Clinical trial NCT02720185
Conditions: Breast Neoplasms Triple Negative Breast Neoplasms
Sponsor: University of Wisconsin, Madison
Start date: 2017-05-03; Completion date: 2022-02-08
Enrollment: 5 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
Primary Objective:

To determine if dasatinib, an inhibitor of the Src family kinases, can prevent the nuclear translocation of the epidermal growth factor receptor (EGFR) in Stage I-III, nuclear EGFR positive, triple negative breast cancers (TNBC).

Secondary Objectives:

1. To examine the safety and tolerability of dasatinib in patients with operable TNBC
2. To explore potential intracellular mechanisms which impact dasatinib effect on cellular localization of EGFR in operable TNBC.
3. To examine the pathologic complete response (pCR) rates to standard neoadjuvant chemotherapy in nEGFR+ TNBC
4. To examine breast cancer recurrence rates and patterns of metastatic recurrent in nEGFR+ TNBC

Detailed description:

Arms: arm 1: Dasatinib 100mg for 7-10 days until day prior to surgery
Interventions: intervention 1: Dasatinib intervention 2: Conventional Surgery intervention 3: Laboratory Biomarker Analysis
Intervention descriptions: intervention 1: 100mg oral once daily dasatinib taken for 7-10 days up to the day prior to planned surgery or research biopsy (neoadjuvant chemotherapy group) intervention 2: Undergo surgery intervention 3: Correlative studies

Protocol documents:
Prot_SAP_000.pdf: 
1 
 
Local Protocol #: UW15114 
 
 
 
 
NCT02720185 
 
TITLE:  A Window of Opportunity Study of Dasatinib in Operable Triple Negative Breast 
Cancers with Nuclear Epidermal Growth Factor Receptor 
 
Coordinating Center: 
University of Wisconsin Carbone Cancer Center  
 
Principal Investigator: 
Kari B. Wisinski, MD 
 
Department of Medicine 
 
Division of Hematology and Medical Oncology 
 
University of Wisconsin Carbone Cancer Center 
1111 Highland Avenue, WIMR 6033 
Madison, WI 53705-2275 
Phone: 608-262-2876 
Fax: 608-265-6905 
kbwisinski@medicine.wisc.edu 
 
Co-Investigators:    
Lee Wilke, MD 
 
Department of Surgery  
 
University of Wisconsin Carbone Cancer Center 
600 Highland Avenue, CSC, H4/722 
Madison, WI 53792 
Phone: 608-265-5852 
Fax: 608-265-2414 
Wilke@surgery.wisc.edu 
 
 
Amye Tevaarwerk, MD 
 
Department of Medicine 
 
Division of Hematology and Medical Oncology 
 
University of Wisconsin Carbone Cancer Center 
1111 Highland Avenue, WIMR 6037 
Madison, WI 53705-2275 
Phone: 608-262-2837 
Fax: 608-265-6905 
at4@medicine.wisc.edu 
 
 
Ruth O’Regan, MD 
 
Department of Medicine 
 
Division of Hematology and Medical Oncology 
 
University of Wisconsin Carbone Cancer Center 
600 Highland Avenue, CSC K4/542 
Madison, WI 53792 
Phone: 608-265-9701 
Fax: 608-265-2414 
roregan@medicine.wisc.edu 


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2 
 
Mark E. Burkard, MD/PhD 
 
Department of Medicine 
 
Division of Hematology and Medical Oncology 
 
University of Wisconsin Carbone Cancer Center 
1111 Highland Avenue, WIMR 6059 
Madison, WI 53705-2275 
Phone: 608-262-2803 
Fax: 608-265-6905 
meburkard@medicine.wisc.edu 
 
 
David Yang, MD 
 
Department of Pathology and Laboratory Medicine  
 
University of Wisconsin Carbone Cancer Center 
600 Highland Avenue, CSC K4/446 
Madison, WI 53792 
Phone: 608-263-5965 
Fax: 608-265-1568 
dyang@pathology.wisc.edu 
 
 
Deric Wheeler, PhD 
 
Department of Human Oncology  
 
University of Wisconsin Carbone Cancer Center 
1111 Highland Avenue, WIMR 3159 
Madison, WI 53705-2275 
Phone: 608-262-7837 
Fax: 608-265-2414 
dlwheeler@humonc.wisc.edu 
 
Statistician: 
 
Program Manager: 
Emmanuel Sampene 
Tammy Koehn 
Department of Biostatics and 
University of Wisconsin Carbone      
Medical Informatics 
Cancer Center  
600 Highland Ave, Box 4675 
600 Highland Ave, K4/6 
Madison, WI 53792-4675 
Madison, WI 53792 
(608)265-8725 
tmkoehn@medicine.wisc.edu  
(608) 265-2414 (Fax) 
 
sampene@biostat.wisc.edu 
 
 
     
 
Participating Sites:  
University of Illinois at Chicago 
 
Funding Sponsor:  
1 R01 CA193004-01A1 
 
Protocol Type / Version # / Version Date: 
[Version 4 / 08/29/2019] 
 
 


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Project Summary/SCHEMA 


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TABLE OF CONTENTS 
 
PROJECT SUMMARY/Project Summary/SCHEMA ....................................................................3 
1. 
OBJECTIVES ......................................................................................................................6 
1.1 
Primary Objectives ...................................................................................................6 
1.2 
Secondary Objectives ...............................................................................................6 
2. 
BACKGROUND .................................................................................................................6 
2.1 
Triple negative breast cancer ...................................................................................6 
2.2 
Epidermal Growth Factor Receptor .........................................................................7 
2.3 
Dasatinib ................................................................................................................13 
2.4 
Rationale ................................................................................................................20 
3. 
PATIENT SELECTION ....................................................................................................21 
3.1 
Eligibility Criteria for nEGFR testing ....................................................................21 
3.2 
Eligibility Criteria for study therapy ......................................................................21 
3.3 
Exclusion Criteria ..................................................................................................22 
3.4 
Inclusion of Women and Minorities ......................................................................23 
4. 
REGISTRATION PROCEDURES ...................................................................................23 
4.1 
Patient Registration ................................................................................................23 
4.2 
General Guidelines .................................................................................................23 
5. 
TREATMENT PLAN ......................................................................................................233 
5.1 
Dasatinib Administration .....................................................................................233 
5.2 
General Concomitant Medication and Supportive Care Guidelines ......................24 
5.3 
Duration of Therapy .............................................................................................244 
5.4 
Duration of Follow Up ...........................................................................................25 
5.5 
Criteria for Removal from Study ...........................................................................25 
6. 
DOSING DELAYS/DOSE MODIFICATIONS................................................................25 
7. 
PHARMACEUTICAL INFORMATION ........................................................................255 
7.1 
Dasatinib ..............................................................................................................255 
8. 
BIOMARKER, CORRELATIVE, AND SPECIAL STUDIES ......................................266 
8.1 
Nuclear EGFR ......................................................................................................266 
8.2 
Exploratory Studies ................................................................................................27 
 
9.         CONFIDENTIALITY AND DATA MANAGEMENT…………………………………27 
10. 
STUDY CALENDAR .......................................................................................................29 
11. 
MEASUREMENT OF EFFECT......................................................................................300 
11.1 
Antitumor Effect – Solid Tumors ........................................................................300 


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12. 
DATA SAFETY AND MONITORING PlAN ................................................................300 
I. 
EXPEDITED REPORTING OF SERIOUS ADVERSE EVENTS .................................333 
13. 
STATISTICAL CONSIDERATIONS.............................................................................366 
REFERENCES ............................................................................................................................388 
APPENDIX A 
PERFORMANCE STATUS CRITERIA ..................................................44 
 
APPENDIX B 
Medication That May Cause QTc Prolongation………………….............45 
 
APPENDIX C 
Medication Diary for Dasatinib…………………………………………..47 
 
 
 


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1. OBJECTIVES 
 
1.1 
Primary Objectives 
 
To determine if dasatinib, an inhibitor of the Src family kinases, can prevent the nuclear 
translocation of the epidermal growth factor receptor (EGFR) in Stage I-III nuclear EGFR 
positive, triple negative breast cancers (TNBC).   
 
