 
and bone remodeling. This effect could potentially result in an increase in bone mineral density 
and a phenotype analogous to osteopetrosis (14). 
2.2.2 
Clinical Pharmacokinetics 
The pharmacokinetics of dasatinib has been evaluated in 229 healthy patients and in 137 patients 
with leukemia. 


 
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02-15-2016 
2.2.2.1 Absorption 
Maximum plasma concentrations (Cmax) of dasatinib are observed between 0.5 and 6 hours 
(Tmax) following oral administration. Dasatinib exhibits dose proportional increases in AUC and 
linear elimination characteristics over the dose range of 15 mg to 240 mg/day. The overall mean 
terminal half-life of dasatinib is 3-5 hours (14).  
Data from a study of 54 healthy patients administered a single, 100-mg dose of dasatinib 
30 minutes following consumption of a high-fat meal resulted in a 14% increase in the mean 
AUC of dasatinib. The observed food effects were not clinically relevant.  
2.2.2.2 Distribution 
In patients, dasatinib has an apparent volume of distribution of 2505 L, suggesting that the drug 
is extensively distributed in the extravascular space. Binding of dasatinib and its active 
metabolite to human plasma proteins in vitro was approximately 96% and 93%, respectively, 
with no concentration dependence over the range of 100-500 ng/mL (14). 
2.2.2.3 Metabolism 
Dasatinib is extensively metabolized in humans, primarily by the cytochrome P450 enzyme 3A4. 
CYP3A4 was the primary enzyme responsible for the formation of the active metabolite. Flavin-
containing monooxygenase 3 (FMO-3) and uridine diphosphate-glucuronosyl transferase (UGT) 
enzymes are also involved in the formation of dasatinib metabolites. In human liver microsomes, 
dasatinib was a weak time-dependent inhibitor of CYP3A4.  
The exposure of the active metabolite, which is equipotent to dasatinib, represents approximately 
5% of the dasatinib AUC. This indicates that the active metabolite of dasatinib is unlikely to play 
a major role in the observed pharmacology of the drug. Dasatinib also had several other inactive 
oxidative metabolites.  
2.2.2.4 Elimination 
Elimination is primarily via the feces. Following a single oral dose of [14C]-labeled dasatinib, 
approximately 4% and 85% of the administered radioactivity was recovered in the urine and 
feces, respectively, within 10 days. Unchanged dasatinib accounted for 0.1% and 19% of the 
administered dose in urine and feces, respectively, with the remainder of the dose being 
metabolites. 
2.2.3 
Clinical Experience with Dasatinib in CML and Ph+ ALL 
Four single-arm multicenter studies were conducted to determine the efficacy and safety of 
dasatinib in patients with CML or Philadelphia chromosome-positive acute lymphoblastic 
leukemia (Ph+ ALL) resistant to or intolerant of treatment with imatinib. Resistance to imatinib 
included failure to achieve a complete hematologic response (within 3–6 months) or major 
cytogenetic response (by month 12) or progression of disease after a previous cytogenetic or 


 
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02-15-2016 
hematologic response. Imatinib intolerance included inability to tolerate 400 mg or more of 
imatinib per day or discontinuation of imatinib because of toxicity. The studies are ongoing. The 
results are based on a minimum of 6 months follow-up after the start of dasatinib therapy. Most 
patients had long disease histories with extensive prior treatment, including imatinib, cytotoxic 
chemotherapy, interferon, and stem cell transplant. The maximum imatinib dose had been 400–
600 mg/day in about one-half of the patients and > 600 mg/day in the other half (14). 
All patients were treated with dasatinib 70 mg, twice a day, on a continuous basis. The median 
durations of treatment were between 2.8 - 5.6 months (14). 
The primary efficacy endpoint in chronic phase CML was major cytogenetic response (MCyR), 
defined as elimination (complete cytogenetic response, CCyR) or substantial diminution (by at 
least 65%, partial cytogenetic response) of Ph+ hematopoietic cells. The primary endpoint in 
accelerated phase, myeloid blast phase, and lymphoid blast phase CML, and Ph+ ALL was major 
hematologic response (MaHR), defined as either a complete hematologic response or no 
evidence of leukemia as defined in Table 1. 
Most cytogenetic responses occurred after 12 weeks of treatment, when the first cytogenetic 
analyses were performed. Hematologic and cytogenetic responses were stable during the 6-
month follow-up of patients with chronic phase, accelerated phase, and myeloid blast phase 
CML. The median durations of major hematologic response were 3.7 months in lymphoid blast 
CML and 4.8 months in Ph+ ALL.  
There were no age- or gender-related response differences (14). 
Table 1: Efficacy in Dasatinib Clinical Studies in CML and Ph+ ALL (All Treated Populations)a 
 
Chronic 
(n=186) 
Accelerated 
(n=107) 
Myeloid 
Blast 
(n=74) 
Lymphoid 
Blast 
(n=42) 
Ph+ ALL 
(n=36) 
Hematologic 
Response Rateb (%) 
MaHR (95% CI) 
n/a 
59 (49–68) 
32 (22–44) 
31 (18–47) 
42 (26–59) 
 
CHR (95% CI) 
90 (85–94) 
33 (24–42) 
24 (15–36) 
26 (14-42) 
31 (16–48) 
 
NEL (95% CI) 
n/a 
26 (18–36) 
8 (3–17) 
5 (0.6–16) 
11 (3.1–26) 
Cytogenetic  
Responsec (%) 
MCyR (95% CI) 
45 (37–52) 
31 (22–41) 
30 (20–42) 
50 (34–66) 
58 (41–74) 
 
CCyR (95% CI) 
33 (26–40) 
21 (14–30) 
27 (17–39) 
43 (28–59) 
58 (41–74) 
a Numbers in bold font are the results of primary endpoint. 
b Hematologic response criteria (all responses confirmed after 4 weeks): 
Major hematologic response: (MaHR) = complete hematologic response (CHR) + no evidence of leukemia (NEL).  
CHR (chronic CML): WBC  institutional ULN, platelets <450,000/mm3, no blasts or promyelocytes in peripheral blood, <5% myelocytes 
plus metamyelocytes in peripheral blood, basophils in peripheral blood  institutional ULN, and no extramedullary involvement. 


 
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CHR (advanced CML/Ph+ ALL): WBC  institutional ULN, ANC 1000/mm3, platelets ≥100,000/mm3, no blasts or promyelocytes in 
peripheral blood, bone marrow blasts 5%, <5% myelocytes plus metamyelocytes in peripheral blood, basophils in peripheral blood 
 institutional ULN, and no extramedullary involvement. 
NEL: same criteria as for CHR but ANC 500/mm3 and <1000/mm3, and/or platelets 20,000/mm3 and 100,000/mm3. 
c Cytogenetic response criteria: complete (0% Ph+ metaphases) or partial (>0%–35%). MCyR (0%–35%) combines both complete and partial 
responses. 
n/a = not applicable. 
 
2.2.4 
Phase 1 Experience of Dasatinib in Solid Tumors other than Melanoma 
In a Phase 1 study (CA180003) conducted by Bristol-Myers Squibb (BMS), dasatinib was 
administered on a twice a day schedule to 42 patients with refractory solid tumor. To date, doses 
up to 160 mg, twice a day, on a 5-day on/2-day off schedule have been administered. A dose of 
120 mg, twice a day, continuous daily schedule is currently under investigation.  
No severe clinical toxicity has been encountered. Gastrointestinal symptoms were reported in 
most patients, fatigue was reported in 17 patients (40%) and rash in 10 patients (24%). Edema, 
lethargy and headache were uncommon and appear to be dose-related. Grade 3 asymptomatic 
hypocalcemia was considered dose-limiting in one subject, Grade 2 rash was considered dose-
limiting in two other patients, and Grade 2 nausea and vomiting (with dysarthria, 
lightheadedness and lethargy in a 49 kg subject taking concurrent diazepam) was considered 
dose-limiting in one subject. 
In another Phase 1 study (CA180021), dasatinib was administered on a once daily schedule to 24 
patients at doses up to 180 mg. Pleural effusions were observed in three patients at the 180 mg 
dose level (one with pneumonia and two with malignant effusion). A dose of 250 mg, once daily, 
is currently under consideration. Hypocalcemia, GI symptoms, and skin rash have been mild and 
infrequent. 
To date, the safety profile in solid tumor patients has been similar to that in chronic phase CML 
patients with the exception of severe myelosuppression, which has not been observed in solid 
tumor patients and is considered related to efficacy against the leukemia as noted above, and 
severe bleeding which is secondary to thrombocytopenia in most instances (14). 
2.2.5 
Phase 2 Experience of Dasatinib in Melanoma Patients 
Kluger et al. (15) have recently reported results of their Phase 2 study in which 39 patients with 
advanced-stage, chemotherapy-naïve, unresectable melanoma received dasatinib twice daily in 
order to assess treatment impact on objective response rates and progression-free survival (PFS), 
as well as, any associated toxicities. The initial dosing of 100 mg, twice daily, was reduced to 70 
mg, twice daily, after treating 17 patients due to dose-limiting toxicities including fatigue, 
dypsea, and pleural effusion. Two patients exhibited partial response to therapy, while an 
additional 3 patients displayed minor responses, with a median PFS of 8 weeks and 13% of 
patients with a PFS of at least 6 months. Although the study failed to meet its clinical endpoints 
of a 30% response rate or 6-month PFS, and it displayed minimal efficacy as a single agent in 
unselected patients, the Kluger study suggested potential biased activity of dasatinib in patients 


 
UPCI 12-048 
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02-15-2016 
without c-KIT mutations. This suggestion will be further inspected in an ongoing Eastern 
Cooperative Oncology Group (ECOG) Phase 2 study investigating the activity of dasatinib in 
patients with c-KIT mutated melanoma. Given the possibility of reduced clinical benefit for 
dasatinib in patients with c-KIT mutant melanomas, and in the absence of additional insights 
from the open ECOG trial, we have chosen to exclude such patients in this protocol. 
Regardless of the melanoma patient cohort selected for inclusion, however, Hersey et al. (16) 
suggest that receptor tyrosine kinases like dasatinib will exert greatest clinical benefit when 
integrated in combination therapy designs, such as the current dasatinib + vaccination schema. 
2.3 
Potential for Vaccine-Induced CD8+ T cell Targeting of Tumor Antigens versus 
Tumor Blood Vessel Antigens 
The finding of T lymphocyte infiltrates in melanomas and other solid tumor types (17-19) 
resulted in a series of studies to evaluate the adoptive transfer of patient-derived T cells in 
customized adoptive immunotherapy (AIT) approaches. Tumor-infiltrating lymphocytes (TIL) 
isolated from cancer patients were expanded to large numbers ex vivo using either rIL-2 or rIL-
2/rIL-4 culture supplementation (20). These lymphocytes were then administered intravenously 
(IV) in combination with systemic rIL-2 therapy. However, the adoptive transfer of TIL 
appeared no more effective than rIL-2 alone in promoting clinical benefit (20, 21). 
Studies performed using such T lymphocytes provided one of the first demonstrations of a T cell 
response to human melanoma through the development of a series of T cell lines. CD8+ T cell 
lines showed specificity for autologous tumor cells in terms of mediating lytic activity and in 
producing IFN- (22, 23). More recent studies have gone on to define a vast array of melanoma-
associated antigens and their corresponding MHC restricting elements that allow for tumor cell 
recognition by specific CD8+ (i.e. HLA-A2-restricted among others; ref. 24) Melanoma cells 
(over)express a broad range of antigens that are capable of being recognized by T cells (25), each 
of which may be considered for integration into clinical vaccine formulations for the treatment of 
patients diagnosed with this disease.  
Cancer vaccines based on tumor-associated antigens (TAA) have been extensively evaluated in 
both translational models and in the clinic (26). Although by most accounts TAA-based vaccines 
have been found to be immunogenic in promoting increased frequencies of Ag-specific T cell 
responses in a large proportion of treated patients, they have only rarely proven curative (27). 
This limitation in efficacy may relate, at least in part, to the heterogeneity of cancer cells found 
within a given tumor lesion, particularly with regard to sub-population immunophenotypes. 
Hence, tumor cells expressing lower levels of MHC molecules, molecules associated with the 
antigen-presentation machinery and/or therapeutically targeted tumor antigens have clear 
survival advantages in the face of specific vaccination (28-30). Indeed, many times patients that 
have exhibited objective clinical responses to immunomodulatory therapies ultimately progress 


 
UPCI 12-048 
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with tumors characterized by defects in their antigen-processing machinery and 
immunophenotype (31, 32). 
A theoretical means by which to promote anti-tumor immunity, while coordinately 
circumventing the immunophenotypic “instability” of cancer cells themselves, involves the 
development of vaccines targeting tumor-associated stromal cells, such as (myo)fibroblasts, 
vascular pericytes, and VEC (33-35). Conditioned by the cancer microenvironment, tumor-
associated blood vessel cells are believed to express phenotypes that distinguish them from 
comparable cell populations found in normal tissues, hypothetically allowing for their biased 
immune targeting by specific T effector cell populations (36-38). Interestingly, the prior 
vaccination of mice with fibroblast or endothelial cell lines has been reported to provide some 
degree of protection against subsequent tumor growth (39, 40). Furthermore, prophylactic 
peptide- and/or recombinant vaccines based on tumor blood vessel-associated antigens (TBVA) 
such as endoglin (CD105), NG2, PDGFR, VEGFR1, or VEGFR2 have been reported to 
provide partial protection against challenge with tumor cell lines that fail to express these 
antigens, presumably based on T cell-mediated anti-neovascular activity in the TME (41-47). In 
a very limited number of studies employing therapeutic models, VEGFR1 and VEGFR2 peptide-
based vaccines were shown capable of modestly slowing the progressive growth of subcutaneous 
(SQ) tumors in mice (44, 47). 
We have recently reported that IL-12 cytokine gene-therapy of established tumors growing in 
HLA-A2 transgenic mice (i.e. these mice exhibit a CD8+ T cell repertoire that mimics that of 
HLA-A2+ humans) results in CD8+ T cell-mediated protective immunity directed at host blood 
vessel cells within the tumor microenvironment (48). CD8+ T cells isolated from effectively-
treated animals recognized a range of HLA-A2-presented peptides derived from TBVA. We 
have more recently shown (49) that vaccination of HHD mice with TBVA-derived peptides 
results in protective immunity against multiple types of solid cancers (including melanoma) 
under conditions that are non-permissive for direct T cell recognition of tumor cells. Indeed, 
when applied in the therapeutic setting, such vaccines can be curative, based on Type-1 CD8+ T 
cell targeting of tumor- (but not normal tissue-) associated blood vessel cells (vascular pericytes 
and/or endothelial cells). Since the efficacious peptides derived from the TBVA DLK1 (delta-
like kinase 1), EphA2, HBB (hemoglobin-), NRP1 (neuropilin 1), RGS5 (regulator of G-
signaling protein 5) and TEM-1 (tumor endothelial marker-1) that bear identical sequences in 
mouse and man, and can be recognized by CD8+ T cells isolated from cancer patients (48), these 
translational data support the use of these specific peptides in vaccine formulations for HLA-A2+ 
patients with solid cancers, including melanoma. 
2.4 
Immune Dysfunction in Patients with Advanced-Stage Solid Cancers 
Type-1 (i.e, Th1 CD4+ T cell and Tc1 CD8+ T cell) responses play critical roles in the rejection 