1.2 
Secondary Objectives 
 
1.2.1 To examine the safety and tolerability of dasatinib in patients with operable TNBC 
 
1.2.2 To explore potential intracellular mechanisms which impact dasatinib effect on cellular 
localization of EGFR in operable TNBC. 
 
1.2.3 To examine the pathologic complete response (pCR) rates to standard neoadjuvant 
chemotherapy in nEGFR+ TNBC 
 
1.2.4    To examine breast cancer recurrence rates and patterns of metastatic recurrence in 
nEGFR+ TNBC.   
 
 
 
2. BACKGROUND 
 
2.1 
Triple negative breast cancer  
 
Breast cancer remains an important global health issue. One of eight U.S. women is diagnosed 
with breast cancer during her lifetime.1 The American Cancer Society estimates that 234,190 new 
cases of invasive breast cancer will be diagnosed in the United States in 2015 with 40,730 deaths. 
2 2 2 2  Triple-negative breast cancer (TNBC) constitutes approximately15% of breast cancers and 
lacks expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal 
growth factor receptor-2 (HER2). TNBC is associated with African-American and Hispanic race, 
younger age at diagnosis, advanced stage, high mitotic index, and BRCA1 mutations.3-7 Due to the 
absence of any targeted therapies, cytotoxic chemotherapy remains the mainstay of medical 
treatment for TNBC, but outcomes are poor compared with other subtypes.8,9 Median survival for 
women with advanced TNBC remains a dismal 13 months.9 Improved understanding of this 
disease to advance our treatment approaches are urgently required.  
 
For stage I-III TNBC, despite optimal local therapy with breast and axillary surgery with or 
without radiation, the risk for distant early recurrence remains significant.  Thus, systemic 
combination chemotherapy is part of the standard of care for the majority of these patients. 
Neoadjuvant (also known as preoperative) chemotherapy is often recommended in this setting 
based on trials that have shown equivalent long term outcomes.10  Furthermore, neoadjuvant 
chemotherapy has been shown to allow for conversion from mastectomy to breast conservation 
in a portion of patients and potentially reduce need for full axillary dissection.11 In addition, 
studies in TNBC have shown that patients who are able to achieve a pathologic complete 


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response (pCR) are likely to have improved survival over those who have residual disease. 
Patient’s with TNBC who achieve a pCR have similar survival to non-TBNC.12 Neoadjuvant 
therapy is now considered a standard of care for many of these patients.  
 
2.2 
Epidermal Growth Factor Receptor  
 
2.2.1 Overview of Epidermal Growth Factor Receptor (EGFR) in TNBC 
 
EGFR is an oncogene that promotes tumor growth.13 Targeting EGFR has been a successful 
therapeutic strategy for multiple solid tumors.14-17  Approximately 50% of TNBCs over-express 
EGFR, which is associated with poor patient prognosis.18-23 However, clinical trials investigating 
the anti-EGFR antibody cetuximab in TNBC have largely been unsuccessful when used alone, and 
only modestly effective in combination with platinum chemotherapy.24,25 The failure of cetuximab 
in TNBC patients may be due to the fact that cetuximab only targets the classical EGFR signaling 
pathway emanating from plasma membrane-bound EGFR. We hypothesize that the failure of 
cetuximab in TNBC patients may be due to the fact that cetuximab only targets classical EGFR 
signaling that emanates from plasma membrane-bound EGFR and not the nEGFR-signaling 
pathway. This represents a fundamental obstacle in the field of EGFR targeting in TNBC, and 
suggests that nEGFR expressing patients will be resistant to cetuximab therapy.   
 
2.2.2 Nuclear EGFR 
 
Emerging data suggests a new mode of EGFR function in which EGFR undergoes nuclear 
translocation in a series of well-defined steps.26-35 At the plasma membrane the dimerized receptor 
is phosphorylated at Y1101 by SFKs and this represents a critical, early event for movement of 
the EGFR to the nucleus.29 These early events lead to receptor internalization to the early 
endosome followed by retrograde trafficking to the Golgi via microtubule-dependent movement 
by interacting with dynein and fusion with the Golgi apparatus through syntaxin 6-mediated 
membrane fusion34. This is followed by COPI-mediated retrograde trafficking from the Golgi to 
the ER where the nuclear localization sequence (NLS)36 of the EGFR interacts with importinβ1.30 
Here the EGFR/importinβ1 complex moves to the outer nuclear membrane (ONM) where 
importinβ1 interacts with nup358 of the nuclear pore complex (NPC) and is shuttled to the inner 
nuclear membrane (INM). Here, EGFR interacts with the Sec61 translocon for nuclear import.32 
Within the nucleus, two functions have been identified for EGFR. Firstly, EGFR serves as a co-
transcription factor regulating genes involved in tumor progression (Cyclin D1, iNOS, B-myb, 
Myc, BCRP, Aurora Kinase A and COX2).37-43 Secondly, nEGFR serves as a kinase that 
phosphorylates proliferating cell nuclear antigen (PCNA) increasing its stability and function in 
the proliferating cell44 and activates DNA-PK within the nucleus to enhance DNA repair.45 These 
identified nuclear functions have been linked with: 1) inverse correlation with overall survival in 
breast, ovarian, oropharyngeal, gallbladder and lung cancer46-51, 2) enhanced tumor growth52-54 
and 3) resistance to therapeutic agents including radiation, gefitinib, cisplatin and 
cetuximab.41,52,55,56 Although there are several reported mechanisms of resistance to cetuximab 
including altered angiogenesis57-59, increased EGFR activity60, increased expression of HER 
family ligands52,61 increased EGFR degradation60,62, oncogenic shift60, epithelial to mesenchymal 
shift63, and PTEN degradation,64 findings from Wheeler et al. suggest that targeting nEGFR can 
sensitize nEGFR positive TNBC cells to cetuximab therapy.29,52,60,65-67 Unfortunately, nEGFR 


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cannot be targeted by cetuximab and since EGFR regulation of genes is kinase-independent, 
gefitinib and erlotinib, small molecule inhibitors of the EGFR, are of little utility.68 
 
 
 
 2.2.3 Preliminary Data  
 
Nuclear EGFR plays a role in cetuximab resistance 
Cetuximab-resistant tumor cell lines were established by exposing the cetuximab sensitive human 
NSCLC line NCI-H226 (H226) and the HNSCC line UMSCC-1 (SCC1) to increasing doses of 
cetuximab for a time period of six-months until single cell resistant clones emerged.60,69 This 
process resulted in six stable resistant clones for the H226 NSCLC line designated HC1, HC4, 
HC5, HC6, HC7 and HC8. The sensitive parental line was designated HP. For the SCC1 HNSCC 
line, six stable resistant clones were generated (SC1, SC2, SC5, SC6, SC7, SC8), and the sensitive 
parental line was designated SP. All clones displayed a robust cetuximab-resistant phenotype when 
challenged with increasing concentrations of cetuximab as 
compared to parental controls. Sequence analysis of the 
EGFR in selected resistant clones indicated no mutations 
developed during the selection process in either the resistant 
or parental cells.60 
 
Cetuximab-resistant clones have increased steady-state 
expression and dependency on the EGFR 
After successful establishment of cetuximab-resistant clones 
in two different model systems, activity of 42 RTKs were 
screened using an antibody-based array in selected clones.60 
Analysis of selected cetuximab-resistant clones (HC1, HC4 
and HC8, SC1 and SC2) indicated that the EGFR steady-state 
expression was dramatically increased, secondary to 
decreased ubiquitination60) as well as its overall activity as 
indicated by phosphorylation of several key tyrosines on the 
EGFR (Figure 1A). Further, experiments using siRNAs 
directed against the EGFR indicated that cetuximab-resistant 
clones remained dependent on the EGFR for proliferation 
(Figure 1B&C). Collectively this data suggested that 
cetuximab-resistant clones upregulated the steady-state 
expression of the EGFR and maintained dependence on the 
EGFR.60 
 
 
 
 
Figure 1: Cetuximab-resistant clones 
have 
increased 
expression 
and 
dependency of EGFR. A) Western blot 
analysis of the EGFR. B&C) siRNAs 
directed against the EGFR decreases 
proliferation 
of 
cetuximab 
resistant 
clones. * P<0.005 