 
UPCI 12-048 
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of tumors (50, 51). Th1 cells secrete IFN- and IL-2, both of which promote cellular immunity, 
in part by providing helper signals for the development of Tc1 (cytotoxic T) lymphocytes (50). 
Th2-type cells produce IL-4 and IL-5, and typically promote a humoral (i.e. antibody-mediated) 
immune response, while Th3 and regulatory T (Treg) cells produce immunosuppressive cytokines 
(IL-10/TFGβ) that can dampen both Type-1 and Type-2 immune responses (51). The majority of 
studies have indicated that the most prevalent cytokine profile observed in patients with 
melanoma is consistent with a Type-2 functional bias in situ (52, 53). In prior studies we found 
little evidence for local Type-1 immune responses within the tumor; i.) there is a minimal 
expression of mRNA encoding for IL-2 and IFN- associated with TIL, and less than 5% of 
tumor infiltrating lymphocytes (TIL) expressed the activation marker, IL-2R (54). TIL were 
defective in their proliferative and cytotoxic capacities (54, 55), and there was also an 
impairment in Type-1 T cell responses against tumor-associated antigens in peripheral blood T 
cells harvested from RCC patients with active disease (52). An analysis of MAGE-6- and 
EphA2-specific CD4+ T cells revealed the predominance of Type-2 responses (i.e. characterized 
by IL-5 but not IFN- production) after in vitro stimulation with autologous dendritic cells (DC) 
pulsed with MAGE-6 and EphA2 peptide epitopes (52). 
Apoptosis may contribute to the immune dysfunction observed for T cells in cancer patients. In 
many solid cancers, a sizeable proportion of tumor-infiltrating lymphocytes (mainly T cells) 
exist in a pro-apoptotic state, and an even greater number of T cells undergo apoptosis following 
in vitro stimulation (56). Peripheral blood T cells isolated from cancer patients are more sensitive 
to activation induced cell death (AICD) versus comparable T cell populations harvested from 
healthy normal donors (57, 58). Most recently we observed that tumor antigen-specific T cells in 
the peripheral blood of melanoma patients with active disease are more sensitive to apoptosis 
when compared to influenza-specific T cells recovered from these same individuals (58).  
In addition to these intrinsic defects associated with anti-tumor T cells in melanoma patients, 
there are also “extrinsic” mechanisms linked to the functional suppression of otherwise 
protective anti-tumor T cells. These include the increased numbers and function of Treg and 
MDSC observed to be enriched in cancer patients (59-62).  
There is growing evidence that CD4+CD25hi+regulatory T cells (Treg) may play an important role 
in suppressing the development of anti-tumor immunity in cancer patients (63). The frequency of 
Treg is elevated in tumor sites and/or the peripheral blood of patients with advanced tumors (59-
62). Treg cells can impair induction of both antigen-specific and nonspecific T cells in cancer 
patients (63) and predict reduced survival in multiple cancer cell types (64, 65). Relevant to 
therapy, experimental models have shown that removal of Treg cells facilitates the therapeutic 
efficacy of protective immunity. Depletion of Treg cells in mouse models using anti-CD25 
antibody enhances anti-tumor activity (66). Furthermore, reducing peripheral blood Treg cell 


 
UPCI 12-048 
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02-15-2016 
numbers in melanoma patients enhances immunocompetence and responsiveness to active 
immunization (67-69). 
MDSC act to suppress T cell effector function, and represent a heterogeneous population of cells 
derived from the myeloid lineage that accumulate in tumor bearing hosts as a result of tumor 
induced alterations in myelopoiesis (70). MDSC have mostly been studied in mice where they 
are identified by the co-expression of CD11b and Gr-1, accumulate in lymphoid organs and 
tumors of tumor bearing mice, and mediate T cell impairment that is reversed with tumor 
removal or with CD11b+ or Gr-1+ cell depletion (71). MDSC have been described in human 
patients with many cancers including kidney cancer (72, 73), where they are defined as cells that 
express the common myeloid marker CD33, but lack markers of mature myeloid cells such as the 
MHC class II molecule HLA-DR. The accumulation of these cells in the peripheral blood of 
advanced-stage cancer patients has been associated with a decrease in the number and 
effectiveness of peripheral blood DC, as well as a decrease in T cell expression of TCR- 
(signaling) chain and IFN- production (70-73). Studies performed in metastatic patients suggest 
these inhibitory effects are mediated via MDSC-derived reactive oxygen species (ROS) and 
MDSC-induced depletion of key biosynthetic amino acids (i.e. arginine, cystine, cysteine and 
tryptophan; refs. (70-74). 
2.5 
Dendritic Cells 
Pre-clinical studies using Dendritic Cells (DC) to stimulate specific anti-tumor T cell responses 
have been encouraging (75-79). DC are the most potent antigen presenting cells (APCs), capable 
of efficiently internalizing and presenting antigen in the context of co-stimulatory signals and 
cytokines that are essential to the induction of effective long-lasting T cell-mediated immunity. 
Animal models have demonstrated that DC, pulsed ex vivo with defined tumor antigens (peptide 
epitopes) or material derived from tumor cells, are capable of eliciting protective, antigen-
specific immune responses (75, 80). Importantly, clinical trials have confirmed that DC pulsed 
with defined (tumor) peptide epitopes or tumor cell lysates induce anti-tumor T cell responses, 
and that occasionally, such immunity coincides with complete tumor regression in late stage 
patients (81-83). 
2.6 
Type-1 Polarized DC 
Two functions of dendritic cells are believed to be important for the ability of DC to induce 
Type-1 Th1 cells and Tc1; i.e. high co-stimulatory activity and high secreted protein levels of 
anti-cancer cytokines, especially IL-12 (79-83). Many previous clinical trials have relied on the 
use of either fully-matured DC exhibiting high stimulatory function but low IL-12 secretion, or 
immature DC that display low stimulatory/high IL-12 secretion functions. Using a novel culture 
method, it is now possible to generate mature DC that exhibit high levels of co-stimulatory 
activity and IL-12 production (84). Such DC are referred to as Type-1 polarized DC or DC1. In 


 
UPCI 12-048 
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02-15-2016 
vitro observations show that DC1 induce up to 50-fold higher frequencies of tumor antigen-
specific CD8+ T cells after in vitro sensitization (the equivalent of vaccination) when compared 
to conventionally matured DCs that have been previously employed in clinical trials (84).  
In addition to their potent immunostimulatory function and high-level production capacity for 
Type-1 polarizing cytokines, DC1 exhibit a stable phenotype that is resistant to tumor-
associated immunosuppressive factors, including IL-10 and PGE2 (84-87). This suggests the 
possibility of using DC1 as a biologic adjuvant to enhance the clinical efficacy of cancer 
vaccines, despite the immune suppressive environment known to exist in many patients with 
melanoma. 
Recently the original DC1 protocol based on fetal bovine serum-supplemented cultures has 
been modified to allow DC1 generation in serum-free media (84). This has facilitated the 
translation of DC1 into ongoing Phase 1/2 clinical trials being performed at the University of 
Pittsburgh Medical Center (UPMC)/University of Pittsburgh Cancer Institute (UPCI) in the 
setting of melanoma (UPCI 03-118, BB-IND 11,754), colorectal cancer (UPCI 05-063; BB-IND 
13,234), and glioma (UPCI 05-115; BB-IND 12,415). Thus far clinical activity for vaccination 
has been observed in 2 of the initial 4 melanoma patients who completed treatment (1PR 12M+ 
in a patient with Stage IIIb disease; 1 SD 11M+ in a patient with stage IV lung disease). The two 
first patients with resected metastatic CRC are undergoing follow-up for time to recurrence and 
immunologic evaluations. We also recently completed a Phase 1/2 DC1 (1-3 x 107 cells) + 
glioma peptide (derived from the EphA2 [specifically EphA2883-891 to be used in the current 
vaccine formulation], IL-13R2, TKL-40 and gp100 antigens) vaccine trial of 22 HLA-A2+ 
patients with malignant glioma (88). More than half of the patients exhibited vaccine-associated 
increases against at least one of the 4 treatment peptides, with increased levels of serum IFN- 
and CXCL10/IP-10 post-vaccination. Nine patients achieved a PFS exceeding 12 months, with 
one patient developing a complete response to treatment. To date, these regimen have proven to 
be well-tolerated, with no untoward side effects observed in any of these settings.  
2.7 
Improved Anti-Tumor Efficacy for Combined Dasatinib + Vaccine Therapy 
Given our own and others’ recent reports supporting the ability of tyrosine kinase inhibitors 
(such as sunitinib and dasatinib) to mitigate at least a portion of the immunosuppressive circuits 
(i.e. regulatory T cells and myeloid-derived suppressor cells) present in cancer patients, we 
hypothesize that the co-application of dasatinib will increase the immunogenicity of specific 
vaccination protocols. 
We recently performed and published proof-of-concept, pre-clinical studies in which we 
observed superior anti-tumor efficacy for therapies combining TKI (sunitinib) + vaccination 
(DC1/peptide) approaches (89). This was particularly evident when sunitinib was applied at the 
time of the initial vaccination or the vaccine booster. Treatment effectiveness was associated 


 
UPCI 12-048 
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02-15-2016 
with the acute loss of (and/or failure to recruit) cells bearing MDSC or Treg phenotypes within 
the melanoma microenvironment and the corollary, prolonged enhancement of specific Type-1 
CD8+ T cell responses in the melanoma-draining lymph node (TDLN) and the melanoma lesion 
itself. Enhanced Type-1 T cell infiltration of tumors was associated with treatment-induced 
expression of VCAM-1 and CXCR3 ligand chemokines (CXCL9/Mig, CXCL10/IP-10 and 
CXCL11/ITAC) in vascular/peri-vascular cells within the tumor microenvironment (TME), with 
the combined therapy benefits negated by administration of blocking antibodies against CXCR3 
or vascular cell adhesion molecule (VCAM)-1. These data support the ability of sunitinib to 
enhance the capacity of vaccines to elicit Type-1 CD8+ T cell responses in tumor-bearing hosts 
and to concomitantly (re)condition the TME to become more receptive to the recruitment and 
prolonged therapeutic action of such vaccine-induced CD8+ T effector cells. More recently we 
have observed similar tendencies when combining dasatinib and DC1/peptide vaccines in murine 
melanoma models (Fig. 1). 
Fig. 1. Combination dasatinib + DC1/peptide-based vaccination results in superior activation of specific CD8+ T 
cells and anti-tumor efficacy in a murine melanoma model. (A) M05 (B16.OVA) melanoma cells were injected 
s.c. into syngenic C57BL/6 mice and allowed to establish for 10 days, at which time animals were randomized for 
tumor size and cohorts of 7 mice/group were left untreated, or they were treated with orally administered dasatinib 
(DAS; 0.1 mg/day for 7 consecutive days), a contralateral s.c. vaccine (VAC) consisting of 106 OVA peptide-pulsed 
dendritic cells (DC)  or a combination of DAS + VAC. Animals were then monitored for tumor growth (B) and at 
day 34, 2 mice/group were sacrificed for analysis of specific T cell (isolated from the tumor-draining lymph node 
(TDLN) of tumor (TIL)) recognition of the OVA peptide based on IFN- secretion as monitored by ELISA (C). Day 
34 tumors were also isolated and analyzed for their content of myeloid-derived suppressor cells (MDSC; bearing a 
CD11b+Gr1+ phenotype) and regulatory T cells (Treg; bearing a CD4+Foxp3+ phenotype) by flow cytometry (D); 
MO5 (B16.OVA) 
injected s.c.  
(2 x 105)
+/- oral Dasatinib (0.1 mg/day) 
on d10-16
+/- s.c. DC/OVA257-262
vaccines d10, d17
Monitor tumor size
TME marker analysis (d34)
Immune monitoring (d34)
A
B
10
20
30
40
50
0
100
200
300
400
500
Untreated
Dasatinib
Vaccine
DAS + VAC
M05 Tumor Size (mm2)
Days Post Tumor 
Inoculation
D
7.3%
CD11b
Gr1
22.6%
2.5%
2.3%
4.3%
8.3%
2.8%
2.1%
CD4
Foxp3
Untreated
Dasatinib
Vaccine
DAS + VAC
C
0
1000
2000
3000
4000
IFN-(pg/ml)
Untreated
Dasatinib
Vaccine
DAS + VAC
Untreated
Dasatinib
Vaccine
DAS + VAC
TDLN
TIL
E
CD31 x
VCAM-1
x DAPI
CD31 x
CXCL10
x DAPI
Untreated
Dasatinib
Vaccine
DAS + VAC
0.3 + 0.2
25.1 + 6.3*
21.8 + 7.9*
81.8 + 6.0**
1.8 + 1.5
17.8 + 8.4*
26.8 + 5.5*
162 + 18**
**
**
**
*
*
*
*
MO5 (B16.OVA) 
injected s.c.  
(2 x 105)
+/- oral Dasatinib (0.1 mg/day) 
on d10-16
+/- s.c. DC/OVA257-262
vaccines d10, d17
Monitor tumor size
TME marker analysis (d34)
Immune monitoring (d34)
A
B
10
20
30
40
50
0
100
200
300
400
500
Untreated
Dasatinib
Vaccine
DAS + VAC
M05 Tumor Size (mm2)
Days Post Tumor 
Inoculation
B
10
20
30
40
50
0
100
200
300
400
500
Untreated
Dasatinib
Vaccine
DAS + VAC
M05 Tumor Size (mm2)
Days Post Tumor 
Inoculation
D
7.3%
CD11b
Gr1
22.6%
2.5%
2.3%
4.3%
8.3%
2.8%
2.1%
CD4
Foxp3
7.3%
CD11b
Gr1
22.6%
2.5%
2.3%
7.3%
CD11b
Gr1
22.6%
2.5%
7.3%
CD11b
Gr1
22.6%
2.5%
2.3%
4.3%
8.3%
2.8%
2.1%
4.3%
8.3%
2.8%
4.3%
8.3%
2.8%
2.1%
CD4
Foxp3
Untreated
Dasatinib
Vaccine
DAS + VAC
C
0
1000
2000
3000
4000
IFN-(pg/ml)
Untreated
Dasatinib
Vaccine
DAS + VAC
Untreated
Dasatinib
Vaccine
DAS + VAC
TDLN
TIL
E
CD31 x
VCAM-1
x DAPI
CD31 x
CXCL10
x DAPI
Untreated
Dasatinib
Vaccine
DAS + VAC
0.3 + 0.2
25.1 + 6.3*
21.8 + 7.9*
81.8 + 6.0**
1.8 + 1.5
17.8 + 8.4*
26.8 + 5.5*
162 + 18**
**
**
**
*
*
*
*


 
UPCI 12-048 
13 
02-15-2016 
percentage of cells bearing the specified phenotype is reported in panel insets) and for expression of VCAM-1 and 
CXCL10 proximal to CD31+ vascular endothelial cells using immunofluorescence microscopy (E); mean +/- SD 
number of cells over 10 high-power fields are reported in panel insets). Three experiments were performed 
independently, each yielding similar data, *p < 0.05 versus untreated; **p < 0.05 versus all other groups.  
Fig. 2. Dasatinib (but not imatinib) pre-treatment of HLA-A2+ human melanoma cell-line Mel526 increases 
degradation of EphA2 and improves tumor cell recognition by EphA2-specific CD8+ T cells. In addition to its 
inhibitory action on src kinases, dasatinib has been reported to inhibit signaling through the receptor tyrosine kinase 
EphA2 (90) which is commonly over-expressed by human melanomas in situ (91) and shown to represent a critical 
oncogene in melanoma tumorigenic potential (92). We treated the human HLA-A2+, EphA2+ with various doses of 
dasatinib or imatinib for 24h and analyzed EphA2 protein by western blotting. Panel A reports the percentage of 
control EphA2 expression in the TKI-treated melanoma cells. This loss of EphA2 protein is due to the proteasome-
dependent degradation of the protein based on the ability of MG-132 and lactacystin to block protein loss (data not 
shown). Using flow cytometry (B), we also showed that cell surface expression of EphA2, but not HLA-A2 class I 
molecules was reduced selectively after treatment with dasatinib. Finally, since proteasomal processing is required 
for the generation of many MHC class I-presented peptides to CD8+ T cells, we next analyzed whether TKI (0.5 
mg/ml) pre-treatment enhanced tumor cell recognition by anti-EphA2 CD8+ T cells based on their secretion of IFN-
after stimulation with these melanoma cells (C). To validate the HLA-A2-restricted nature of T cell recognition, 
we showed that addition of the anti-HLA-A2 blocking antibody BB7.2 to T cell/melanoma cultures ablated specific 
IFN- production by the CD8+ T cells. *p < 0.05 versus control untreated tumor cells (Control) and dasatinib-treated 
tumor cells + BB7.2 antibody (DAS/BB7.2).  
0
50
100
150
200
250
300
350
T Cells Only
Control
Control/BB7.2
DAS
DAS/BB7.2
SUT
SUT/BB7,2
IMAT
IMAT/BB7.2
IFN-(pg/ml)
+ Mel526 
pre-treated with:
*
A
B
0
20
40
60
80
100
120
DAS
IMAT
% Control EphA2 Protein
0
0.1
0.5
1.0
[TKI] (g/ml)
*
*
Control Ab
EphA2 Ab
EphA2 Ab + IMAT 0.5 M
EphA2 Ab + DAS 0.5 M
EphA2
HLA-A2
Events
C
0
50
100
150
200
250
300
350
T Cells Only
Control
Control/BB7.2
DAS
DAS/BB7.2
SUT
SUT/BB7,2
IMAT
IMAT/BB7.2
IFN-(pg/ml)
+ Mel526 
pre-treated with:
*
A
B
0
20
40
60
80
100
120
DAS
IMAT
% Control EphA2 Protein
0
0.1
0.5
1.0
[TKI] (g/ml)
*
*
0
20
40
60
80
100
120
DAS
IMAT
% Control EphA2 Protein
0
0.1
0.5
1.0
[TKI] (g/ml)
*
*
Control Ab
EphA2 Ab
EphA2 Ab + IMAT 0.5 M
EphA2 Ab + DAS 0.5 M
EphA2
HLA-A2
Events
Control Ab
EphA2 Ab
EphA2 Ab + IMAT 0.5 M
EphA2 Ab + DAS 0.5 M
Control Ab
EphA2 Ab
EphA2 Ab + IMAT 0.5 M
EphA2 Ab + DAS 0.5 M
EphA2
HLA-A2
Events
C


 
UPCI 12-048 
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Fig. 3. Proposed mechanism of action for dastinib + vaccine co-therapy. Dasatinib is proposed to mediate 
beneficial anti-tumor impact based on a series of effects including its acute inhibition of immunoregulatory cells 
(MDSC and Treg) and normalization of blood vasculature within the tumor microenvironment (TME), leading to 
reduced hypoxia and interstitial fluid pressure (A). Stromal cells within the TME alter their expression of 
chemokines and adhesion molecules, such as CXCL10 and VCAM-1 that are required for recruitment and 
infiltration of protective CXCR3+VLA4+ Type-1 CD8+ T cells (B). Dasatinib also mediates direct effects on EphA2+ 
melanoma cells, by increasing the proteolytic processing of this protein into T cell recognized peptide epitopes, 
leading to enhanced anti-tumor activity mediated by anti-EphA2 TIL in vivo (C).  
 
3. 
STUDY RATIONALE 
3.1 
Importance and Rationale for the Trial 
Current therapeutic approaches available for patients with advanced-stage melanoma remain 
inadequate, and existing approaches including those involving immunotherapy with cytokines 
and/or targeted strategies have resulted in disappointingly low rates of durable and complete 
responses. Correcting immune dysfunction in advanced-stage melanoma patients using TKI such 
as dasatinib is proposed to relicense the patient’s immune system to respond optimally to specific 
immunization. The integration of antigens expressed by tumor-associated blood vessel cells 
provides a means to selectively target the genetically-/antigenically-heterogeneous population of 
tumor cells in the advanced-stage melanoma patient. 
The current trial represents a pilot, randomized, Phase 2 study to determine the activity and 
safety of intradermal administration of αDC1s loaded with a mixture of six TBVA-derived 
VCAM-1
CXCL10  
Pericyte
VEC
Vaccine-Induced 
Tc1 (CD8+ T cell)
EphA2
Peptide
MHC I
EphA2
Protein
MEL
Degrade
Regulatory Cells
Vascular Norm.
Hypoxia, HIF1/2
Kill
Cytokines
Tumor
Blood 
Vessels
Tc1 infiltrate
A
B
C
VCAM-1
CXCL10  
Pericyte
VEC
Vaccine-Induced 
Tc1 (CD8+ T cell)
EphA2
Peptide
MHC I
EphA2
Protein
MEL
Degrade
Regulatory Cells
Vascular Norm.
Hypoxia, HIF1/2
Kill
Cytokines
Tumor
Blood 
Vessels
Tc1 infiltrate
A
B
C


 
UPCI 12-048 
15 
02-15-2016 
peptides at the time of, or a cycle prior to, starting study treatment with the TKI dasatinib. 
3.2 
Rationale for Vaccine Dose, Route of Administration and Schedule 
We propose to analyze CD8+ T cell responses against the TBVA DLK1, EphA2, HBB, NRP1, 
RGS5, and TEM1 in peripheral blood of HLA-A2+ melanoma patients prior to, during the course 
of, and one month after the last dose of dasatinib. Based on the strong Type-1 polarizing 
potential of DC1 in vitro, we hypothesize that these vaccines will enhance Type-1 CD8+ T cell 
responses against at least 3 of the 6 peptides included in the vaccine (particularly when patients 
receive concurrent dasatinib administration, as this removes the regulatory action of MDSC/Treg 
suppressor cells). 
We chose to evaluate a mixture of six TBVA-derived epitopes to be applied to DC1s as an 
intradermal vaccine injection into 28 HLA-A2+ patients with advanced-stage melanoma. This 
choice is based on the finding of superior anti-tumor efficacy in HLA-A2 transgenic tumor 
models for the pooled peptide vaccine approach and the relevance of the previously identified 
TBVA-derived peptides (which share sequence identity in both human and mouse TBVA) in the 
HLA-A2+ human patient setting (48, 49). Our combinational vaccine + dasatinib modeling 
suggest that optimal therapeutic benefit against established M05 melanoma occurred when 
sunitinib administration was initiated at the time of initial vaccination or at the time of boosting. 
Systemic review and meta-analysis of previous DC-based vaccine trials in cancer patients 
suggests that: i.) vaccine-induced T cell responses are associated with beneficial clinical 
outcome; ii.) mature DC (such as DC1) were superior activators of specific immunity and a 
better clinical prognosis when compared to immature DC; iii.) while a threshold dose of DC is 
required in the vaccine in order to promote specific immunity, a vast increase in DC number over 
that threshold did not generally yield superior efficacy (93-98). A recent study by Verdijk et al. 
suggests that intradermal delivery of DC-based vaccines in patients with advanced stage 
melanoma was clinically equitable to the delivery of these cells directly into lymph nodes (95), 
while a report from Lesterhuis and colleagues argues that intradermal delivered DC-based 
vaccines were superior to intranodal delivered vaccines in promoting melanoma-specific T cell 
activation in vivo (96). In vivo tracking of intradermal injected DC in melanoma patients 
suggests that approximately 4% of the administered DC actually migrate to tissue-draining 
lymph nodes and that the delivery of approximately 5 x 105 (vaccine) DC are needed to promote 
clinically-meaningful levels of antigen-specific T cells (95). By extrapolation, these figures 
indicate that intradermal injection of a vaccine containing approximately 107 mature antigen-
loaded DC1 would be anticipated to provide a quasi-optimal degree of immune stimulation that 
may be associated with clinical benefit. Pre-clinical, clinical, and mathematical modeling all 
suggest that optimal vaccine-induced immunity and benefit to the tumor-bearing host can be best 
achieved through repeated immunization (3-5 vaccines) provided over a regular-interval 


 
UPCI 12-048 
16 
02-15-2016 
schedule (93-98). Since there is no consensus in the literature for an optimal time interval 
between the individual vaccinations, we have adopted a protocol involving 4 intradermal 
vaccines every 2 weeks, which is a commonly employed schedule for DC-based vaccines (96). 
4. 
STUDY DESIGN 
This is a single-center, prospective randomized, pilot, Phase 2 trial evaluating the activity, safety 
and immune effects of dasatinib given in combination with an autologous type-1 polarized DC 
vaccine. Dasatinib will be administered at the standard dose and schedule recommended by the 
FDA (70 mg BID). The autologous type-I DC vaccine will be administered either prior to, or 
concomitant with, the inititation of dasatinib administration. Patients will be vaccinated 
intradermally with the αDC1/peptide mixture on days 1 and 15 of every cycle on an outpatient 
basis in the University of Pittsburgh Clinical and Translational Research Center (UPCI-CTRC). 
For those patients starting therapy with vaccine alone, dasatinib will be initiated on day 29 after 
receiving the first immunization. Unless patients are removed from study, they will be treated for 
at least 6 cycles or disease progression. In cases where there is continued clinical benefit and no 
additional vaccine product is available, patients can continue to be treated on single agent 
dasatinib. Additional leukapheresis procedures may be performed at the discretion of the 
Investigator/sub-investigator for patients that derive benefit but require the production of 
additional vaccine. 
4.1 
Leukapheresis 
Leukapheresis (3 hours, 180 minutes) is a minimal risk procedure. Prior to the procedure each 
subject’s venous access will be evaluated. If a subject does not have acceptable venous access a 
pheresis catheter will need to be put in place. All selected patients will undergo a single limited 
leukapheresis once they have been deemed eligible and prior to the first course of vaccination. If 
the patients receive benefit from the vaccine, but require additional cells for vaccine production, 
the leukapheresis may be repeated at the discretion of the investigator. The additional 
leukapheresis will take about two hours (120 minutes) to complete. One time of the subject's 
blood volume will be processed per procedure. The product is delivered immediately to the 
UPCI Immunologic Monitoring and Cellular Products Laboratory (IMCPL) from the UPCI-
CTRC. Additional leukapheresis procedures may be performed at the discretion of the 
Investigator/sub-investigator for patients that derive benefit but require production of additional 
vaccine. 
Leukapheresed product will be immediately processed as described in the Chemistry, 
Manufacturing, and Control section of BB-IND 11754, and a part of it will be used for the first 
vaccination course (Week 1). The remainder of the product will be cryopreserved as described. If 
cytopenia (WBC < 2000/mm3 or platelets < 40,000/mm3) develops during, or as a result of, 
leukapheresis, the procedure will be postponed until recovery. This will not be considered an 
adverse event. Samples from each cell product will be obtained for hemoglobin, hematocrit, total 


 
UPCI 12-048 
17 
02-15-2016 
WBC, and differential and platelet count. 
4.2 
Vaccine 
4.2.1 
Formulation 
Dendritic cells (DC) are derived from autologous (the subject’s own) adherent mononuclear cells 
(monocytes) in the peripheral blood obtained from leukapheresis. In this case, “biologic product” 
and “biologic substance” are the same. We prefer the term “biologic product.” 
The vaccine will be manufactured in the UPCI-IMPCL, under cGMP conditions. 
4.2.2 
Storage and preparation 
The final product is placed in vials with labels identifying each unique vaccine lot and 
cryopreserved. DCs used in the vaccine will be suspended in 5% human serum albumin (HSA) 
and delivered to the clinic for administration. For preparation of the vaccines, the labeled vials of 
cryopreserved αDC1 are removed from storage in liquid nitrogen and quickly thawed in a 37C 
water bath. After 3 washes in sterile medium, thawed αDC1 will be suspended in saline with 5% 
human serum albumin (HSA), placed in sterile syringe for administration to the subject and 
delivered to the clinic for administration, depending on time of notification, it can be as early as 
10 am. Each syringe will be labeled with a custom-designed label, identifying the subject and the 
vaccine. Both saline and HSA are clinical grade (NDC 0944-0490-01 and NDC 0338-0049-31) 
and available from UPMC hospital pharmacy as injection products for humans.  
4.2.3 
Administration 
The autologous type-I DC vaccine will be administered intradermally either prior to, or 
concomitant with, the initiation of dasatinib administration. The injections will be performed on 
an outpatient basis in the UPCI-CTRC. 
4.2.4 
Anticipated Adverse Events (Vaccine) 
Dendritic cell-based vaccines have been extensively evaluated in thousands of cancer patients 
over the past 15 years (25, 27, 79, 81, 82, 95, 96) and found to be safe and extremely well-
tolerated. The experience of the UPMC/UPCI in trials involving vaccination with DC1 has 
similarly shown that this cellular therapy is safe (88). Since the toxicities observed for the TKI 
are not of an immunologic nature, we do not anticipate any increase in adverse event frequency 
or severity for the dasatinib + vaccine cohort (Arm B) to exceed the level previously observed 
for dasatinib alone. 
Dendritic cell vaccines carry a potential risk of development of autoimmune disease (such as 
lupus or vitiligo). Whereas development of vitiligo has been reported with similar vaccines 
administered to melanoma subjects, and correlated with positive response to vaccination, other 
side-effects have not been reported. Based on the previous trials with DC-based vaccines, the 
chances of developing an allergic reaction to DC-based vaccines are rare. There is a chance that 


 
UPCI 12-048 
18 
02-15-2016 
subjects could have an allergic reaction to the vaccine. 
4.3 
Dasatinib 
Dasatinib is provided by BMS in 3 different strengths: 
Strength 
Description 
5 mg 
white, round, film coated tablet 
20 mg 
white to off-white, biconvex, round, film coated tablet with either “20” or 
“BMS” debossed on one side and “527” on the other side 
50 mg 
white to off-white, biconvex, oval, film coated tablet with either “50” or 
“BMS” debossed on one side and “528” on the other side 
 
4.3.1 
Packaging and Labeling 
Dasatinib is supplied as 5 mg, 20 mg, and 50 mg film-coated tablets containing dasatinib with 
lactose monohydrate, microcrystalline cellulose, croscarmellose sodium, hydroxypropyl 
cellulose, and magnesium stearate. The tablet coating contains hydroxypropyl methylcellulose, 
titanium dioxide, and polyethylene glycol (triacetin in the 5 mg film-coated tablet). Tablets for 
clinical studies are supplied in high-density polyethylene bottles containing a desiccant and 
cotton. The bottles are heat-induction sealed with child resistant caps.  
Each bottle is labeled in an open label manner. Labels contain, at a minimum, the following 
information: product name, tablet strength, batch number, directions for use, storage conditions, 
and appropriate caution statements. 
4.3.2 
Storage 
Bottles containing dasatinib tablets should be stored at 15° - 25°C. 
The investigational product should be stored in a secure area according to local regulations. The 
investigator is responsible for ensuring that it is dispensed only to study patients and only from 
official study sites by authorized personnel, as dictated by local regulations. 
The investigator is responsible for ensuring that the investigational product is stored under the 
appropriate environmental conditions (temperature, light, and humidity). 
If concerns regarding the quality or appearance of the investigational product arise, do not 
dispense the investigational product, and contact BMS immediately. 
4.3.3 
Handling and Dispensing 
Procedures for proper handling and disposal of anticancer drugs should be considered. 
Dasatinib tablets consist of a core tablet (containing the active drug substance), surrounded by a 
film coating to prevent exposure of pharmacy and clinical personnel to the active drug substance. 