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Cetuximab-resistant clones have increased 
nEGFR 
To investigate why cetuximab-resistant clones 
maintain dependency on the EGFR, yet are 
resistant 
to 
cetuximab, 
we 
performed 
biochemical 
fractionation 
and 
automated 
quantitative IHC analysis (AQUA, data not 
shown) and found that cetuximab-resistant 
clones exhibit increased nEGFR expression 
relative to cetuximab-sensitive parental controls 
(HP and SP, respectively) (Figure 2A). To 
determine if this increased nEGFR resulted in 
upregulation of known nEGFR gene targets we 
analyzed expression of Cyclin D1, B-myb, iNOS 
and Aurora Kinase A by quantitative PCR 
(qPCR). These results, relative to the cetuximab-
sensitive 
parental 
controls 
(HP 
and 
SP, 
respectively), 
showed 
robust 
increased 
expression (Figure 2B). Collectively these findings 
indicate that cetuximab-resistant clones, in two 
independent model systems with multiple clones, have 
increased nEGFR and upregulation of known nEGFR 
target genes.  
Nuclear EGFR plays a role in resistance to cetuximab 
therapy 
 
To isolate the effects of nEGFR and its role in 
resistance, we designed a retrovirus expressing wild-
type human EGFR fused to a SV40 nuclear localization 
sequence tag (NLS) (EGFR-NLS/Myc52, Figure 3A). 
We infected the cetuximab-sensitive parental lines, HP 
and SP, with a retrovirus containing either the WT-
EGFR or EGFR-NLS vectors. From these experiments 
we obtained one WT-EGFR and two EGFR-NLS-
expressing clones (EGFR-NLS1 and EGFR-NLS2) for 
the LSCC model (Figure 3B) and one WT-EGFR and 
one EGFR-NLS expressing clones (EGFR-NLS1) for 
the HNSCC model (Figure 3E) that were highly 
localized to the nucleus. To determine if increased 
nEGFR had functionality we measured the expression 
of four known EGFR target genes, cyclin D1, COX2, 
iNOS and B-myb: All four genes had greater than six-
fold increase in expression (data not shown).52 
 
Figure 
2: 
Cetuximab-resistant 
clones 
have 
increased nEGFR. A) Western blot analysis of EGFR 
from cytoplasmic (NN) and nuclear (nuc) cellular 
fractions. B) Quantitative PCR (qPCR) for known 
nEGFR target genes.  
Figure 3: nEGFR leads to resistance to 
cetuximab. A) Schematics for viral vectors 
of EGFRWT and EGFR fused to NLS 
(EGFR-NLS). B&E) Nuclear (Nuc) fraction 
showing increased expression of EGFR in 
the nucleus in the EGFR-NLS clones. C&F) 
Proliferation assays showing EGFR-NLS 
clones were more resistant to cetuximab. D) 
EGFR-NLS 
clones 
are 
resistant 
to 
cetuximab in vivo. * P<0.005 


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Next, we used these model systems to test the hypothesis that directing EGFR to the nucleus would 
result in increased resistance to cetuximab therapy. The results indicated that, relative to the 
wildtype control, each clone exhibited an approximate 50—75% increase in resistance to 
cetuximab treatment in vitro (Figure 3C&F)52.  These results were expanded for the lung system 
in xenografts models challenged with cetuximab for the time indicated (Figure 3D).52 Further, 
analysis of established NSCLC, HSNCC and TNBC cell lines, exhibited a correlation between 
nEGFR levels and intrinsic resistance to cetuximab (data not shown).52,65 Collectively, these data 
support a role for nEGFR in cetuximab resistance.  
 
Cetuximab-resistant clones have increased SFK activity 
To further investigate activated pathways in cetuximab-resistant models we performed proteomic 
analysis measuring activity, by phosphorylation, using intracellular phospho-arrays (data not 
shown). Results of this analysis indicated that all cetuximab-resistant clones, in both models of 
acquired resistance, had marked increased phosphorylation of SFKs on Y419, a critical tyrosine 
site in the activation loop of the kinase (data not shown).66,67 Further, SFK kinase assays 
demonstrated robust activation in resistant clones tested (data not shown).67 Collectively these 
results suggest that cetuximab-resistant clones have increased SFK activity relative to cetuximab 
sensitive parental controls.  
 
SFKs regulate EGFR translocation to the nucleus by phosphorylation of tyrosine 1101 
 
Our 
findings 
in 
acquired 
resistance 
to 
cetuximab52,67 indicated that cetuximab-resistant 
cells have 1) increased nEGFR52,66 and 2) 
increased SFK activity.66,67 To understand if 
SFKs played a direct role in nuclear translocation 
of the EGFR we analyzed known SFK 
phosphorylation sites on the EGFR (Y845 and 
Y1101).70,71 This analysis showed increased 
phosphorylation of Y845 and Y1101 in 
cetuximab-resistant clones (Figure 1A). To 
determine whether phosphorylation of one or 
both of these tyrosines were involved in EGFR 
nuclear translocation, the MCF-7 cell line was 
transfected with wild-type (WT) EGFR or the 
following EGFR mutants: EGFR-Y845F or 
EGFR-Y1101F (Figure 4A). To induce nuclear 
translocation of the EGFR we treated the transfected cells with EGF for 45 min before nuclear 
fractionation. The results of this experiment indicated that EGF induced nuclear translocation of 
EGFR in both EGFR-WT and EGFR-Y845F mutant cells (Figure 4B). However, EGF- induced 
EGFR nuclear translocation was diminished ~70% in EGFR-Y1101F mutant cells as compared to 
EGFR-WT expressing cells.29 Collectively, these data suggest that the phosphorylation of Y1101 
is important for the nuclear translocation of EGFR and suggests that SFK blockade may represent 
a unique opportunity to block nEGFR translocation and thus function.  
 
 
 
Figure 4: Tyrosine 1101 of the EGFR plays a 
critical role in EGFR nuclear translocation. A) 
Western blot analysis of cytosolic lysate showing 
equal expression of WT, Y845F and Y1101 mutant 
stable clones. B) Western blot analysis of nuclear 
lysate showing Y1101F is defective in nuclear 
translocation of the EGFR as compared to WT and 
Y845F.  


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TNBC cell lines and human tumors express 
nuclear localized EGFR 
 
Six 
established 
(BT549, 
MBAMD231, 
SUM149, 
SUM159, 
SUM229 
and 
MDAMB468) 
TNBC 
cell 
lines 
were 
evaluated for EGFR and SFK expression 
(Figure 5A). Further, all TNBC cell lines 
expressing EGFR also expressed variant 
levels of nEGFR (Figure 5B). Immunogold 
labeling of the EGFR and subsequent 
transmission 
electron 
microscopy 
(EM) 
validated nEGFR expression (Figure 5C). 
Given that nEGFR was expressed in 
established TNBC cell lines, we probed a 
human tissue microarray (TMA) containing 
74 TNBC patient tumors for EGFR expression 
and localization. Analysis of tumors stained 
for EGFR via immunohistochemistry (IHC) 
indicated that 20% of the tumors expressed nEGFR (Figure 5D). Together, these results indicated 
that established human TNBC lines and TNBC patients express nEGFR.  
 
TNBC cells are resistant to cetuximab therapy, but 
dependent on EGFR for proliferation  
 
To determine the role of EGFR in TNBC 
proliferation, 
genetic 
ablation 
studies 
were 
performed in six TNBC cell lines using an EGFR 
directed siRNA pool: SUM159, MDAMB231, 
SUM229, SUM149, SUM102 and MDAMB468 
(depicted) (Figure 6A and B). Loss of EGFR 
expression led to a 23-50% reduction in cell 
proliferation as compared to cells treated with vehicle 
or non-targeting (NT) siRNA  (Figure 6). Each cell 
line challenged with increasing doses of cetuximab 
(from 0.01nM to 100nM) demonstrated only minor 
reductions in proliferation indicating cetuximab 
resistance (Figure 6C). SUM102 was sensitive to 
cetuximab therapy (data not shown).  These results 
indicate that TNBC cell lines depend on EGFR for 
proliferation but are resistant to cetuximab. 
       