 
UPCI 12-048 
19 
02-15-2016 
If tablets are crushed or broken, pharmacy and clinical personnel should wear disposable 
chemotherapy gloves. Personnel who are pregnant should avoid exposure to crushed and/or 
broken tablets. 
The Investigator (or assigned designee, i.e., study pharmacist) will dispense the proper number 
of each strength tablet to the subject to satisfy dosing requirements for the study. The containers 
provided to the subject should be labeled with proper instructions for use. The lot numbers, 
dosing start dates and the number of tablets for each dosage strength must be recorded on the 
drug accountability pages of record for the site. The subject must be instructed to return all 
unused dasatinib in the provided packaging at each subsequent visit. 
4.3.4 
Drug Ordering and Accountability 
Initial Orders of dasatinib should be requested by completing the Dasatinib (Sprycel®) Drug 
Supply Form for Investigator Sponsored Studies and submitting the request form electronically 
via e-mail at least 5-7 business days before the expected delivery date. Deliveries will be made 
Tuesday through Friday. 
Initial drug supply will be provided for a 12-week treatment period per subject. 
Please send drug supply form to: srcsupply@bms.com. 
Re-supply requests should be sent to the appropriate email address (see Section 4.3.4). Please 
check “Re-supply” on the drug supply form. Re-supply requests should be submitted at least 5-7 
business days before the expected delivery date. Deliveries will be made Tuesday through 
Friday. 
4.3.5 
Safety of Dasatinib in Clinical Studies in CML and Ph+ ALL 
The data described below reflect exposure to dasatinib in 911 patients with leukemia from 1 
Phase 1 and 5 Phase 2 clinical studies. The median duration of therapy was 6 months (range 0–
19 months).  
The majority of dasatinib-treated patients experienced adverse drug reactions at some time. Drug 
was discontinued for adverse drug reactions in 6% of patients in chronic phase CML, 5% in 
accelerated phase CML, 11% in myeloid blast phase CML, and 6% in lymphoid blast phase 
CML or Ph+ ALL.  
The most frequently reported serious adverse events (SAEs) included pyrexia (9%), pleural 
effusion (8%), febrile neutropenia (7%), gastrointestinal bleeding (6%), pneumonia (6%), 
thrombocytopenia (5%), dyspnea (4%), anemia (3%), cardiac failure (3%), and diarrhea (2%).  
All treatment-emergent adverse events (excluding laboratory abnormalities), regardless of 
relationship to study drug, that were reported in at least 20% of the patients in dasatinib clinical 
studies are shown in Table 2. 


 
UPCI 12-048 
20 
02-15-2016 
 
Table 2. Adverse Events Reported ≥20% in Clinical Studies in CML and Ph+ ALL 
 
Table 3. CTC Grades 3/4 Laboratory Abnormalities in Clinical Studies in CML and Ph+ ALL 
 
Chronic Phase 
(n=488) 
Accelerated Phase 
(n=186) 
Myeloid 
Blast Phase 
(n=132) 
Lymphoid Blast 
Phase and 
Ph+ ALL 
(n=105) 
 
Percent (%) of Patients 
Hematology Parameters 
 
 
 
 
 
Neutropenia 
49 
74 
83 
81 
 
Thrombocytopenia 
48 
83 
82 
83 
 
Anemia 
18 
70 
70 
51 
Biochemistry Parameters 
 
 
 
 
 
Hypophosphatemia 
11 
13 
23 
21 
 
Hypocalcemia 
2 
9 
20 
15 
Preferred Term 
All Patients 
(n=911) 
Chronic 
Phase 
(n=488) 
Accelerated 
Phase 
(n=186) 
Myeloid 
Blast 
Phase 
(n=132) 
Lymphoid 
Blast Phase 
and 
Ph+ ALL 
(n=105) 
All 
Grades 
Grades 3/4 
Grades 3/4 
Percent (%) of Patients 
Fluid Retention 
50 
9 
6 
6 
23 
9 
 
Superficial Edema 
36 
1 
0 
2 
3 
2 
 
Pleural Effusion 
22 
5 
3 
3 
14 
8 
Diarrhea 
49 
5 
3 
10 
8 
6 
Headache 
40 
2 
2 
2 
4 
6 
Hemorrhage 
40 
10 
3 
18 
23 
17 
Musculoskeletal Pain 
39 
4 
2 
3 
6 
13 
Pyrexia 
39 
5 
1 
5 
13 
9 
Fatigue 
39 
3 
2 
4 
4 
8 
Skin Rash 
35 
1 
1 
1 
1 
4 
Nausea 
34 
1 
<1 
0 
5 
2 
Dyspnea 
32 
6 
5 
7 
11 
9 
Cough 
28 
<1 
<1 
1 
1 
0 
Infection 
(including bacterial, 
viral, fungal, non-
specified) 
34 
7 
4 
8 
15 
13 
Infection/Inflammation 
26 
1 
1 
1 
5 
1 
Abdominal Pain 
25 
2 
1 
2 
4 
6 
Pain 
26 
2 
<1 
1 
5 
4 
Vomiting 
22 
1 
1 
2 
2 
2 
Febrile Neutropenia 
9 
8 
2 
11 
17 
20 


 
UPCI 12-048 
21 
02-15-2016 
 
Chronic Phase 
(n=488) 
Accelerated Phase 
(n=186) 
Myeloid 
Blast Phase 
(n=132) 
Lymphoid Blast 
Phase and 
Ph+ ALL 
(n=105) 
 
Elevated SGPT (ALT) 
1 
4 
7 
11 
 
Elevated SGOT (AST) 
1 
2 
5 
8 
 
Elevated Bilirubin 
<1 
1 
5 
8 
 
Elevated Creatinine 
0 
2 
1 
1 
CTC grades: neutropenia (Grade 3 ≥0.5–1.0 × 109/L, Grade 4 <0.5 × 109/L); thrombocytopenia (Grade 3 ≥10–50 × 
109/L, Grade 4 <10 × 109/L); anemia (hemoglobin ≥65–80 g/L, Grade 4 <65 g/L); elevated creatinine (Grade 3 >3–6 
× upper limit normal range (ULN), Grade 4 >6 × ULN); elevated bilirubin (Grade 3 >3–10 × ULN, Grade 4 >10 × 
ULN); elevated SGOT or SGPT (Grade 3 >5–20 × ULN, Grade 4 >20 × ULN); hypocalcemia (Grade 3 <7.0–6.0 
mg/dL, Grade 4 <6.0 mg/dL); hypophosphatemia (Grade 3 <2.0–1.0 mg/dL, Grade 4 <1.0 mg/dL).  
 
4.3.6 
Laboratory Abnormalities 
Myelosuppression was commonly reported in all patient populations. The frequency of Grade 3 
or 4 neutropenia, thrombocytopenia, and anemia was higher in patients with advanced CML or 
Ph+ ALL than in chronic phase CML. Myelosuppression was reported in patients with normal 
baseline laboratory values as well as in patients with pre-existing laboratory abnormalities. 
(Table 3) 
In patients who experienced severe myelosuppression, recovery generally occurred following 
dose interruption and/or reduction; permanent discontinuation of treatment occurred in 1% of 
patients.  
Grade 3 or 4 elevations of transaminases or bilirubin and Grade 3 or 4 hypocalcemia and 
hypophosphatemia were reported in patients with all phases of CML but were reported with an 
increased frequency in patients with myeloid or lymphoid blast CML and Ph+ ALL. Elevations 
in transaminases or bilirubin were usually managed with dose reduction or interruption. Patients 
developing Grade 3 or 4 hypocalcemia during the course of dasatinib therapy often had recovery 
with oral calcium supplementation (Table 3). 
4.3.7 
Anticipated Adverse Events (Dasatinib) 
4.3.7.1 Myelosuppression 
Treatment with dasatinib is associated with severe (NCI CTC Grade 3 or 4) thrombocytopenia, 
neutropenia, and anemia. Their occurrence is more frequent in patients with advanced CML or 
Ph+ ALL than in chronic phase CML. Complete blood counts should be performed weekly for 
the first month and then monthly thereafter, or as clinically indicated. Myelosuppression was 
generally reversible and usually managed by withholding dasatinib temporarily or dose reduction 
(14).  
4.3.7.2 Bleeding Related Events 
In addition to causing thrombocytopenia in human patients, dasatinib caused platelet dysfunction 


 
UPCI 12-048 
22 
02-15-2016 
in vitro. Severe CNS hemorrhages, including fatalities, occurred in 1% of patients receiving 
dasatinib. Severe gastrointestinal hemorrhage occurred in 7% of patients and generally required 
treatment interruptions and transfusions. Other cases of severe hemorrhage occurred in 4% of 
patients. Most bleeding events were associated with severe thrombocytopenia.  
Patients were excluded from participation in dasatinib clinical studies if they took medications 
that inhibit platelet function or anticoagulants. Caution should be exercised if patients are 
required to take medications that inhibit platelet function or anticoagulants. 
4.3.7.3 Fluid Retention 
Dasatinib is associated with fluid retention, which was severe in 9% of patients, including pleural 
and pericardial effusion reported in 5% and 1% of patients, respectively. Severe ascites and 
generalized edema were each reported in 1%. Severe pulmonary edema was reported in 1% of 
patients. Patients who develop symptoms suggestive of pleural effusion such as dyspnea or dry 
cough should be evaluated by chest X-ray. Severe pleural effusion may require thoracentesis and 
oxygen therapy. Fluid retention events were typically managed by supportive care measures that 
include diuretics or short courses of steroids (14).  
4.3.7.4 QT Prolongation 
In vitro data suggest that dasatinib has the potential to prolong cardiac ventricular repolarization 
(QT interval). In single-arm clinical studies in patients with leukemia treated with dasatinib, the 
mean QTc interval changes from baseline using Fridericia’s method (QTcF) were 3–6 msec; the 
upper 95% confidence intervals for all mean changes from baseline were <8 msec. Nine patients 
had QTc prolongation reported as an adverse event. Three patients (<1%) experienced a QTcF 
>500 msec.  
Dasatinib should be administered with caution to patients who have or may develop prolongation 
of QTc. These include patients with hypokalemia or hypomagnesemia, patients with congenital 
long QT syndrome, patients taking anti-arrhythmic medicines or other medicinal products that 
lead to QT prolongation, and cumulative high-dose anthracycline therapy. Hypokalemia or 
hypomagnesemia should be corrected prior to dasatinib administration (14). 
5. 
STUDY TREATMENT PLAN 
No investigational or commercial agents or therapies other than those described below may be 
administered with the intent to treat the patient's malignancy. 
5.1 
Dasatinib Administration 
All patients will receive dasatinib at a starting dose of 70 mg twice daily by mouth in the 
outpatient setting. Dasatinib will be supplied as 50 mg and 20 mg tablets. Patients will take 1 of 
the 50 mg tablets and 1 of the 20 mg tablets twice daily, approximately every 12 hours, at the 
same time each day. Dasatinib may be taken with or without food. Patients are to swallow the 


 
UPCI 12-048 
23 
02-15-2016 
tablets whole. The tablets should not be cut, crushed or dissolved. Study medication will be self-
administered and patients will be asked to keep a daily diary of the time and dose of drug 
ingested. 
Patients on Arm A will start dasatinib administration on cycle 2, day 1 (week 5), while those 
patients in Arm B will start dasatinib administration on cycle 1, day 1 (week 1). Study treatment 
will continue for at least 6 cycles or disease progression. In case of vaccine depletion patients 
may continue on dasatinib alone and there is evidence of clinical benefit. 
The dosing time may be adjusted as required for subject convenience. If doses are missed for 
toxicity, they should not be replaced. If a dose is not taken due to an error, it may be taken up to 
12 hours later. If vomiting occurs within 30 minutes of intake, that dose may be repeated. 
Patients will be provided with a diary in which to record any changes in dosing of study drug. 
Dasatinib should not be taken with grapefruit containing food or beverages. 
REGIMEN DESCRIPTION 
Agent 
Premedications/
Precautions 
Dose 
Route 
Schedule 
Cycle 
Length 
αDC1 Vaccine None 
107 cells 
Intradermal 
injection 
Arms A and B: every 2 weeks 
starting on Cycle 1, day1 
28 days 
(4 weeks) 
Dasatinib 
Take with or 
without food 
70 mg 
Orally, 
twice a day 
Arm A: Daily starting on 
Cycle 2, day 1 
Arm B: daily starting on Cycle 
1, day 1 
 
5.2 
Vaccine administration 
The DC vaccine will be administered by a single intradermal injection of approximately 107 cells 
(a minimum of 5 x 106 cells is allowable due to manufacturing limitations), with all the DCs 
being administered on days 1and 15 of each cycle. The intradermal administration will be in the 
vicinity of the four nodal drainage groups of the four extremities and performed on an outpatient 
basis in the UPCI-CTRC. Study treatment will continue for at least 6 cycles or disease 
progression. 
5.3 
Duration of Study Treatment 
In the absence of treatment delays due to adverse events, treatment may continue for at least 6 
cycles until one of the following criteria applies: 
 
Disease progression, 
 
Intercurrent illness that prevents further administration of treatment, 
 
Unacceptable adverse event(s), 


 
UPCI 12-048 
24 
02-15-2016 
 
Patient decides to withdraw from the study, or 
 
General or specific changes in the patient's condition render the patient unacceptable for 
further treatment in the judgment of the investigator/sub-investigator. 
 
Study is terminated 
 
Loss of ability to freely provide consent 
Patients may continue on dasatinib alone in case the vaccine product is no longer available and 
there is evidence of clinical benefit. 
5.4 
Duration of Follow Up 
Patients will be followed for 1 year (+2 months) per standard of care visits or through medical 
records review after removal from study treatment or until death, whichever occurs first. Patients 
removed from study for unacceptable adverse event(s) will be followed until resolution or 
stabilization of the adverse event. 
6. 
STUDY CALENDAR 
Schedules shown in the Study Calendar below are provided as an example and should be 
modified as appropriate. 
 
Screening evaluations are to be conducted within 1 week of leukapheresis. Scans and x-rays 
must be done < 4 weeks prior to baseline. In the event that the patient’s condition is 
deteriorating, laboratory evaluations should be repeated within 48 hours prior to initiation of the 
next cycle of therapy. 
Week 
Up to -1d 
from 
baseline 
Baseline 
1 
2 
3 
4 
5e 
6e 
7e 
8e 
Off Study 
Treatmentf 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
 
HLA-A2 Screeninga 
X 
 
 
 
 
 
 
 
 
 
 
BRAF, c-KIT mutationa 
X 
 
 
 
 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
 
 
 
 
Physical exam 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
Vital signs 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
Height 
X 
 
 
 
 
 
 
 
 
 
 
Weight 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
Performance status 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
CBC w/diff, platelets 
X 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Serum chemistryb 
X 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
B-HCGc 
X 
 
 
 
 
 
 
 
 
 
 
EKG (as indicated) 
X 
 
 
 
 
 
 
 
 
 
 
Leukapheresis 
 
Xi 
 
 
 
 
 
 
 
 
 


 
UPCI 12-048 
25 
02-15-2016 
Week 
Up to -1d 
from 
baseline 
Baseline 
1 
2 
3 
4 
5e 
6e 
7e 
8e 
Off Study 
Treatmentf 
Vaccine production 
 
X 
 
 
 
 
 
 
 
 
 
Dasatinib 
Arm A 
Arm B  
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
X 
X 
 
X 
X 
 
X 
X 
 
X 
X 
 
DC Vaccinee 
 
 
X 
 
X 
 
X 
 
X 
 
 
Immune monitoring-PBMC 
Xg 
 
X 
 
X 
 
X 
 
X 
Tumor Biopsy 
Xg 
 
 
 
X 
 
 
 
 
AE evaluation 
 
 
X 
X 
Tumor measurements 
X 
 
Tumor measurements are repeated every 8 weeks. 
Documentation (radiologic) must be provided for 
patients removed from study for progressive disease. 
X 
Radiologic evaluation 
X 
 
Radiologic measurements should be performed every 
8 weeks. 
Xh 
a: Not necessary if already known. 
b: Albumin, alkaline phosphatase, total bilirubin, bicarbonate, BUN, calcium, chloride, creatinine, glucose, LDH, 
phosphorus, potassium, total protein, SGOT [AST], SGPT [ALT], sodium, magnesium. 
c: Serum pregnancy test (women of childbearing potential). 
d: Screening is to be performed in the UPCI-CTRC. 
e: Treatment with DC vaccine and dasatinib will continue for at least 6 cycles (or until disease progression), the weeks of 
those cycles will follow the tests and procedures listed for weeks 5 through 8 above for each subsequent cycle. 
f: Four weeks after the last dasatinib administration. 
g: Can happen at any time prior to start of study treatment. 
h: If removed from the study for reasons other than DP. 
i: Additional leukapheresis procedures may be performed at the discretion of the Investigator/sub-investigator for patients 
that derive benefit but require production of additional vaccine. 
 
There is a window of ±7 calendar days available for scheduling/rescheduling treatment and/or procedures at the discretion of 
the Sub-investigator, and as discussed with the Investigator if a course is missed or a subject's treatment and/or testing day(s) 
need to be rescheduled due to the subject’s inability to comply with the study calendar (this includes but is not limited to: 
hospitalizations, business, vacation plans, travel from long distances for study treatment, in advance of the scheduled date to 
allow ready access to the result(s), reduce financial burden on the subject [i.e. non-UPMC insurance coverage] or reduce 
travel inconvenience, illness, transportation issues, holidays, family emergencies, etc.). 
 
7. 
DOSE-LIMITING TOXICITIES 
7.1 
Definition of Dose-Limiting Toxicity 
Toxicities will be scored according to the NCI Common Terminology Criteria for Adverse 
Events (NCI CTCAE) v4.0. 
Dose-limiting toxicity (DLT) is defined as the following regimen-related events experienced 
during Cycle 1 and 2 for Arm A and during Cycle 1 for Arm B: 
 
Grade 4 neutropenia or thrombocytopenia which lasts more than 7 days; 
 
Grade 3 or 4 febrile neutropenia; or 
 
Grade 3 or greater non-hematological toxicities; this includes grade 3 or greater diarrhea, 
nausea or vomiting which last more than 7 days despite adequate treatment (with 
loperamide for diarrhea, 5HT3 antagonists, steroids and dopamine antagonist for N/V). 


 
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7.2 
Dasatinib Dosing delays/modifications 
Dose Level 
Dasatinib Dose 
0 
70 mg twice a day 
-1 
50 mg twice a day 
-2 
100 mg once a day 
 
This study will use the CTCAE (Common Terminology Criteria for Adverse Events) version 4.0 
for toxicity and serious adverse event reporting. A copy can be downloaded from the CTEP 
homepage http://ctep.cancer.gov. 
7.2.1 
Grades 1 or 2 hematologic or non-hematologic toxicities 
No routine dose interruptions or reductions will occur for Grades 1 or 2 toxicities. If the subject 
experiences an intolerable Grade 2 toxicity, dasatinib may be reduced 1 dose level without 
temporarily withholding study drug. If the toxicity resolves to baseline, dasatinib may be 
escalated to the previous dose level. If the intolerable toxicity recurs after re-escalation, dasatinib 
should be reduced 1 dose level and re-escalation of dose should not occur. 
7.2.2 
Grades 3 or 4 hematologic or non-hematologic toxicities 
If the subject experiences a clinically significant Grade 3 non-hematologic toxicity or any Grade 
4 non-hematologic toxicity or Grade 3 or 4 neutropenia or thrombocytopenia, dasatinib must be 
withheld until the toxicity resolves to Grade 1 or less or returns to baseline at the start of therapy, 
and the dose must be reduced 1 dose level when drug is restarted. If the clinically significant 
Grade 3, or Grade 4 non-hematologic toxicity, or Grade 3 or 4 neutropenia or thrombocytopenia 
recurs on the lower dose of dasatinib, drug must be withheld until the toxicity resolves to Grade 
1 or less or returns to baseline, and the dose must be reduced to level -2 when restarted. If the 
clinically significant Grade 3, or Grade 4 non-hematologic toxicity, or Grade 3 or 4 neutropenia 
or thrombocytopenia recurs on dose level -2 of dasatinib, the subject’s treatment with dasatinib 
will be discontinued but follow-up for tumor progression and survival will continue. 
7.3 
Vaccine Dosing delays/modifications 
Although past experience at UPCI and other clinical centers performing DC-based vaccinations 
suggests that immunization with DC1-based vaccines will be well-tolerated and have little if any 
serious adverse side effects, to ensure patient safety we will closely monitor for any potential 
autoimmunity. No dose modifications will be allowed. 
8. 
PATIENT SELECTION 
8.1 
Inclusion Criteria 
 
Patients must be HLA-A2+ and have histologically confirmed melanoma that is 
metastatic (Stage IV) or unresectable Stage IIIB/C and for which standard curative or 
palliative measures do not exist or are no longer effective. 


 
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 
Patients must have measurable disease by RECIST 1.1, defined as at least one lesion that 
can be accurately measured in at least one dimension (longest diameter to be recorded for 
non-nodal lesions and short axis for nodal lesions) as > 20 mm with conventional 
techniques or as > 10 mm with spiral CT scan, MRI, or calipers by clinical exam. See 
Section 11 for the evaluation of measurable disease. 
 
Patients should have at least 2 subcutaneous, intracutaneous, and accessible tumor 
deposits, lymph node or other site available for biopsy purposes. Patients that have one 
biopsiable site that can be amenable to 2 biopsies (pre- and post-) will be considered 
eligible. 
 
Prior chemotherapy, immunotherapy, or targeted therapy is allowed as long as it did not 
include dasatinib. 
 
Age > 18 years. Because no dosing or adverse event data are currently available on the 
use of dasatinib in patients < 18 years of age, children are excluded from this study, but 
will be eligible for future pediatric trials. 
 
ECOG performance status < 2 (Karnofsky > 60%, see Appendix A). 
 
Life expectancy of greater than 12 weeks. 
 
Patients must have normal organ and marrow function as defined below: 
o 
Leukocytes ≥ 3,000/µL 
o 
absolute neutrophil count ≥ 1,500/µL 
o 
absolute lymphocyte count ≥ 500/µL 
o 
platelets ≥ 100,000/µL 
o 
total bilirubin within normal institutional limits 
o 
AST(SGOT)/ALT(SGPT) ≤ 2.5 X institutional upper limit of normal 
o 
Creatinine ≤ 2.0 X institutional upper limit of normal 
 
Serum magnesium, potassium and adjusted (or ionized) calcium ≥ the institutional lower 
limit of normal. (Supplementation of electrolytes prior to screening is allowed). 
 
Sexually active women and men of childbearing potential must agree to use an effective 
method of birth control during the course of the study and for up to 3 months following 
the last dose of the study drug, in a manner such that risk of pregnancy is minimized. 
Surgical sterilization, intrauterine device, birth control pills, or barrier method (e.g. 
condom and/or diaphragm with spermicidal agents) are acceptable forms of birth control. 
Women of childbearing potential must have a negative pregnancy test (serum) within 7 
days prior to treatment. A pregnancy test is not required for registration. Women who 
have not menstruated for more than 2 years will be considered postmenopausal, thus not 
of childbearing potential. 
 
Ability to understand and the willingness to sign a written informed consent document. 
8.2 
Exclusion Criteria 
 
Patients who have had chemotherapy or radiotherapy within 4 weeks (6 weeks for 
nitrosoureas or mitomycin C) prior to entering the study or those who have not recovered 
from adverse events due to agents administered more than 4 weeks earlier. 
 
Patients with documented c-KIT mutations. 
 
Patients who are receiving any other investigational agents. 
 
Patients with known active brain metastases should be excluded. Patients with treated 
brain metastases with documented stability for 4 weeks are eligible. 


 
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 
History of allergic reactions attributed to compounds of similar chemical or biologic 
composition to dasatinib or any of the components of the vaccine being administered as 
part of this study. 
 
Women who are pregnant or nursing/breastfeeding. 
 
History of significant bleeding disorder unrelated to cancer, including: 
o 
Diagnosed congenital bleeding disorders (e.g., von Willebrand’s disease) 
o 
Diagnosed acquired bleeding disorder within one year (e.g., acquired anti-factor 
VIII antibodies) 
 
Patients currently taking medications that inhibit platelet function (i.e., aspirin, 
dipyridamole, epoprostenol, eptifibatide, clopidogrel, cilostazol, abciximab, ticlopidine, 
and any non-steroidal anti-inflammatory drug) because of a potential increased risk of 
bleeding from dasatinib. 
 
Patients currently taking anticoagulants (warfarin, heparin/low molecular weight heparin 
[e.g., danaparoid, dalteparin, tinzaparin, enoxaparin]) because of a potential increased 
risk of bleeding from dasatinib. 
 
Diagnosis of unstable angina or myocardial infarction within 6 months of study entry. 
 
Patients currently taking one or more of the following drugs that are generally accepted to 
have a risk of causing Torsades de Pointes: 
o 
quinidine, procainamide, disopyramide 
o 
amiodarone, sotalol, ibutilide, dofetilide 
o 
erythromycins, clarithromycin 
o 
chlorpromazine, haloperidol, mesoridazine, thioridazine, pimozide 
o 
cisapride, bepridil, droperidol, methadone, arsenic, chloroquine, domperidone, 
halofantrine, levomethadyl, pentamidine, sparfloxacin, lidoflazine. 
 
Diagnosed or suspected congenital long QT syndrome. 
 
Prolonged QTc interval on pre-entry electrocardiogram (> 450 msec) within 30 days prior 
to study registration. 
 
Any history of clinically significant ventricular arrhythmias (such as ventricular 
tachycardia, ventricular fibrillation, or Torsades de pointes) 
 
Uncontrolled intercurrent illness including, but not limited to, ongoing or active infection, 
symptomatic congestive heart failure, unstable angina pectoris, cardiac arrhythmia, or 
psychiatric illness/social situations that would limit compliance with study requirements. 
 
HIV-positive patients on combination antiretroviral therapy are ineligible because of the 
potential for pharmacokinetic interactions with dasatinib. In addition, these patients are at 
increased risk of lethal infections when treated with marrow-suppressive therapy. 
Appropriate studies will be undertaken in patients receiving combination antiretroviral 
therapy when indicated. 
 
8.3 
Inclusion of Women and Minorities 
Both men and women of all races and ethnic groups are eligible for this trial. 
8.4 
General Concomitant Medication and Supportive Care Guidelines 
In general, concomitant medications and therapies deemed necessary for the supportive care and 
safety of the patient are allowed. The administration of growth factors (e.g. procrit, neupogen) is 


 
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allowed. ASCO guidelines for providing growth factor support are recommended. The 
administration of any other anticancer agents including chemotherapy and biologic agents is not 
permitted. The administration of interferons is not allowed. The use of other concurrent 
investigational drugs is not allowed. 
8.4.1 
Bisphosphonates 
Intravenous bisphophonates will be withheld for the first 8 weeks of treatment due to the risk of 
hypocalcemia. After the need for Ca2+ supplementation has been assessed and levels documented 
to be > lower limit of normal, patients on prior bisphosphonate may be restarted with caution at 
the investigator’s discretion. 
8.4.2 
Antacids 
Short-acting antacid agents may be taken, but it is recommended that these not be taken from 2 
hours before to 2 hours after dosing of dasatinib. In vitro solubility data indicate that dasatinib 
may have decreased solubility and absorption at pH > 4. Until further data are available, patients 
should try to avoid taking proton pump inhibitors and H2 antagonists. 
8.4.3 
Medications that prolong the QT interval 
Ideally, patients should not take medications known to prolong the QT interval. However, should 
the Investigator believe that beginning therapy with a potentially QT prolonging medication 
(other than the ones explicitly prohibited) is vital to an individual subject’s care, the Investigator 
must check that the subject’s prior on-therapy ECG has not shown a QTcF  480 msec or an 
increase in QTc  60 msec over the baseline value. Additional ECG(s) will be done one week 
later or more at the Investigator’s discretion to ensure the subject’s safety. 
8.4.4 
Medications that affect CYP3A4 
Caution is warranted when administering dasatinib to patients taking drugs that are highly 
dependent on CYP3A4 for metabolism and have a narrow therapeutic index. Systemic exposures 
to these medications could be increased while receiving dasatinib. In in vitro studies, dasatinib is 
a strong inhibitor of the human CYP3A4 enzyme and a weak inhibitor of CYP1A2, CYP2D6 and 
CYP2C19. Dasatinib shows time-dependent inhibition of CYP3A4; however, there appears to be 
a low probability for drug-drug interactions due to metabolism-dependent CYP3A4 inactivation. 
Results from an in vitro hPXR trans-activation study suggest that dasatinib has little potential to 
induce CYP3A4 through the activation of hPXR. 
Until information regarding exposure-toxicity and exposure-response relationships is available 
with dasatinib, concomitant CYP3A4 inhibitors and inducers should be avoided since they could 
alter the systemic exposure to dasatinib. Incubations with recombinant human CYP450 isozymes 
suggest that dasatinib is primarily metabolized by the CYP3A4 enzyme. Many other enzymes 
appear capable of metabolizing dasatinib, including CYP1A1, 2C9, 2E1, FMO3, 1B1, 2B6, 2A6, 
2C8, and 4A1; however, it is unknown at this time what contributions these enzymes may have 


 
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to the total metabolic clearance of dasatinib. 
Based on pre-clinical data, dasatinib may increase the likelihood of bleeding. Hence, patients 
undergoing surgical procedures, including dental procedures, should be instructed to inform their 
doctors of this potential increased risk. Patients requiring surgery should stop dasatinib at least 1 
day prior to surgery, when feasible. As dasatinib has the potential to cause hemorrhage and its 
effect on wound healing is unclear, a minimum 2 week wait time after surgery is recommended 
and further do not resume medication until recommended by the attending surgeon based on the 
post-operative course. 
Palliative radiation is allowed provided that the irradiated lesion is not the sole target lesion for 
response assessment. 
The following medications are prohibited: 
 
St. John's wort (Hypericum perforatum): May decrease dasatinib plasma concentrations 
unpredictably. Patients receiving dasatinib should not take St. John's wort. 
 
H2 Blockers/Proton Pump Inhibitors:  Long-term suppression of gastric acid secretion by 
H2 blockers or proton pump inhibitors (eg, famotidine and omeprazole) is likely to 
reduce dasatinib exposure. The concomitant use of H2 blockers or proton pump inhibitors 
with dasatinib is not recommended. The use of antacids should be considered in place of 
H2 blockers or proton pump inhibitors in patients receiving dasatinib therapy (see Section 
5.2.2, antacids).  
9. 
RESEARCH SAMPLES 
Patient peripheral blood and tumor biopsies will be obtained at various time points prior to, and 
after, the initiation of therapy. 
If the patient is determined to express the HLA-A2 antigen on their peripheral blood cells, to 
express a wild-type phenotype, and they pass all additional inclusion/exclusion criteria, after 
written consent, they may be entered on trial. 
Those patients determined to express c-KIT mutations will be excluded from study, while BRAF 
mutational status will be used to stratify patients during randomization to ensure a balanced 
proportion of patients with the mutation on both arms. 
9.1 
Biopsy tissue 
Melanoma biopsies will be obtained prior to the first vaccination (baseline) and week 5 (date of 
the third vaccination). Patients should have at least 2 subcutaneous, intracutaneous, and 
accessible tumor deposits, lymph node or other site available for biopsy purposes or one 
biopsiable site that can be amenable to 2 biopsies (pre- and post-) will be considered eligible. 
 