Figure 6: TNBC cell lines are dependent on 
EGFR for proliferation, but are intrinsically 
resistant to cetuximab. A and B) Six TNBC cell 
lines (MDAMB468 depicted) were incubated with 
siEGFR, non-targeting (NT) siRNA, or vehicle for 
72-96 hr prior to performing proliferation assays. C) 
Cells were treated with cetuximab at indicated doses 
for the same time course. Proliferation is plotted as 
a percentage of growth relative to vehicle treated 
cells (n=3). Whole cell lysate was harvested from 
all cell lines at the same time point to confirm 
knockdown of EGFR. Data points are represented 
as mean±s.e.m. **p<0.01.  
Figure 5: TNBC cell lines and human tumors express 
nuclear localized EGFR. A) TNBC cells express EGFR 
and SFKs. B and C) TNBC cells express nEGFR as 
indicated by nuclear fractionation (B) and immunogold 
labeling and EM (C). D) 20% of Human TNBC tumors 
express nEGFR. IHC staining for EGFR was performed on 
a total of 74 TNBC patient tumor sections.  


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Nuclear EGFR trafficking is abrogated by SFK inhibition leading 
to increased EGFR membrane expression  
Since Y1101 phosphorylation by SFKs is necessary for EGFR 
nuclear translocation (Figure 7A) we utilized the SFK inhibitor 
dasatinib to determine if it could abrogate EGFR translocation from 
the membrane to nucleus. Treatment of TNBC cells SUM149, 
SUM229 and MDAMB468 with dasatinib led to potent decreases in 
nEGFR expression (see arrow) and robust increases in non-nuclear 
EGFR levels (see arrow) at 72 hours post treatment (SUM 148 and 
MDAMB468 depicted, Figure 7B). Additionally, significant 
increases in cell surface EGFR levels were detected by flow 
cytometry 72 hours post treatment. These results were also verified 
using genetic approaches with SFK mutants and a negative regulator 
of Src (known as the SLAP protein) to ensure these findings were 
not off target effects of dasatinib.  Increase in membrane EGFR 
was also seen after dasatinib treated mice with MDAM8468 
xenografts (Figure 7C).  These results indicate that SFK blockade 
decreases nEGFR and increases 
plasma membrane EGFR levels.  
 
 
Cetuximab resistant TNBC cell 
lines are sensitized to Cetuximab 
resistant TNBC cell lines are 
sensitive 
upon 
inhibition 
of 
nEGFR translocation 
 
Since SFK inhibition of nEGFR 
increased plasma membrane EGFR levels we 
hypothesized that cells would now be more sensitive 
to cetuximab. To investigate this possibility, we pre-
treated 
TNBC 
cells 
(SUM129, 
SUM149, 
MBAMB231, MDAMB468 depicted) with dasatinib 
or vehicle for 24 hours, the time point where 
increased surface level EGFR was detected, and 
subsequently treated cells with increasing doses of 
cetuximab therapy for an additional 72 hours (Figure 
8). Analysis of cellular proliferation post treatment 
indicated that TNBC cell lines that received dasatinib 
for 24 hours prior to cetuximab treatment 
demonstrated significant reductions in proliferation 
over a wide range of cetuximab doses. Collectively, 
these data suggest that the blockade of nEGFR 
translocation via SFK inhibition can increase TNBC 
cell sensitivity to cetuximab.  
 
 
Figure 7:  Therapeutic inhibition of SFKs blocks nEGFR 
translocation in TNBC cell lines A)Western blot analysis of 
cytoplasmic and nuclear fractions demonstrates that mutation of 
Y1101 prevents nEGFR translocation. B) Dasatinib (25nM) inhibits 
nEGFR translocation (red arrows) and increases  nonnuclear EGFR 
levels (green arrows). SFK blockade via dasatinib increases EGFR 
surface levels measured by flow cytometry. C) Tumors harvested from 
mice treated with dasatinib have decreased nEGFR and increased 
plasma membrane EGFR levels. 
Figure 8. Therapeutic inhibition of SFK 
activity 
can 
sensitize 
TNBC 
cells 
to 
cetuximab19. 
Six 
TNBC 
cell 
lines 
(MDAMB468 depicted) were pretreated with 
vehicle or dasatinib (25 nM) for 24 hr prior to 
adding cetuximab to the growth medium at the 
indicated doses for an additional 72 hr. 
Proliferation assays were performed and plotted 
as a percentage of growth relative to vehicle 
treated cells (n=3). Data points are represented 
as mean±s.e.m. **p<0.01.  


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2.3 
Dasatinib  
 
Dasatinib is an oral tyrosine kinase inhibitor that targets BCR/ABL, Src, c-Kit, ephrin and other 
receptors.72 It was first approved in 2006 for chronic phase-chronic myelogenous leukemia (CML) 
who are intolerant or resistant to prior therapy including imatinib.73 It was also approved in 2010 
for treatment of newly diagnosed patients with Philadelphia chromosome + CML during the 
chronic phase.74 Since then, dasatinib approval has now been extended to use in all phases of CML 
in patient who are intolerant of imatinib and Philadelphia chromosome +  acute lymphoblastic 
leukemia (ALL) with resistance or intolerance to prior therapy.75  
 
At nanomolar concentrations, dasatinib is shown to inhibit BCR/ABL, SRC family (Src, Lck, Yes, 
Fyn), c-KIT, EPHA2, and PDGFRβ.  In vitro, dasatinib was found to be active in leukemic cell 
lines overexpressing BCR/ABL that were sensitive and resistant to imatinib. In addition, dasatinib 
was found to overcome BCR/ABL kinase domain mutations leading to imatinib resistance by 
activating alternate signaling pathways involving SRC family kinases and multi-drug resistance 
gene expression. 
 
Pharmacokinetics 
 
Maximum plasma concentrations of dasatinib were observed between 0.5-6hours after oral 
administration. There is a dose proportional increase in AUC and linear elimination characteristics 
between 15mg per day to 240mg per day. Overall terminal half-life of 3 to 5 hours.  Volume of 
distribution if 2.5L and is extensively distributed in the extravascular space. Binding of the drug 
and its active metabolite to human plasma proteins in vitro was approximately 96 and 93%, 
respectively.  
 
Metabolism:  Dasatinib is extensively metabolized in humans by the CYP3A4 enzyme (a 
cytochrome P450 enzyme) leading to its active metabolite. Flavin-containing monooxygenase 3 
and uridine diphosphate-glucoronosyltransferase enzymes are also involved in the formation of 
dasatinib metabolites. The active metabolite of dasatinib, which is equipotent to dasatinib, is about 
5% of the AUC, indicating that the active metabolite does not play a significant in the 
pharmacology of the drug.Dasatinib is a weak inhibitor of CYP3A4 enzyme. It does not inhibit 
CYP1A2, 2A6, 2C8, 2C9, 2C19, 2D6, or 2E1 at clinically relevant concentrations. It does not 
induce the CYP enzymes in humans. 
 
Elimination:  After a single oral dose of [14C]-labeled dasatinib, 4% was recovered in the urine and 
85% recovered in the feces within 10 days. Unchanged dasatinib was found in urine at a rate of 
0.1% and 19% in feces. 
 
Effects of age and gender:  There are no clinically relevant effects of age and gender on the 
pharmacokinetics of dasatinib. 
 


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Hepatic impairment:  Differences in Cmax and AUC are not clinically relevant and dose 
adjustments are not needed in hepatic impairment.  Dasatinib administered in patients with Child 
Pugh B and Child Pugh C at doses of 50 and 20mg, respectively, and compared to patients with 
normal hepatic function at 70mg. Patients with Child Pugh B hepatic impairment had decreases in 
dose-normalized Cmax and AUC by 47% and 8%, respectively. Patients with Child Pugh C hepatic 
impairment had decreases in dose-normalized Cmax and AUB by 43% and 28%, respectively. 
 
Renal impairment:  There are no clinical studies with dasatinib in renal impairment. Less than 4% 
of dasatinib and its metabolites are excreted by the kidney. 
 