 


 
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9.2 
Blood samples 
At least 3 weeks prior to study treatment, peripheral blood will be obtained for the screening of 
patient HLA-A2 expression status (if not already known) and for baseline testing. Peripheral 
blood will be obtained every 2 weeks on trial beginning Cycle 1 week 3. 
10. CORRELATIVE STUDIES 
We expect to observe that HLA-A2/TBVA peptide dextramer+ CD8+ T cells (i.e. CD8+ T cells 
imaged by flow cytometry using a fluorescently-labeled, antigen-specific probe) will exhibit 
higher frequencies in the peripheral blood and a greater propensity to produce IFN- after the 
initiation of DC1-based vaccines. Since dasatinib is expected to alter the recruiting capacity of 
the tumor microenvironment based on activation of VCAM-1 expression on the tumor-associated 
vascular endothelial cells and locoregional production of CXCR3 ligand chemokines, it is very 
possible (and likely clinically-preferred) that the frequency of TBVA-specific CD8+ T cells will 
selectively decline in the patients peripheral blood if the combined therapy is performing as 
expected based on our preclinical modeling data (given recruitment away out of the blood and 
into the tumor lesion). Based on a recent publication by Ernstoff and colleagues (99), circulating 
levels of the CXCR3 ligand CXCL10 (aka IP-10), as well as, the Okada and colleagues Phase 
1/2 trial results for DC1/peptide vaccination of glioma patients (88) may become elevated 
under treatment conditions in patients that are more prone to exhibit objective clinical response 
to effective immunotherapy. As a consequence, we will analyze levels of serum CXCL10 before, 
during and after combined vaccine + dasatinib therapy to determine correlation with TBVA-
specific CD8+ T cells in the blood versus tumor over time post-treatment. 
10.1 
Immune Monitoring Analysis of TBVA-specific CD8+ T cell responses (Primary 
Endpoint). 
Rationale and Hypothesis: Translation and clinical vaccine trials have demonstrated that 
DC/peptide-based vaccines effectively activate specific CD8+ T cells in tumor-bearing hosts that 
may be detected in peripheral blood, and that individuals that exhibit objective clinical response 
to such vaccine therapies tend to derive from the cohort of patients that display detectable 
increases in T cell responses post-vaccination (25-27). Our own pre-clinical studies support the 
effectiveness of DC1/peptide vaccination to elicit protective/therapeutic T cell-mediated 
immunity in melanoma models in vivo (49), supporting the hypothesis that DC1/peptide 
vaccination of advanced stage melanoma patients will result in increased quantities of specific 
CD8+ T cells in patient peripheral blood and that those individuals in which improved response 
to many peptides can be observed are those that are more likely to demonstrate clinical benefit.  
Method: Using fluorescently-labeled HLA-A2/peptide dextramer probes and intracellular 
staining for the Type-1 cytokine IFN-, we can simultaneously determine how the frequency of 
CD8+ T cells specific for TBVA peptides changes over time post-vaccination and how many of 


 
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these T cells are Type-1 effector T cells. By screening for in vitro reactivity of T cells against 
melanoma antigen-derived peptides, we will also be able to assess how vascular-targeting T cell 
responses may promote the corollary ability of the immune system to recognize melanoma-
associated peptide epitopes (i.e. “epitope spreading”) that is hypothesized by many to be a strong 
surrogate for the clinical efficacy of immunotherapeutic approaches (25-27).  
10.2 
Quantitation of CD8+ T cells, Treg, MDSC and blood vessels in melanoma biopsy 
tissue  
Rationale and Hypothesis: Tumor progression is believed to be linked to the accumulation of 
suppressor cell populations (both MDSC and Treg) and strong pro-angiogenic signals, as well as, 
“prevention” of Type-1 T cell recruitment within the tumor microenvironment (65-74). Our 
preliminary data in murine melanoma models support the ability of dasatinib (particularly when 
combined with DC1/peptide vaccines) to counteract these biologic endpoints in vivo (Figs. 1, 3). 
We hypothesize that such changes may also be evidenced in effectively treated melanoma 
patients by analyzing melanoma biopsies taken post- versus pre-treatment and that the greatest 
“normalization” of the tumor microenvironment will be observed after treatment with combined 
dasatinib + vaccine therapy.  
Method: Immunofluorescence microscopy will be used to analyze tumor sections of melanoma 
biopsies for expression of the markers CD8(T effector cells), CD11b + CD33 + lack of HLA-
DR (lineage-negative MDSC), CD11b + CD15 + lack of CD14 (neutrophilic MDSC), CD11b + 
CD14 + lack of CD15 (myeloid MDSC), CD4 + Foxp3 (Treg cells) and CD31 + NG2 (blood 
vessels). After staining and washing, sections will be covered in Gelvatol (Monsanto, St. Louis, 
MO) and a coverslip applied. Positively-stained cells will be quantitated by analyzing the images 
at a final magnification of x20 using Metamorph Imaging software (Molecular Devices, 
Sunnyvale, CA). 
10.3 
Treg analysis in PBMC 
Rationale and Hypothesis: Cancer patients have commonly also been shown to have elevated 
populational frequencies of Treg (based on the CD4+Foxp3+ phenotype) circulating in the 
peripheral blood. Alternate TKI, such as sunitinib, have been shown capable of reducing 
peripheral blood Treg levels within the first 4 week cycle of drug administration, in concert with 
a rebound in Type-1 T cell numbers and function in PBMC (59). Based on our pre-clinical data 
(Fig. 1), we hypothesize that dasatinib with provide a similar effect in melanoma patients and 
that those patients exhibiting the greatest degree of Treg reduction post-therapy will be more 
likely to respond favorably against the peptide epitopes contained in the vaccine formulation.  
Method: Peripheral blood cells will be analyzed by flow cytometry using specific antibodies 
against CD3 (all T cells), CD4 + Foxp3 (Treg), CD4 + CD25hi (Treg) Results will expressed as 
percentage of CD25+hi/Foxp3+ cells out of total CD3+/CD4+ viable cells.  


 
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10.4 
MDSC Analysis in PBMC 
Rationale and Hypothesis: Similar to Treg, levels of cells expressing an MDSC phenotype have 
been reported to be elevated in the peripheral blood of cancer patients, including patients with 
advanced-stage melanoma (60, 61). TKI, such as sunitinib and dasatinib can reduce the 
frequency of such suppressor cells to a variable degree when used as a therapy (ref. 60, Fig. 1). 
We hypothesize that melanoma patients treated with dasatinib will exhibit reduction in MDSC 
frequencies in PBMC, with the degree of loss correlating with the patient’s ability to respond 
favorably against the peptide epitopes contained in the vaccine formulation.  
Method: Analysis of MDSC percentages in patient PBMC will be performed using flow 
cytometry and anti-human antibodies against CD11b, CD11c, CD14, CD15, CD33 and HLA-DR 
as previously described (100).  
10.5 
EphA2 Protein Levels in tumor biopsies 
Rationale and Hypothesis: We have previously shown that drug treatments (including dasatinib 
in vitro) that promote the proteasome-dependent degradation of the tumor (and tumor vascular 
endothelial) cell-associated protein EphA2 lead to improved recognition by specific CD8+ T cells 
(refs. 101, 102 and Fig. 2). We hypothesize that administration of dasatinib to melanoma patients 
will promote the loss of EphA2 protein within the tumor lesion, leading to an enhancement in the 
sensitivity of EphA2+ cells in the tumor microenvironment to EphA2-specific CD8+ T cells that 
have been activated as a consequence of DC1/peptide-based vaccination.  
Method: Western blotting and immunofluorescence microscopy will be used to quantitate EphA2 
protein expression in melanoma biopsies pre- versus post-vaccination.  
10.6 
CXCL10 Levels in Patient Serum  
Rationale and Hypothesis: We and others have recently shown that therapeutic CD8+ T cells 
require the production of CXCR3 ligand chemokines within the tumor microenvironment in 
order to effectively home to these disease sites. Two recent clinical trials, including our 
DC1/glioma peptide vaccination trial in patients with brain tumors strongly support CXCL10 
(aka IP-10) as a chemokine associated with superior clinical outcome to immune-based therapy 
(88, 99). We hypothesize that this will also be the case in our DC1/TBVA peptide vaccinated 
patients with melanoma where Type-1 CXCR3+ responder T cells require a gradient of 
CXCL10/IP-10 (as detected in serum) in order to traffick to tumor sites in vivo.  
Method: Patient serum levels of CXCL10 will be monitored using Luminex fluorescent bead 
technology according to manufacturer's protocol. 
11. MEASUREMENT OF EFFECT 
Although objective tumor response is not the primary endpoint of this trial, patients with 
measurable disease will be assessed by standard criteria. For the purposes of this study, patients 


 
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should be re-evaluated every 8 weeks. In addition to a baseline scan, confirmatory scans will also 
be obtained ≥ 4 weeks following initial documentation of an objective response. 
11.1 
Antitumor Effect  
For the purposes of this study, patients should be re-evaluated for response every 8 weeks. In 
addition to a baseline scan, confirmatory scans should also be obtained no less than 4 weeks 
following initial documentation of objective response. 
Response and progression will be evaluated in this study using the new international criteria 
proposed by the revised Response Evaluation Criteria in Solid Tumors (RECIST) guideline 
(version 1.1; ref. 104). Changes in the largest diameter (uni-dimensional measurement) of the 
tumor lesions and the shortest diameter in the case of malignant lymph nodes are used in the 
RECIST criteria. 
11.1.1 
Definitions 
Evaluable for toxicity. All patients will be evaluable for toxicity from the time of their first 
treatment. 
Evaluable for objective response. Only those patients who have measurable disease present at 
baseline, have received at least one cycle of therapy, and have had their disease re-evaluated will 
be considered evaluable for response. These patients will have their response classified 
according to the definitions stated below. (Note:  Patients who exhibit objective disease 
progression prior to the end of cycle 1 will also be considered evaluable.) 
Evaluable Non-Target Disease Response. Patients who have lesions present at baseline that are 
evaluable but do not meet the definitions of measurable disease, have received at least one cycle 
of therapy, and have had their disease re-evaluated will be considered evaluable for non-target 
disease. The response assessment is based on the presence, absence, or unequivocal progression 
of the lesions.  
11.1.2 
Disease Parameters 
Measurable disease. Measurable lesions are defined as those that can be accurately measured in 
at least one dimension (longest diameter to be recorded) as > 20 mm by chest x-ray or as > 10 
mm with CT scan, MRI, or calipers by clinical exam. All tumor measurements must be recorded 
in millimeters (or decimal fractions of centimeters). 
Note: Tumor lesions that are situated in a previously irradiated area might or might not be 
considered measurable. If the investigator thinks it appropriate to include them, the conditions 
under which such lesions should be considered must be defined in the protocol. 
Malignant lymph nodes. To be considered pathologically enlarged and measurable, a lymph node 
must be > 15 mm in short axis when assessed by CT scan (CT scan slice thickness recommended 
to be no greater than 5 mm). At baseline and in follow-up, only the short axis will be measured 


 
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and followed. 
Non-measurable disease. All other lesions (or sites of disease), including small lesions (longest 
diameter < 10 mm or pathological lymph nodes with ≥ 10 to < 15 mm short axis), are considered 
non-measurable disease. Bone lesions, leptomeningeal disease, ascites, pleural/pericardial 
effusions, lymphangitis cutis/pulmonitis, inflammatory breast disease, and abdominal masses 
(not followed by CT or MRI), are considered as non-measurable. 
Note:  Cystic lesions that meet the criteria for radiographically defined simple cysts should not 
be considered as malignant lesions (neither measurable nor non-measurable) since they are, by 
definition, simple cysts. 
‘Cystic lesions’ thought to represent cystic metastases can be considered as measurable lesions, 
if they meet the definition of measurability described above. However, if non-cystic lesions are 
present in the same patient, these are preferred for selection as target lesions. 
Target lesions. All measurable lesions up to a maximum of 2 lesions per organ and 5 lesions in 
total, representative of all involved organs, should be identified as target lesions and recorded 
and measured at baseline. Target lesions should be selected on the basis of their size (lesions 
with the longest diameter), be representative of all involved organs, but in addition should be 
those that lend themselves to reproducible repeated measurements. It may be the case that, on 
occasion, the largest lesion does not lend itself to reproducible measurement in which 
circumstance the next largest lesion which can be measured reproducibly should be selected. A 
sum of the diameters (longest for non-nodal lesions, short axis for nodal lesions) for all target 
lesions will be calculated and reported as the baseline sum diameters. If lymph nodes are to be 
included in the sum, then only the short axis is added into the sum. The baseline sum diameters 
will be used as reference to further characterize any objective tumor regression in the measurable 
dimension of the disease. 
Non-target lesions. All other lesions (or sites of disease) including any measurable lesions over 
and above the 5 target lesions should be identified as non-target lesions and should also be 
recorded at baseline. Measurements of these lesions are not required, but the presence, absence, 
or in rare cases unequivocal progression of each should be noted throughout follow-up.  
11.1.3 
Methods for Evaluation of Measurable Disease 
All measurements should be taken and recorded in metric notation using a ruler or calipers. All 
baseline evaluations should be performed as closely as possible to the beginning of treatment and 
never more than 4 weeks before the beginning of the treatment. 
The same method of assessment and the same technique should be used to characterize each 
identified and reported lesion at baseline and during follow-up. Imaging-based evaluation is 
preferred to evaluation by clinical examination unless the lesion(s) being followed cannot be 


 
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imaged but are assessable by clinical exam. 
Clinical lesions. Clinical lesions will only be considered measurable when they are superficial 
(e.g., skin nodules and palpable lymph nodes) and  10 mm diameter as assessed using calipers 
(e.g., skin nodules). In the case of skin lesions, documentation by color photography, including a 
ruler to estimate the size of the lesion, is recommended. 
Chest x-ray. Lesions on chest x-ray are acceptable as measurable lesions when they are clearly 
defined and surrounded by aerated lung. However, CT is preferable. 
Conventional CT and MRI. This guideline has defined measurability of lesions on CT scan based 
on the assumption that CT slice thickness is 5 mm or less. If CT scans have slice thickness 
greater than 5 mm, the minimum size for a measurable lesion should be twice the slice thickness. 
MRI is also acceptable in certain situations (e.g. for body scans).  
Use of MRI remains a complex issue. MRI has excellent contrast, spatial, and temporal 
resolution; however, there are many image acquisition variables involved in MRI, which greatly 
impact image quality, lesion conspicuity, and measurement. Furthermore, the availability of MRI 
is variable globally. As with CT, if an MRI is performed, the technical specifications of the 
scanning sequences used should be optimized for the evaluation of the type and site of disease. 
Furthermore, as with CT, the modality used at follow-up should be the same as was used at 
baseline and the lesions should be measured/assessed on the same pulse sequence. It is beyond 
the scope of the RECIST guidelines to prescribe specific MRI pulse sequence parameters for all 
scanners, body parts, and diseases. Ideally, the same type of scanner should be used and the 
image acquisition protocol should be followed as closely as possible to prior scans. Body scans 
should be performed with breath-hold scanning techniques, if possible. 
PET-CT. At present, the low dose or attenuation correction CT portion of a combined PET-CT is 
not always of optimal diagnostic CT quality for use with RECIST measurements. However, if 
the site can document that the CT performed as part of a PET-CT is of identical diagnostic 
quality to a diagnostic CT (with IV and oral contrast), then the CT portion of the PET-CT can be 
used for RECIST measurements and can be used interchangeably with conventional CT in 
accurately measuring cancer lesions over time. Note, however, that the PET portion of the CT 
introduces additional data which may bias an investigator if it is not routinely or serially 
performed.  
Ultrasound. Ultrasound is not useful in assessment of lesion size and should not be used as a 
method of measurement. Ultrasound examinations cannot be reproduced in their entirety for 
independent review at a later date and, because they are operator dependent, it cannot be 
guaranteed that the same technique and measurements will be taken from one assessment to the 
next. If new lesions are identified by ultrasound in the course of the study, confirmation by CT or 
MRI is advised. If there is concern about radiation exposure at CT, MRI may be used instead of 


 
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CT in selected instances. 
Endoscopy, Laparoscopy. The utilization of these techniques for objective tumor evaluation is 
not advised. However, such techniques may be useful to confirm complete pathological response 
when biopsies are obtained or to determine relapse in trials where recurrence following complete 
response (CR) or surgical resection is an endpoint. 
Cytology, Histology. These techniques can be used to differentiate between partial responses 
(PR) and complete responses (CR) in rare cases (e.g., residual lesions in tumor types, such as 
germ cell tumors, where known residual benign tumors can remain). 
The cytological confirmation of the neoplastic origin of any effusion that appears or worsens 
during treatment when the measurable tumor has met criteria for response or stable disease is 
mandatory to differentiate between response or stable disease (an effusion may be a side effect of 
the treatment) and progressive disease. 
FDG-PET. While FDG-PET response assessments need additional study, it is sometimes 
reasonable to incorporate the use of FDG-PET scanning to complement CT scanning in 
assessment of progression (particularly possible 'new' disease). New lesions on the basis of FDG-
PET imaging can be identified according to the following algorithm: 
 
Negative FDG-PET at baseline, with a positive FDG-PET at follow-up is a sign of PD 
based on a new lesion. 
 