Drug interactions 
 
Drugs that may increase dasatinib plasma concentrations include CYP3A4 inhibitors: In a trial 
evaluating pharmacokinetics of dasatinib, it was found that dasatinib 20mg daily with 
ketoconazole 200mg twice daily (a potent CYP3A4 inhibitor) showed that Cmax and AUC were 
increased by four- and five-fold, respectively. Use of dasatinib in combination with a CYP3A4 
inhibitor may increase dasatinib levels and recommended to be avoided. Patients taking dasatinib 
should be monitored for toxicity and a dose reduction is recommended if CYP3A4 inhibitor cannot 
be discontinued.  Common medications that inhibit P450 system and grapefruit juice may increase 
plasma concentrations of dasatinib and should be avoided. Use of an alternative medication with 
no or minimal enzyme inhibition potential should be considered. If dasatinib must be used in 
conjunction with a CYP3A4 inhibitor, a dose decreased should be considered with 
recommendation to reduce dose 20mg daily if starting at 100mg daily. Reduced doses of dasatinib 
are predicted to adjust for area under the curve to the range observed if no CYP3A4 inhibitor was 
used. However, there is no clinical data to support these dose adjustments in patients.  
 
Drugs that may decrease dasatinib plasma concentrations include CYP3A4 inducers, antacids and 
proton pump inhibitors: Concominant use of a potent CYP3A4 inducer (rifampin) and dasatinib 
showed the mean Cmax and AUC of dasatinib were decreased by 81% and 82%, respectively. If a 
CYP3A4 inducer must be utilized, consider an alternative drug that has less CYP3A4 activity. If 
dasatinib must be used with a CYP3A4 inducer, a dose increase is recommended. Common 
medications that induce P450 system and St. John’s Wort may decrease plasma concentrations of 
dasatinib unpredictably. If patients must use a strong CYP3A4 inducer, pharmacokinetics suggest 
a dose increase. Patient should be monitored for drug toxicity. 
 
Drugs that may have plasma concentration altered by dasatinib:  Mean Cmax and AUC of 
simvastatin were increased by 37% and 20%, respectively, when administered in combination. 
CYP3A4 substrates known to have a narrow therapeutic index should be administered in caution 
while taking dasatinib.  
 
 


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Adverse Events 
 
In the overall population of 2712 dasatinib-treated patients, 88% of patients experienced adverse 
reaction at some time and 19% had adverse reactions that led to treatment discontinuation.72-81 In 
a randomized trial in patients with newly diagnosed chronic phase CML, 16% of patients had to 
discontinue dasatinib due to adverse reactions at 60 month follow up and 39% of patients 
discontinued dasatinib for all reasons.78   The reported frequency of specific adverse events is in 
tables below.  Significant adverse events include:  
 
Myelosuppression: Dasatinib is associated with severe (Grade 3 or 4) thrombocytopenia, 
neutropenia, and anemia. This is generally reversible and managed by holding dasatinib or dose 
reducing, depending on severity. This was commonly reported in all patient populations. 
Severe adverse reaction rate was higher in patients in patients with advanced CML than in chronic 
phase CML.  These occurred earlier and more frequently in patients with advanced CML or Ph+ 
ALL than in chronic phase CML. In patients on dasatinib for CML, it is recommended for CBC 
monitoring for every 1-2 weeks for first 2-3 months and, thereafter, as clinically indicated. 
 
Bleeding related events: Dasatinib is associated with thrombocytopenia and platelet dysfunction 
in vitro. Most bleeding events in studies were associated with severe thrombocytopenia. Co-
administration of medications that inhibit platelet function or anticoagulation may increase risk of 
hemorrhage. In all CML or PH+ ALL studies: ≥ Grade 3 CNS hemorrhages, including fatalities, 
occurred in <1% of patients; ≥ Grade 3 gastrointestinal hemorrhage, including fatalities, occurred 
in 4% of patients and required treatment interruptions and transfusions; 
≥ Grade 3 other hemorrhages occurred in 2% of patients. 
 
Fluid retention: Dasatinib may cause fluid retention. Evaluation of patients who develop 
symptoms of pleural effusion or other fluid retention, such as new or worsened dyspnea, pleuritic 
chest pain, or dry cough with chest X-ray or additional diagnostic imaging as clinically indicated. 
Management includes supportive care measures as diuretics or steroids. Severe pleural effusions 
are managed by thoracentesis and oxygen. Dasatinib was either dose reduced or held depending 
on severity of adverse reaction.  At 5 years of follow-up with chronic dasatinib use, grade 3 or 4 
fluid retention was reported in 5-8% of CML and ALL pts. 
 
Cardiovascular events: Dasatinib has been associated with cardiovascular events.  At 5 years 
follow-up, patients on continuous dasatinib for chronic phase CML reported cardiac ischemic 
events in 3.9%, cardiac related fluid retention in 8.5%, arrhythmias or palpitations in 7.0% and 
Transient ischemic attacks in 0.8%. 
 
Pulmonary arterial hypertension: Dasatinib may be associated with an increased risk of 
developing pulmonary arterial hypertension, occurring at any time after initiation, including after 
more than 1 year of treatment. Symptoms included dyspnea, fatigue, hypoxia, fluid retention. 


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These were found to be reversible after discontinuation of dasatinib and recommend to be 
permanently discontinued 
 
QT prolongation: Based on in vitro studies, dasatinib potentially prolonged cardiac ventricular 
repolarization. This was more evident in patients with hypokalemia, hypomagnesemia, congenital 
long QT syndrome, patient on anti-arrhythmic medicine or other medications that also prolong 
QT, and cumulative high-dose anthracycline therapy. In a study of 2440 patients at all doses of 
dasatinib, <1% of patients reported QT prolongation with 1% experiences a QT prolongation of 
>500ms. Maximum changes in QTc from baseline ranged from 7.0 to 13.4ms 
 
Severe dermatological reactions: Cases of severe mucocutaneous dermatologic reaction have been 
reported, including Stevens-Johnson syndrome and erythema multiforme. 
 
Embryo-fetal toxicity: Limited human data suggests that dasatinib may cause fetal harm with 
patient is pregnancy. Resulting complication include hydrops fetalis, fetal leukopenia, and fetal 
thrombocytopenia. Recommend that females of reproductive potential to avoid pregnancy and 
recommend the use of contraception while taking dasatinib and for 30 days after final dose. 
 
Table 1. Non-laboratory adverse reactions in ≥10% of patients (or those of special interest) in a 
randomized trial of dasatinib vs imatinib in patients with newly diagnosed chronic phase CML 
with minimum follow-up of 60 months (N=258).72 Dasatinib dose of 100mg daily 
Adverse reaction 
All grades (%) 
Grade 3 or 4 (%)  
Fluid retention 
38 
5 
Pleural effusion 
28 
3 
Superficial local edema 
14 
0 
Pulmonary hypertension 
5 
1 
Generalized edema 
4 
0 
Pericardial effusion 
4 
1 
CHF/cardiac dysfunction* 
2 
<1 
Pulmonary edema 
1 
0 
Diarrhea 
22 
1 
Musculoskeletal pain 
14 
0 
Rash$ 
14 
0 
Headache 
14 
0 
Abdominal pain 
11 
0 
Fatigue 
11 
<1 
Nausea 
10 
0 
Myalgia 
7 
0 
Arthralgia 
7 
0 
Hemorrhage 
8 
1 
GI bleeding 
2 
1 
CNS bleeding 
<1 
0 
Other bleeding^ 
6 
0 


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Vomiting 
5 
0 
Muscle spasms 
5 
0 
*Cardiac dysfunction includes acute cardiac failure, congestive cardiac failure, cardiomyopathy, 
diastolic dysfunction, decreased EF, and LV dysfunction 
$Rash includes erythema, erythema multiforme, rash, generalized rash, macular rash, popular 
rash, pustular rash, skin exfoliation, and vasicular rash 
^Other bleeding includes conjunctival hemorrhage, ear hemorrhage, ecchymosis, epistaxis, eye 
hemorrhage, gingival bleeding, hematoma, hematuria, intra-abdominal hematoma, petechiae, 
scleral hemorrhage, uterine hemorrhage, and vaginal hemorrhage 
 
Table 2.  Grade 3/4 laboratory abnormalities in patients with newly diagnosed chronic phase 
CML (N=258) with minimum 60 months follow-up.72 Dasatinib dose of 100mg daily 
 
Laboratory Abnormality  
 
Percentage (%) 
of Patients  
Hematologic  
 
Neutropenia  
29 
Thrombocytopenia 
22 
Anemia 
13 
Non-Hematologic  
 
Hypophosphatemia 
7 
Hypokalemia 
0 
Elevated SGPT (ALT) 
4 
Elevated SGOT (AST) 
<1 
Elevated Bilirubin 
<1 
Elevated Creatinine  
1 
 
At 60 months of continuous dasatinib use, there were 26 deaths in dasatinib-treated patients 
(10.1%). 
 
Table 3.  Additional adverse reactions from pooled data from clinical trials72 
Adverse 
reaction 
≥10% 
1%-10% 
0.1%-<1% 
<0.1% 
GI disorders 
 
Mucosal 
inflammation, 
dyspepsia, 
abdominal 
distension, 
constipation, 
gastritis, colitis, 
oral soft tissue 
disorder 
Ascites, dysphagia, 
anal fissure, upper 
GI ulcer, esophagitis, 
pancreatitis, GERD 
Protein losing 
gastroenteropathy, 
ileus, acute 
pancreatitis, anal 
fistula 
General 
disorders and 
administration 
Peripheral 
edema, 
face 
Asthenia, chest 
pain, chills 
Malaise, other 
superficial edema 
Gait disturbance 


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site-conditions 
edema 
Skin and 
subcutaneous 
tissue disorders 
 
Alopecia, acne, 
dry skin, 
hyperhidrosis, 
urticarial, 
dermatitis 
Pigmentation 
disorder, skin ulcer, 
bullous conditions, 
photosensitivity, nail 
disorder, 
neutrophilic 
dermatosis, 
panniculitis, palmar-
planta 
erythrodysesthesia, 
hair disorder 
Leukocytoclastic 
vasculitis, skin 
fibrosis 
Respiratory, 
thoracic, and 
mediastinal 
disorders 
 
Lung infiltration, 
pneumonitis, 
cough 
Asthma, 
bronchospasm, 
dysphonia, 
pulmonary arterial 
hypertension 
ARDs, pulmonary 
embolism 
Nervous system 
disorders 
 
Neuropathy, 
dizziness, 
dysgeusia, 
somnolence 
Amnesia, tremor, 
syncope, balance 
disorder 
Convulsion, 
CVA, TIA, optic 
neuritis, VII nerve 
palsy, dementia, 
ataxia 
Blood and 
lymphatic 
system 
disorders 
 
 
Lymphadenopathy, 
lymphopenia 
pure red cell 
aplasia 
Musculoskeletal 
and connective 
tissue disorders 
 
Muscular 
weakness, 
musculoskeletal 
stiffness 
Rhabdomyolysis, 
tendonitis, muscle 
inflammation, 
osteonecrosis, 
arthritis 
 
Investigations 
 
Weight increase, 
weight decrease 
Blood creatine 
phosphokinase 
increased, gamma-
glutamyltransferase 
increased 
 
Infection and 
infestations 
 
Pneumonia, 
URI/inflammation, 
herpes virus 
infection, 
entercolitis 
infection, sepsis 
 
 
Metabolism and 
nutrition 
disorders 
 
Appetite 
disturbances, 
hyperuricemia 
Hypoalbuminemia, 
tumor lysis 
syndrome, 
dehydration, 
Diabetes mellitus 


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hypercholesterolemia 
Cardiac 
disorders 
 
Arrhythmia, 
palpitations 
Angina pectoris, 
cardiomegaly, 
pericarditism 
ventricular 
arrhythmia, T-wave 
abnormality, 
troponin increase 
Cor pulmonale, 
myocarditis, acute 
coronary 
syndrome, cardiac 
arrest, PR 
prolongation, 
coronary artery 
disease, 
pleuropericarditis 
Eye disorders 
 
Visual disorder 
(visual 
disturbance, 
blurred vision, 
reduced visual 
acuity), dry eye 
Conjunctivitis, visual 
impairment 
photophobia, 
increased lacrimation 
 
Vascular 
disorders 
 
Flushing, 
hypertension 
Hypotension, 
thrombophlebitism 
thrombosis 
Livedo reticularis, 
DVT, embolism 
Psychiatric 
disorders 
 
Insomnia, 
depression 
Anxiety, labile 
affect, confusion, 
decreased libido 
 
Pregnancy, 
puerperium, and 
perinatal 
conditions 
 
 
 
Abortion 
Reproductive 
system and 
breast disorders 
 
 
Gynecomastia, 
menstrual disorder 
 
Injury, poising, 
and prodecureal 
complications 
 
Contusion 
 
 
Ear and 
labyrinth 
disorders 
 
Tinnitus 
Vertigo, hearing loss 
 
Hepatobiliary 
disorders 
 
 
Cholestasis, 
cholecystitis, 
hepatitis 
 
Renal and 
urinary 
disorders 
 
 
Urinary frequency, 
renal failure, 
proteinuria 
Renal impairment 
Immune system 
disorders 
 
 
Hypersensitivity 
 
Endocrine 
disorders 
 
 
Hypothyroidism 
Hyperthyroidism, 
thyroiditis 


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In a 2011 phase II study, 44 patients with triple 
negative breast cancer (locally advanced or 
metastatic) were treated with single agent dasatinib. 
Results showed that initial dosing of 100mg BID was 
poorly tolerated and required dose reduction to 70mg 
BID, which was better tolerated. While single agent 
dasatinib had limited activity (objective response rate 
was 4.7%), dasatinib was tolerated at 70mg BID. 
While no grade 4 adverse events were reported, there 
were grade 3 adverse reactions related to fatigue, 
diarrhea, pleural effusions, and dyspnea, occurring in 
>5% of patients. Mean duration of drug use was 1.6 
months, limited by development of adverse 
reactions.82   
 
2.4 
Rationale 
 
Figure 9 demonstrates our working hypothesis for 
cetuximab resistance in TNBC.  The goal of this 
window clinical trial is to evaluate in women with 
TNBC that harbors nuclear EGFR whether dasatinib 
can be used to re-establish EGFR on the plasma 
membrane.   
 
A window of opportunity design allows this important 
research question to be asked in a treatment- naïve 
population of patients as exposure to prior therapies 
can impact mechanisms of resistance in tumors. If 
successful, this will provide proof-of-principle for a 
clinical trial of the combination of dasatinib and 
cetuxumab in nEGFR positive TNBC.  Notably, the 
combination has already been demonstrated to be safe in phase I studies83, so rapid translation of 
this data to the next step clinical trial is feasible.    
 
Since many patients with Stage I-III TNBC are treated upfront with neoadjuvant chemotherapy 
rather than primary breast surgery, this study will allow for the window of dasatinib to be given 
prior to either of these modalities.  Potential interactions with  with neoadjuvant chemotherapy 
by dasatinib will be avoided with a minimum 2 day washout period between dasatinib use and 
start of neoadjuvant therapy. This window of time to chemotherapy initiation is often needed in 
standard practice to obtain pre-chemotherapy testing and insurance approvals.  A research biopsy 
will be obtained in these patients at the end of dasatinib to meet the primary endpoint of this 
study. 
 
Figure 9: Dual targeting of both nEGFR and 
plasma membrane-bound EGFR is necessary 
for the complete inhibition of EGFRs oncogenic 
functions in TNBC19: A working hypothesis: SFK 
phosphorylation of Y1101 of the EGFR is a crucial, 
early event, in nuclear translocation of the 
EGFR18,20. We have previously shown that TNBC 
tumors resistant to cetuximab have dependence on 
the nEGFR pathway (Figure 9A)19. Blockade of 
EGFR translocation to the nucleus leads to 
increased expression on the membrane of the tumor 
cell where it is available for inhibition by 
cetuximab (Figure 9B). 


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3. PATIENT SELECTION 
 
3.1 
Eligibility Criteria for nEGFR testing 
 
3.1.1 Patients must have histologically or cytologically confirmed Stage I- III triple negative 
breast cancer.   
3.1.1.1   ER and PR must be <10% by standard assay methods 
3.1.1.2   HER2 must be either 0, 1+ by immunohistochemistry (if 2+, in situ 
hybridization method used to define HER2) OR have HER2: 17 centromere 
signal of <2.0 using a standard in situ hybridization method.  
 