No FDG-PET at baseline and a positive FDG-PET at follow-up:  If the positive FDG-
PET at follow-up corresponds to a new site of disease confirmed by CT, this is PD. If the 
positive FDG-PET at follow-up is not confirmed as a new site of disease on CT, 
additional follow-up CT  scans are needed to determine if there is truly progression 
occurring at that site (if so, the date of PD will be the date of the initial abnormal FDG-
PET scan). If the positive FDG-PET at follow-up corresponds to a pre-existing site of 
disease on CT that is not progressing on the basis of the anatomic images, this is not PD. 
 
FDG-PET may be used to upgrade a response to a CR in a manner similar to a biopsy in 
cases where a residual radiographic abnormality is thought to represent fibrosis or 
scarring. The use of FDG-PET in this circumstance should be prospectively described in 
the protocol and supported by disease-specific medical literature for the indication. 
However, it must be acknowledged that both approaches may lead to false positive CR 
due to limitations of FDG-PET and biopsy resolution/sensitivity. 
Note: A ‘positive’ FDG-PET scan lesion means one which is FDG avid with an uptake greater 
than twice that of the surrounding tissue on the attenuation corrected image. 
 
 


 
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11.1.4 
Response Criteria 
11.1.4.1 Evaluation of Target Lesions 
Complete Response (CR): 
Disappearance of all target lesions. Any pathological lymph nodes 
(whether target or non-target) must have reduction in short axis to <10 mm. 
Partial Response (PR): 
At least a 30% decrease in the sum of the diameters of target 
lesions, taking as reference the baseline sum diameters. 
Progressive Disease (PD): 
At least a 20% increase in the sum of the diameters of target 
lesions, taking as reference the smallest sum on study (this includes the baseline sum if that is the 
smallest on study). In addition to the relative increase of 20%, the sum must also demonstrate an 
absolute increase of at least 5 mm. (Note:  the appearance of one or more new lesions is also 
considered progressions). 
Stable Disease (SD): Neither sufficient shrinkage to qualify for PR nor sufficient increase to 
qualify for PD, taking as reference the smallest sum diameters while on study. 
11.1.4.2 Evaluation of Non-Target Lesions 
Complete Response (CR): 
Disappearance of all non-target lesions and normalization of tumor 
marker level. All lymph nodes must be non-pathological in size (< 10 mm short axis). 
Note: If tumor markers are initially above the upper normal limit, they must normalize for a 
patient to be considered in complete clinical response. 
Non-CR/Non-PD: 
Persistence of one or more non-target lesion(s) and/or maintenance of 
tumor marker level above the normal limits. 
Progressive Disease (PD): 
Appearance of one or more new lesions and/or unequivocal 
progression of existing non-target lesions. Unequivocal progression should not normally trump 
target lesion status. It must be representative of overall disease status change, not a single lesion 
increase.  
Although a clear progression of “non-target” lesions only is exceptional, the opinion of the 
treating physician should prevail in such circumstances, and the progression status should be 
confirmed at a later time by the review panel (or Investigator). 
11.1.4.3 Evaluation of Best Overall Response 
The best overall response is the best response recorded from the start of the treatment until 
disease progression/recurrence (taking as reference for progressive disease the smallest 
measurements recorded since the treatment started). The patient's best response assignment will 
depend on the achievement of both measurement and confirmation criteria. 
 
 


 
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For Patients with Measurable Disease (i.e., Target Disease) 
Target 
Lesions 
Non-Target 
Lesions 
New 
Lesions 
Overall 
Response 
Best Overall Response when Confirmation is 
Required* 
CR 
CR 
No 
CR 
≥4 wks. Confirmation** 
CR 
Non-CR/Non-PD 
No 
PR 
≥4 wks. Confirmation** 
CR 
Not evaluated 
No 
PR 
PR 
Non-CR/Non-PD/ 
not evaluated 
No 
PR 
SD 
Non-CR/Non-
PD/not evaluated 
No 
SD 
Documented at least once ≥4 wks. from 
baseline** 
PD 
Any 
Yes or No 
PD 
no prior SD, PR or CR 
Any 
PD*** 
Yes or No 
PD 
Any 
Any 
Yes 
PD 
* 
See RECIST 1.1 manuscript for further details on what is evidence of a new lesion. 
** 
Only for non-randomized trials with response as primary endpoint. 
*** 
In exceptional circumstances, unequivocal progression in non-target lesions may be accepted as disease 
progression. 
 
Note: 
Patients with a global deterioration of health status requiring discontinuation of treatment without 
objective evidence of disease progression at that time should be reported as “symptomatic 
deterioration.”  Every effort should be made to document the objective progression even after 
discontinuation of treatment. 
 
For Patients with Non-Measurable Disease (i.e., Non-Target Disease) 
Non-Target Lesions 
New Lesions 
Overall Response 
CR 
No 
CR 
Non-CR/non-PD 
No 
Non-CR/non-PD* 
Not all evaluated 
No 
not evaluated 
Unequivocal PD 
Yes or No 
PD 
Any 
Yes 
PD 
*  ‘Non-CR/non-PD’ is preferred over ‘stable disease’ for non-target disease since SD is 
increasingly used as an endpoint for assessment of efficacy in some trials so to assign 
this category when no lesions can be measured is not advised 
 
11.1.5 
Duration of Response 
Duration of overall response: The duration of overall response is measured from the time 
measurement criteria are met for CR or PR (whichever is first recorded) until the first date that 
recurrent or progressive disease is objectively documented (taking as reference for progressive 
disease the smallest measurements recorded since the treatment started). 
The duration of overall CR is measured from the time measurement criteria are first met for CR 
until the first date that progressive disease is objectively documented. 
 
Duration of stable disease: Stable disease is measured from the start of the treatment until the 
criteria for progression are met, taking as reference the smallest measurements recorded since the 
treatment started, including the baseline measurements.  
 
 


 
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11.1.6 
Progression-Free Survival 
Progression-free survival (PFS) is defined as the duration of time from start of treatment to time 
of progression or death, whichever occurs first. 
12. STATISTICAL CONSIDERATIONS 
12.1 
Study Design 
The primary objective of this study is to evaluate the effects of combination therapy of dasatinib 
and vaccine on immune response rate. Here a patient who responded to at last 3 of the 6 peptides 
is considered to have a positive immune response. The secondary objectives of this trial include 
evaluation of clinical response rate, overall survival (OS), progression free survival (PFS), and 
immunological endpoints, which include number of CD8+ T cells, MDSC/Treg regulatory cells 
and blood vessels in tumor lesions, level of EphA2 protein expressed within the tumor lesion and 
the level of the CXCL10/IP-10 chemokine in patient serum pre- versus post-treatment.  
We expect that the proportion of patients that are not evaluable for immune endpoint will be 10% 
or less. To ensure a total of 28 evaluable patients, up to 31 evaluable patients will be randomized 
in 1:1 ratio to receive either: 
A. Vaccine alone starting in the first 28-day cycle followed by vaccine combined with daily 
dasatinib starting on the first day of the second cycle (Arm A) 
B. Vaccine combined with daily dasatinib starting on the first day of cycle 1 (Arm B) 
The UPCI randomizer will be used for the randomization. The randomization will be stratified 
by BRAF mutation status. 
12.2 
Safety Monitoring 
The dose limiting toxicity (DLT) is defined in section 7.2.2. Bayesian analysis will be used to 
continuously monitor the DLTs in both arms to ensure that the study will continue enrollment 
without interruption as long as the underlying true DLT rate is 20% or less. The posterior 
probability (PPr) will be calculated from the study’s accumulated data on DLTs. A weak prior 
distribution of Beta (0.1, 1.9) was used. If PPr > 0.7, that is, there is a 70% or higher chance that 
more than 20% of the patients will experience DLTs, the enrollment of the study arm will stop 
pending a review of both the data and the study’s implementation.  
The following table listed the suspension rule based on the Bayesian analysis calculation.  
Bayesian rule for suspending enrollment 
Suspend accrual if n patients experience a DLT 
in N patients treated 
2 
2-4 
3 
5-9 
4 
10-14 
 
The operating characteristics of this study design can be expressed in terms of probability of 
early suspending the enrollment for the combination arm under the assumptions of various the 


 
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true rates of SAEs. The following table calculated these probabilities based on the simulation of 
10,000 hypothetical trials. 
Frequentist properties of Bayesian rule for suspending enrollment 
Probability of DLTs 
Probability of suspending the enrollment 
Median sample size for the combination arm 
0.05 
0.02 
14 
0.1 
0.09 
14 
0.15 
0.22 
14 
0.2 
0.36 
14 
0.25 
0.53 
13 
0.3 
0.67 
8 
0.35 
0.79 
7 
0.4 
0.88 
4 
 
12.3 
Sample Size/Accrual Rate 
We estimated an accrual rate of approximately 1-2 patients per month therefore, we the accrual 
will be completed within 20 months. We plan a 12 month follow up after the last patient entered 
the study. Therefore, the study will be completed within 32 months after the initiation of the 
study.  
The power calculation for the proposed study is based on the comparison of the positive immune 
response rate between the two treatment arms. Based on a preliminary data of 15 patients, the 
immune response rate for the vaccine only arm is close to 0%. With 14 evaluable patients in each 
arm, we will have 80% power to distinguish a difference between 5% immune response rate (of 
the vaccine arm) and 50% immune response rate (of the combination arm) with 1-sided Fisher 
exact test (= 0.025). 
12.4 
Data Analysis 
12.4.1 
Analysis Sets 
Evaluable Patients are patients who meet all of the protocol inclusion/exclusion criteria and 
begin treatment with the protocol assigned regimen. All evaluable patients will be used in the 
analysis of safety, immune response, clinical response, OS and PFS. 
12.4.2 
Baseline Characteristics 
Baseline characteristics on all evaluable patients will be provided on demographic variables (age, 
sex, race/ethnicity), performance status, laboratory parameters, prior treatments, and disease 
characteristics, including tumor size, number of nodes involved, and metastatic sites. 
12.4.3 
Safety Profile 
NCI CTCAE version 4.0 will be used to evaluate the serious adverse events (SAEs) in each cycle 
of the treatment, and for 30 days beyond the last protocol specified treatment. Sever AEs rate for 
each treatment arm will be calculated and the corresponding exact 95% confidence interval (CI) 
will be provided. All adverse events that are determined to be possibly, probably or definitely 
related to treatment will be tabulated according to grade and type (according to the NCI CTCAE, 


 
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Version 4.0). For each adverse event category, frequencies will be tabulated by treatment group 
according to the highest grade per patient within 30 days after any study treatment. 
12.4.4 
Efficacy Analysis 
The immune response rate, defined as proportion of patients that responded to  3 out of the 6 
peptides, for each study arm will be calculated with 95% exact CI. The clinical response rate for 
each study arm will be estimated by the percentage of patients achieving CR or PR by RECIST 
criteria, with corresponding exact 95% CI. Both immune response rate and clinical response rate 
of the two treatment groups will be compared using Fisher exact test. As an exploratory analysis, 
we will also evaluate and compare the immune response for the B-raf mutant carrier and non-
carrier in similar manner as described above.  
The Kaplan-Meier estimate of PFS and OS with corresponding 95% confidence band will be 
provided for each dose level. The corresponding median survival time (with 95% confidence 
limits) will be determined, along with OS and PFS estimates at selected time points. The exact 
log rank test will be used to compare the PFS and OS between the two study arms.  
Exploratory analysis will be conducted to study the association between the positive immune 
response and:  
a. Objective clinical response. 
b. CD8+ T cell infiltration in tumor after cycle 1. 
c. Reduction in suppressor cells in the tumor and blood.  
d. Reduction in blood vessel density in the tumor after cycle 1. 
e. Reduction in EphA2 protein expression in tumor after cycle 1. 
f. Increased level of the CXCR3 ligand chemokine CXCL 10/IP-10 in patient serum after 
cycle 1.  
Chi-square (or Fisher exact) test will be used to test the association between immune response 
and the categorical outcomes (e.g. objective clinical response). Wilcoxon test will be used to 
compare the continuous outcomes (e.g. CD8+ T cell infiltration, suppressor cell populations, 
tumor blood vessel density, EphA2 protein expression, chemokine level) between the immune 
responders and non-responders. 
12.5 
Reporting and Exclusions 
12.5.1 
Evaluation of toxicity 
All patients will be evaluated for toxicity from the time of their first treatment with either 
vaccine alone or dasatinib with vaccine. 
12.5.2 
Evaluation of response 
All patients included in the study must be assessed for response to treatment, even if there are 
major protocol treatment deviations or if they are ineligible. Each patient will be assigned one of 
the following categories: 1) complete response, 2) partial response, 3) stable disease, 4) 


 
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progressive disease, 5) early death from malignant disease, 6) early death from toxicity, 7) early 
death because of other cause, or 9) unknown (not assessable, insufficient data). [Note:  By 
arbitrary convention, category 9 usually designates the “unknown” status of any type of data in a 
clinical database.] 
All of the patients who met the eligibility criteria (with the possible exception of those who 
received no study medication) should be included in the main analysis of the response rate. 
Patients in response categories 4-9 should be considered to have a treatment failure (disease 
progression). Thus, an incorrect treatment schedule or drug administration does not result in 
exclusion from the analysis of the response rate. Precise definitions for categories 4-9 will be 
protocol specific.  
All conclusions should be based on all eligible patients. Sub-analyses may then be performed on 
the basis of a subset of patients, excluding those for whom major protocol deviations have been 
identified (e.g., early death due to other reasons, early discontinuation of treatment, major 
protocol violations, etc.). However, these sub-analyses may not serve as the basis for drawing 
conclusions concerning treatment efficacy, and the reasons for excluding patients from the 
analysis should be clearly reported. The 95% confidence intervals should also be provided. 
13. DATA SAFETY MONITORING PLAN 
Investigator/Sub-investigators, regulatory, CRS management, clinical research coordinators, 
clinical research associates, data managers, and clinic staff meet monthly in disease center Data 
Safety Monitoring Boards (DSMB) to review and discuss study data to include, but not limited 
to, the following: 
 
serious adverse events 
 
subject safety issues 
 
recruitment issues 
 
accrual 
 
protocol deviations 
 
unanticipated problems 
 
breaches of confidentiality 
 
Minutes from the DSMB meetings are available to anyone unable to attend the center DSMB. 
 