3.1.2 No prior therapy for current breast cancer 
 
3.1.3 Meet criteria for neoadjuvant chemotherapy or primary breast surgery, as determined by 
primary oncologist and surgeon.   
 
3.1.4 Women age ≥18 years.   
 
3.2 
Eligibility Criteria for study therapy 
 
3.2.1 nEGFR positive 
 
3.2.2 ECOG performance status ≤1 (see Appendix A). 
 
3.2.3 Patients must have normal organ and marrow function as defined below: 
 
− 
leukocytes 
≥3,000/mcL 
− 
absolute neutrophil count 
≥1,500/mcL 
− 
platelets 
≥150,000/mcL 
− 
total bilirubin 
<1.25x institutional upper limit of normal 
− 
AST(SGOT)/ALT(SGPT) 
≤2.5 × institutional upper limit of normal 
 
 
− 
creatinine 
within normal institutional limits 
OR 
− 
creatinine clearance 
≥60 mL/min/1.73 m2 for patients with creatinine levels 
above institutional normal. 
 
3.2.4 Women of child-bearing potential and men must agree to use adequate contraception 
(hormonal or barrier method of birth control; abstinence) prior to study entry and for the 
duration of study participation and for 30 days after the final dose.  Should a woman 
become pregnant or suspect she is pregnant while she or her partner is participating in 
this study, she should inform her treating physician immediately.   
 
3.2.5 Ability to understand and the willingness to sign a written informed consent document. 
 


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3.3 
Exclusion Criteria 
 
3.3.1 Patients who are receiving any other investigational agents. 
 
3.3.2 Patients not able to swallow oral medications or with gastrointestinal conditions that may 
impact absorption of dasatinib.   
 
3.3.3 History of allergic reactions attributed to compounds of similar chemical or biologic 
composition to dasatinib.   
 
3.3.4 Patients receiving any medications or substances that are moderate or strong inhibitors or 
inducers of CYP3A4 are ineligible.  Because the lists of these agents are constantly 
changing, medications should be reviewed by the UW Pharmacy Research Center for any 
contraindicated medications.  As part of the enrollment/informed consent procedures, the 
patient will be counseled on the risk of interactions with other agents, and what to do if 
new medications need to be prescribed or if the patient is considering a new over-the-
counter medicine or herbal product.  
 
3.3.5 H2 antagonists and proton pump inhibitors are not allowed  
 
3.3.6 Anticoagulants (ie. Coumadin, heparin, anti-Xa inhibitors) and anti-platelet agents (ie. 
aspirin) are not allowed.  NSAIDS and acetaminophen are allowed on study. 
 
3.3.7 Medications known to prolong QTC are not allowed (See Appendix B)   
 
3.3.8 No history of prolonged QTC or cardiomyopathy unless normal QTC and ejection 
fraction confirmed within 1 month prior to study entry.    
 
3.3.9 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. 
 
3.3.10 Pregnant women are excluded from this study because dasatinib is a pregnancy category 
D agent with the potential for teratogenic or abortifacient effects.  Because there is an 
unknown but potential risk for adverse events in nursing infants secondary to treatment of 
the mother with dasatinib, breastfeeding should be discontinued if the mother is treated 
with dasatinib and not resumed until at least 2 weeks after the final dose.   
 
3.3.11 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. 
 
3.3.12  Contraindication to repeat breast biopsy (neoadjuvant chemotherapy group) 
 


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3.4 
Inclusion of Women and Minorities 
 
Women and Minorities are not excluded from participation in this trial.  
 
 
4. REGISTRATION PROCEDURES 
 
4.1 
Patient Registration 
 
Eligible patients will be entered on study using the University of Wisconsin Carbone Cancer Center 
OnCore Database. 
 
To register a patient, the following documents should be completed by the study coordinator or data 
manager and maintained in the subject study chart:  
• Copy of required eligibility tests.  
• Signed patient consent form.  
• Signed HIPAA authorization form. 
 
The research coordinator at the site will then register the subject into the OnCore database prior to 
starting study treatment. OnCore will assign the unique subject number.  
 
There will not be access to the OnCore database registration until documented IRB approval is 
obtained and entered into the OnCore database. 
 
 
4.2 
General Guidelines 
 
Following registration, patients should begin protocol treatment within 30 days.  Issues that 
would cause treatment delays should be discussed with the Principal Investigator.  If a patient 
does not receive protocol therapy following registration, the patient’s registration on the study 
may be canceled.   
 
 
5. TREATMENT PLAN 
 
5.1 
Dasatinib Administration 
 
Treatment will be administered on an outpatient basis.  No investigational or commercial agents 
or therapies other than those described below may be administered with the intent to treat the 
patient's malignancy. 
 
Regimen Description 
Agent 
Premedications; 
Precautions 
Dose 
Route 
Schedule 
Cycle 
Length* 
Dasatinib 
None 
 
100mg 
Oral 
Once daily 7-10 days 
 
 


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*Dose should be continued up to the day prior to planned surgery or research biopsy 
(neoadjuvant chemotherapy group)   
 
Dasatinib should be taken at the same time daily (+/- 4 hours).  It may be taken with or without 
food, in the morning or evening.  If a dose is missed, it should not be made up.  Do not crush or 
cut tablets.   
 
Patients should be informed that the tablet contains lactose, in the event they are lactose 
intolerant 
 
The patient will be requested to maintain a medication diary of each dose of medication 
(Appendix C).  The medication diary will be returned to clinic staff at the end of the therapy. 
 
5.2 
General Concomitant Medication and Supportive Care Guidelines 
 
Because there is a potential for interaction of dasatinib with other concomitantly administered 
drugs, the case report form must capture the concurrent use of all other drugs, over-the-counter 
medications, or alternative therapies.  The Principal Investigator should be alerted if the patient is 
taking any agent known to affect or with the potential for drug interactions. 
 
Drug interactions most commonly associated in the absorption and metabolism of dasatinib are 
ones that affect the CYP3A4, both inducers and inhibitors, antacids, and H2 antagonists along 
with proton-pump inhibitors.  
 
CYP3A4:  Moderate or Strong CYP3A4 inducer or inhibitors are not allowed.  CYP3A4 
substrates may be administered, but caution and close monitoring should occur if the substrate 
has a narrow therapeutic index.   
 
Proton pump inhibitors and H2 antagonists are not allowed.  If antacid therapy is needed, the 
antacid dose should be administered at least 2 hours prior to or after the dose of dasatanib.   
 
Anticoagulants and anti-platelet agents are not allowed.  NSAIDS and acetaminophen are 
allowed on study.  However, NSAIDS should be discontinued as per standard of care for pre-
surgical management.     
 
5.3 
Duration of Therapy 
 
Treatment may continue for a maximum of 10 days or until one of the following criteria applies: 
 
• 
Disease progression, 
 
• 
Intercurrent illness that prevents further administration of treatment, 
 
• 
Unacceptable adverse event(s), 
 


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• 
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. 
 
5.4 
Duration of Follow Up 
 
Patients treated with dasatinib will be followed for adverse events for 30 (+/-14) days after breast 
surgery date (if surgery first) or through completion of cycle 1 of chemotherapy (if neoadjuvant 
chemotherapy given).  Patients removed from study therapy for unacceptable adverse event(s) 
will be followed until resolution or stabilization of the adverse event.  All patients will have 
long-term follow up until 2 years after primary breast surgery.   
 
5.5 
Criteria for Removal from Study 
 
Patients will be removed from study when any of the criteria listed in Section 5.4 applies.  The 
reason for study removal and the date the patient was removed must be documented in the Case 
Report Form. 
 
6. DOSING DELAYS/DOSE MODIFICATIONS 
 
Due to the limited therapeutic intent of this clinical trial, study drug will be discontinued for any 
grade 3 or higher adverse event or any intolerable grade 2 event if the event is attributed at least 
possible or greater to dasatinib.  No dose modifications are allowed on study.  Once study drug is 
discontinued for an adverse event it may not be resumed. 
 