All toxicities encountered during the study will be evaluated on an ongoing basis according to 
the NCI Common Toxicity Criteria version 4.0. All study treatment associated adverse events 
that are serious, at least possibly related and unexpected will be reported to the IRB. Any 
modifications necessary to ensure subject safety and decisions to continue, or close the trial to 
accrual are also discussed during these meetings. If any literature becomes available which 
changes the risk/benefit ratio or suggests that conducting the trial is no longer ethical, the IRB 
will be notified in the form of an Unanticipated Problem submission and the study may be 


 
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terminated. 
All study data reviewed and discussed during these meetings will be kept confidential. Any 
breach in subject confidentiality will be reported to the IRB in the form of an Unanticipated 
Problem submission. The summaries of these meetings are forwarded to the UPCI DSMC which 
also meets monthly following a designated format. 
For all research protocols, there will be a commitment to comply with the IRB’s policies for 
reporting unanticipated problems involving risk to subjects or others (including adverse events). 
DSMC progress reports, to include a summary of all serious adverse events and modifications, 
and approval will be submitted to the IRB at the time of renewal. 
Protocols with subjects in long-term (survival) follow-up or protocols in data analysis only, will 
be reviewed twice a year rather than monthly by the disease center DSMB. 
Both the UPCI DSMC as well as the individual disease center DSMB have the authority to 
suspend accrual or further investigate treatment on any trial based on information discussed at 
these meetings. 
All records related to this research study will be stored in a locked environment. Only the 
researchers affiliated with the research study and their staff will have access to the research 
records. 
14. ADVERSE EVENT REPORTING 
14.1 
Adverse event definitions 
Adverse event (AE) means any untoward medical occurrence associated with the use of a drug in 
humans, whether or not considered drug related. 
Suspected adverse reaction means any adverse event for which there is a reasonable possibility 
that the drug caused the adverse event. Suspected adverse reaction implies a lesser degree of 
certainty about causality than adverse reaction, which means any adverse event caused by a drug. 
Reasonable possibility, for the purpose of IND safety reporting, means there is evidence 
to suggest a causal relationship between the drug and the adverse event. 
Life-threatening adverse event or life-threatening suspected adverse reaction is considered "life-
threatening" if, in the view of the sponsor-investigator, its occurrence places the patient or 
subject at immediate risk of death. It does not include an adverse event or suspected adverse 
reaction that, had it occurred in a more severe form, might have caused death. 
Serious adverse event or serious suspected adverse reaction is considered "serious" if, in the 
view of the sponsor-investigator, it results in any of the following outcomes: death, a life-
threatening adverse event, inpatient hospitalization or prolongation of existing hospitalization, a 
persistent or significant incapacity or substantial disruption of the ability to conduct normal life 


 
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functions, or a congenital anomaly/birth defect. 
Important medical events that may not result in death, be life-threatening, or require 
hospitalization may be considered serious when, based upon appropriate medical 
judgment, they may jeopardize the patient or subject and may require medical or surgical 
intervention to prevent one of the outcomes listed in this definition. Examples of such 
medical events include allergic bronchospasm requiring intensive treatment in the 
emergency room or at home, blood dyscrasias or convulsions that do not result in 
inpatient hospitalization, or the development of drug dependency or drug abuse. 
Unexpected adverse event or unexpected suspected adverse reaction is considered "unexpected" 
if it is not listed in the investigator brochure or is not listed at the specificity or severity that has 
been observed; or, if an investigator brochure is not required or available, is not consistent with 
the risk information described in the general investigational plan or elsewhere in the current 
application, as amended. 
For example, under this definition, hepatic necrosis would be unexpected (by virtue of 
greater severity) if the investigator brochure referred only to elevated hepatic enzymes or 
hepatitis. Similarly, cerebral thromboembolism and cerebral vasculitis would be 
unexpected (by virtue of greater specificity) if the investigator brochure listed only 
cerebral vascular accidents. “Unexpected,” as used in this definition, also refers to 
adverse events or suspected adverse reactions that are mentioned in the investigator 
brochure as occurring with a class of drugs or as anticipated from the pharmacological 
properties of the drug, but are not specifically mentioned as occurring with the particular 
drug under investigation. 
14.2 
Recording/Reporting requirements 
14.2.1 
Eliciting AE Information 
Research subjects will be routinely questioned about AEs at study visits. 
14.2.2 
Recording Requirements 
All observed or volunteered adverse events (serious or non-serious) and abnormal test findings, 
regardless of study group or suspected causal relationship to the study drug(s) will be recorded in 
the subjects’ case histories. For all adverse events, sufficient information will be pursued and/or 
obtained so as to permit 1) an adequate determination of the outcome of the event (i.e., whether 
the event should be classified as a serious adverse event) and; 2) an assessment of the casual 
relationship between the adverse event and the study drug(s). 
AEs or abnormal test findings felt to be associated with the investigational drug or study 
treatment(s) will be followed until the event (or its sequelae) or the abnormal test finding 
resolves or stabilizes at a level acceptable to the Sponsor-Investigator. 


 
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14.2.2.1 Abnormal Test Findings 
An abnormal test finding will be classified as an adverse event if one or more of the following 
criteria are met: 
 
The test finding is accompanied by clinical symptoms. 
 
The test finding necessitates additional diagnostic evaluation(s) or medical/surgical 
intervention; including significant additional concomitant drug treatment or other 
therapy. 
Note: simply repeating a test finding, in the absence of any of the other listed criteria, 
does not constitute an AE. 
 
The test finding leads to a change in study drug dosing or discontinuation of subject 
participation in the clinical study. 
 
The test finding is considered an AE by the Sponsor-Investigator of the IND application. 
 
14.2.2.2 Causality and severity assessment 
The Sponsor-Investigator of the IND application will promptly review documented adverse 
events and abnormal test findings to determine 1) if the abnormal test finding should be 
classified as an adverse event; 2) if there is a reasonable possibility that the adverse event was 
caused by the study drug(s); and 3) if the adverse event meets the criteria for a serious adverse 
event. 
If the Sponsor-Investigator’s final determination of causality is “unknown and of questionable 
relationship to the study drug(s)”, the adverse event will be classified as associated with the use 
of the study drug(s) for reporting purposes. If the Sponsor-Investigator’s final determination of 
causality is “unknown but not related to the study drug(s)”, this determination and the rationale 
for the determination will be documented in the respective subject’s case history. 
14.3 
Reporting of Suspected Adverse Reactions 
All events meeting the definition of a serious adverse event should be recorded on a MedWatch 
3500A Form 
(http://www.fda.gov/downloads/AboutFDA/ReportsManualsForms/Forms/UCM048334.pdf) or 
departmental SAE form. Copies should be sent to: 
1. Investigator-Sponsor 
2. crssafetysubmissions@upmc.edu 
3. Local Institutional Review Board per institutional reporting requirements 
 
In addition to completing appropriate patient demographic and suspect medication information, 
the report should include as applicable the following information that is available at the time of 
report within the Event Description (section 5) of the MedWatch 3500A form: 
• CTCAE term(s) and grade(s) 
• Current status of study drug 


 
UPCI 12-048 
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• All interventions to address the AE (testing and result, treatment and response) 
• Hospitalization and/or discharge dates 
• Event relationship to study drug 
 
Follow-up reports: 
Additional information may be added to a previously submitted report by adding to the original 
MedWatch 3500A report and submitting it as follow-up or creating supplemental summary 
information and submitting it as follow-up with the original MedWatch 3500A form. 
14.4 
Review of Safety Information: Sponsor Responsibilities[1] 
The sponsor must promptly review all information relevant to the safety of the drug obtained or 
otherwise received by the sponsor from foreign or domestic sources, including information 
derived from any clinical or epidemiological investigations, animal or in vitro studies, reports in 
the scientific literature, and unpublished scientific papers, as well as reports from foreign 
regulatory authorities and reports of foreign commercial marketing experience for drugs that are 
not marketed in the United States. 
14.5 
Reporting of Adverse Reactions to the FDA 
14.5.1 
Written IND safety reports 
The Sponsor-Investigator will submit a written IND Safety Report (i.e., completed FDA Form 
3500 A) to the responsible new drug review division of the FDA for any observed or volunteered 
adverse event that is determined to be a serious and unexpected, suspected adverse reaction. 
Each IND Safety Report will be prominently labeled, “IND Safety Report”, and a copy will be 
provided to all participating investigators (if applicable) and sub-investigators. 
Written IND Safety Reports will be submitted to the FDA as soon as possible and, in no event, 
later than 15 calendar days following the Sponsor-Investigator’s receipt of the respective adverse 
event information and determination that it meets the respective criteria for reporting. 
For each written IND Safety Report, the Sponsor-Investigator will identify all previously 
submitted IND Safety Reports that addressed a similar suspected adverse reaction experience and 
will provide an analysis of the significance of newly reported, suspected adverse reaction in light 
of the previous, similar report(s) or any other relevant information. 
Relevant follow-up information to an IND Safety Report will be submitted to the applicable 
review division of the FDA as soon as the information is available and will be identified as such 
(i.e., “Follow-up IND Safety Report”). 
If the results of the Sponsor-Investigator’s follow-up investigation show that an adverse event 
that was initially determined to not require a written IND Safety Report does, in fact, meet the 
requirements for reporting; the Sponsor-Investigator will submit a written IND Safety Report as 
                                                 
[1] 21 CFR Sec. 312.50 


 
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soon as possible, but in no event later than 15 calendar days, after the determination was made. 
14.5.2 
Telephoned IND safety reports – fatal or life-threatening suspected adverse reactions 
In addition to the subsequent submission of a written IND Safety Report (i.e., completed FDA 
Form 3500A), the Sponsor-Investigator will notify the responsible review division of the FDA 
by telephone or facsimile transmission of any unexpected, fatal or life-threatening suspected 
adverse reaction.  
The telephone or facsimile transmission of applicable IND Safety Reports will be made as soon 
as possible but in no event later than 7 calendar days after the Sponsor-Investigator’s receipt of 
the respective adverse event information and determination that it meets the respective criteria 
for reporting. 
14.6 
Reporting adverse events to the responsible IRB 
In accordance with applicable policies of the University of Pittsburgh Institutional Review Board 
(IRB), the Sponsor-Investigator will report, to the IRB, any observed or volunteered adverse 
event that is determined to be 1) associated with the investigational drug or study treatment(s); 
2) serious; and 3) unexpected. Adverse event reports will be submitted to the IRB in accordance 
with the respective IRB procedures. 
Applicable adverse events will be reported to the IRB as soon as possible and, in no event, later 
than 10 calendar days following the sponsor-investigator’s receipt of the respective information. 
Adverse events which are 1) associated with the investigational drug or study treatment(s); 2) 
fatal or life-threatening; and 3) unexpected will be reported to the IRB within 24 hours of the 
Sponsor-Investigator’s receipt of the respective information. 
Follow-up information to a reported adverse event will be submitted to the IRB as soon as the 
relevant information is available. If the results of the Sponsor-Investigator’s follow-up 
investigation show that an adverse event that was initially determined to not require reporting to 
the IRB does, in fact, meet the requirements for reporting; the Sponsor-Investigator will report 
the adverse event to the IRB as soon as possible, but in no event later than 10 calendar days, after 
the determination was made. 
15. WITHDRAWAL OF SUBJECTS DUE TO ADVERSE EVENTS 
Therapy will be immediately discontinued for any grade 4 toxicity that becomes apparent or any 
toxicity outlined in the Regimen Limiting Toxicities Section 9.1.4. Study treatment will be 
reinitiated for any grade 3 (except toxicities outlined in the Regimen Limiting Toxicities) 
toxicity pending the reversal of such toxicity after withholding treatment. In the event of any 
adverse effect, appropriate medical treatment will be instituted and study treatment will be 
discontinued if the above toxicity remains. 
 
 


 
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16. QUALITY CONTROL AND QUALITY ASSURANCE 
Independent monitoring of the clinical study for protocol and Guidelines on Good Clinical 
Practice compliance will be conducted periodically (i.e., at a minimum of annually) by qualified 
staff of the Education and Compliance Office – Human Subject Research, Research Conduct and 
Compliance Office, University of Pittsburgh. 
The Investigator (i.e., the study site principal investigator) and the University of Pittsburgh and 
University of Pittsburgh Medical Center will permit direct access of the study monitors and 
appropriate regulatory authorities to the study data and to the corresponding source data and 
documents to verify the accuracy of this data. 
17. DATA HANDLING AND RECORD-KEEPING 
The Investigator (i.e., the study site principal investigator) will maintain records in accordance 
with Good Clinical Practice. 
The investigator will retain the specified records and reports for up to 2 years after the marketing 
application is approved for the investigational drug; or, if a marketing application is not 
submitted or approved for the investigational drug, until 2 years after investigations under the 
IND have been discontinued and the FDA so notified. 
18. ETHICS 
18.1 
Institutional Review Board (IRB) Approval 
The investigator (i.e., the study site principal investigator) will obtain, from the University of 
Pittsburgh Institutional Review Board (IRB), prospective approval of the clinical protocol and 
corresponding informed consent form(s); modifications to the clinical protocol and 
corresponding informed consent forms, and advertisements (i.e., directed at potential research 
subjects) for study recruitment, if applicable. 
The only circumstance in which a deviation from the current IRB-approved clinical 
protocol/consent form(s) may be initiated in the absence of prospective IRB approval is to 
eliminate an apparent immediate hazard to the research subject(s). In such circumstances, the 
investigator will promptly notify the University of Pittsburgh IRB of the deviation. 
The University of Pittsburgh IRB operates in compliance with FDA regulations at 21 CFR Parts 
50 and 21 CFR 56, and in conformance with applicable International Conference on 
Harmonization (ICH) Guidelines on Good Clinical Practice. 
In the event that the University of Pittsburgh IRB requires, as a condition of approval, substantial 
changes to a clinical protocol submitted under an FDA-accepted IND application, or in the event 
of an sponsor’s decision to modify the previously accepted clinical protocol, the sponsor will 
submit (i.e., in advance of implementing the change) a Protocol Amendment to the IND 
describing any change that significantly affects the safety of subjects, the scope of the 


 
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investigation, or the scientific quality of the study. Examples of protocol changes requiring the 
submission of a Protocol Amendment include: 
 Any increase in drug dosage or duration of exposure of individual subjects to the 
investigational drug beyond that described in the current protocol, or any significant 
increase in the number of subjects under study. 
 Any significant change in the design of the protocol (such as the addition or deletion of a 
control group). 
 The addition of a new test or procedure that is intended to improve monitoring for, or 
reduce the risk of, a side effect or AE; or the dropping of a test intended to monitor the 
safety of the investigational drug. 
18.2 
Ethical and Scientific Conduct of the Clinical Study 
The clinical study will be conducted in accordance with the current IRB-approved clinical 
protocol; ICH Guidelines on Guidelines on Good Clinical Practice; and relevant policies, 
requirements, and regulations of the University of Pittsburgh IRB, University of Pittsburgh and 
University of Pittsburgh Medical Center, Commonwealth of Pennsylvania, and applicable federal 
agencies. 
18.3 
Subject Informed Consent 
The investigator (i.e., the study site principal investigator) will make certain that an appropriate 
informed consent process is in place to ensure that potential research subjects, or their authorized 
representatives, are fully informed about the nature and objectives of the clinical study, the 
potential risks and benefits of study participation, and their rights as research subjects. The 
investigator, or a sub-investigator(s) designated by the sponsor, will obtain the written, signed 
informed consent of each subject, or the subject’s authorized representative, prior to performing 
any study-specific procedures on the subject. The date and time that the subject, or the subject’s 
authorized representative, signs the informed consent form and a narrative of the issues discussed 
during the informed consent process will be documented in the subject’s case history. The 
investigator or sub-investigator will retain the original copy of the signed informed consent form, 
and a copy will be provided to the subject, or to the subject’s authorized representative. 
The investigator will make certain that appropriate processes and procedures are in place to 
ensure that ongoing questions and concerns of enrolled subjects are adequately addressed and 
that the subjects are informed of any new information that may affect their decision to continue 
participation in the clinical study. In the event of substantial changes to the clinical study or the 
risk-to-benefit ratio of study participation, the investigator will obtain the informed consent of 
enrolled subjects for continued participation in the clinical study. 
 
 


 
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APPENDIX A 
 
Performance Status Criteria 
 
ECOG Performance Status Scale 
Karnofsky Performance Scale 
Grade 
Descriptions 
Percent 
Description 
0 
Normal activity. Fully active, able to 
carry on all pre-disease performance 
without restriction. 
100 
Normal, no complaints, no evidence of 
disease. 
90 
Able to carry on normal activity; minor 
signs or symptoms of disease. 
1 
Symptoms, but ambulatory. 
Restricted in physically strenuous 
activity, but ambulatory and able to 
carry out work of a light or sedentary 
nature (e.g., light housework, office 
work). 
80 
Normal activity with effort; some signs 
or symptoms of disease. 
70 
Cares for self, unable to carry on 
normal activity or to do active work. 
2 
In bed <50% of the time. Ambulatory 
and capable of all self-care, but 
unable to carry out any work 
activities. Up and about more than 
50% of waking hours. 
60 
Requires occasional assistance, but is 
able to care for most of his/her needs. 
50 
Requires considerable assistance and 
frequent medical care. 
3 
In bed >50% of the time. Capable of 
only limited self-care, confined to bed 
or chair more than 50% of waking 
hours. 
40 
Disabled, requires special care and 
assistance. 
30 
Severely disabled, hospitalization 
indicated. Death not imminent. 
4 
100% bedridden. Completely 
disabled. Cannot carry on any self-
care. Totally confined to bed or chair. 
20 
Very sick, hospitalization indicated. 
Death not imminent. 
10 
Moribund, fatal processes progressing 
rapidly. 
5 
Dead. 
0 
Dead.