Adverse events (AEs) will be graded using the National Cancer Institute Common Terminology 
Criteria for Adverse Events (CTCAE) (version 4.0).   Attribution of the AE should be also 
recorded as: 
- 
Definite – The AE is clearly related to the study treatment. 
- 
Probable – The AE is likely related to the study treatment. 
- 
Possible – The AE may be related to the study treatment. 
- 
Unlikely – The AE is doubtfully related to the study treatment. 
- 
Unrelated – The AE is clearly NOT related to the study treatment. 
 
Although impact on surgery or chemotherapy is not anticipated, any delays in planned breast 
surgery or initiation of neoadjuvant chemotherapy by >1 week related to the study drug or any 
unanticipated complications from surgery reported by the surgery team will also be recorded and 
reviewed at regular DOWG meetings.     
 
7. PHARMACEUTICAL INFORMATION 
 
7.1 
Dasatinib  
 
Availability:  Dasatinib is a commercial agent which will be purchased for the investigational 
use in this study.   


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 Product description:  Tablet – White to off-white, biconvex, oval, film-coated tablet with 
“BMS” debossed on one side and medication dosage on other side. Available preparations in 
20mg, 50mg, 70mg, 80mg, 100mg, and 140 mg. 
 
Solution preparation:  Not applicable – oral medication available in dose required in study 
 
Storage requirements:  should be stored at 20°C to 25°C (68°F to 77°F); excursions permitted 
between 15°C and 30°C (59°F and 86°F). 
 
Handling and Disposal: Dasatinib is an antineoplastic product. 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. However, if tablets are 
inadvertently crushed or broken, pharmacy and clinical personnel should wear disposable 
chemotherapy gloves. Personnel who are pregnant should avoid exposure to crushed or broken 
tablets. 
 
Stability:  Include the stability of the original dosage form, reconstituted solution, and final 
diluted product, as applicable. 
 
Route of administration:  Take orally, once a day. If a patient misses a dose, they should take 
the next dose at its regular schedule. They should not take multiple doses at once. To be taken 
with or without food, swallowed whole without crushing, and not with grapefruit juice. 
 
Agent Ordering:  Ordering and distribution will be handled by the UW Pharmaceutical Research 
Center 
 
8. BIOMARKER, CORRELATIVE, AND SPECIAL STUDIES 
 
8.1 
Nuclear EGFR  
 
nEGFR determination on the diagnostic biopsy will be performed by immunohistochemistry 
utilizing a protocol developed by the Wheeler lab and interpreted by a pathologist (D. Yang) who 
has been collaborating with the Wheeler lab in evaluating nEGFR for over 5 years.  Briefly, 5 
micron tissue sections from the patient’s diagnostic biopsy with be de-paraffinized and subjected 
to citrated buffer antigen retrieval followed by overnight incubation with EGFR polyclonal 
antibody (sc-03, Santa Cruz Biotechnology Inc., Santa Cruz, CA) at 4C.  Peroxidase conjugated 
secondary antibody is then applied for 1 hour followed by standard DAB staining and hematoxylin 
counterstaining.  Each case will be evaluated for nEGFR expression by counting 300 tumor cells 
at 400x magnification by light microscopy.  Tumor cells with strong nucleolar DAB staining will 
be counted as positive and cases with ≥20% positive cells will be categorized as nEGFR positive.  
To ensure preservation of sufficient biopsy tissue for clinical care, no more than three 5um sections 
will be cut from the tissue block for nEGFR immunohistochemical evaluation. 
 
In patients with nEGFR positive tumors treated with dasatinib who after dasatinib are proceeding 
with neoadjuvant chemotherapy, a research biopsy of the primary breast tumor will be obtained 
after 7-10 days of dasatinib.  This biopsy will be palpation or ultrasound guided and include 2-4 


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27 
cores using a standard breast biopsy needle.  Samples will be formalin-fixed and paraffin 
embedded, coded and stored in the TRIP laboratory.  The risks of a breast biopsy are overall low 
and include risk for bleeding, infection and pain.  For non-UWCCC sites, please refer to study 
procedure manual for sample shipping instructions and tissue requirements.         
 
After completion of the window trial, a tissue microarray (TMA) comprised of all tissue samples 
will be prepared.  This will include all nEGFR positive cases who consented to study.  Tumor 
tissue will include the initial breast cancer tumor biopsies (thus, pretreatment or baseline samples),  
follow up tumor tissue from surgical specimens or biopsies..The TMA will be constructed to 
evaluate the effect of dasatinib therapy on plasma membrane and nEGFR expression.  Sections of 
the TMA will be fluorescently triple stained with anti-EGFR, anti-E-cadherin, and DAPI per 
previously established protocol84 and images will be acquired through the VECTRA imaging 
system (PerkinElmer, Waltham, MA). Following multispectral de-convolution, co-localization of 
EGFR with DAPI or E-cadherin will be assessed by fluorescence intensity to quantitatively 
determine nuclear and plasma membrane EGFR expression, respectively.  Again, to ensure 
preservation of sufficient biopsy and resection tissue for clinical care, no more than 50% of the 
tissue available in each paraffin block will be utilized for TMA construction. If there is not 
sufficient tumor material for creation of a TMA, unstained slides will be used.    
 
8.2 
Exploratory Studies 
 
If consent is obtained for future studies, additional tumor tissue (blocks or slides) remaining will 
be stored for future studies.  This may include studies evaluating mechanisms of resistance to 
dasatinib therapy impact on EGFR location or other studies related to cancer.   
 
Please see section 9.0 further information on storage for future research. 
 
9. CONFIDENTIALITY & DATA MANAGEMENT 
 
To protect the confidentiality of subjects in this study the tissue samples will be coded. The key 
will be saved in computer files that are protected via passwords and access rights. The key will 
include: subject name, medical record number, procedure dates or pathology accession number 
(biopsy and surgery), the sample identification codes (one per subject per sample), and the 
nEGFR status of the biopsy samples. For subjects who are deemed eligible and are registered to 
the trial their subject ID number will also be listed on the key. The rest of the research data will 
also be saved in secure computer files or in the OnCore database which is only accessible via 
username, password, and access rights.  
 
After subjects sign the informed consent form, the biopsy samples will be obtained and coded 
prior to the samples being provided to the Wheeler lab or the Translational Research Initiatives 
in Pathology (TRIP) lab. Both of these labs are only accessible to individuals involved with the 
research completed in those labs. Subjects whose biopsy tissue is nEGFR negative will not be 
eligible for the dasatinib component of the trial but the remaining slides of tumor tissue collected 
will be analyzed as a control cohort for the nEGFR testing and any tissue remaining after 
completion of the nEGFR testing for these subjects will be destroyed.  
 


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28 
Subjects whose tissue is nEGFR positive, and who meet all other eligibility criteria, will be 
enrolled in the dasatinib treatment component of the study. Further samples of their original 
biopsy tissue, as well as tissue from their definitive surgery (post dasatinib treatment or post neo-
adjuvant chemotherapy) will be obtained (for those subjects who go directly to surgery following 
the dasatinib treatment). For those patients receiving dasatinib prior to neoadjuvant 
chemotherapy, a research biopsy will be obtained the day after 7-10 days of dasatinib. This tissue 
will be coded and stored in the TRIP lab.  Archived tumor tissue may be obtained from surgical 
pathology by either the UWCCC research office or the staff in the TRIP lab. The UWCCC 
research staff or the TRIP lab would then code the tissue. For those subjects who did not consent 
to future banking any excess tissue obtained from pathology, but not required for study related 
testing, would be returned to surgical pathology. For non-UWCCC sites, please refer to study 
procedure manual for sample shipping instructions and tissue requirements. 
 
After study related testing is completed the tissue samples from subjects who consent to future 
banking would be stored in the Wheeler lab. These samples will be kept until they are exhausted. 
The link between the study and the samples will be broken one year after completion of the 
primary analysis. A limited data set would be kept associated with these samples as listed in the 
table below. Subjects would be permitted to withdraw their samples from the study or the bank 
up until the time that the link is broken. They would withdraw their samples by informing the 
study PI.  
 
Table 4. Identifiers and information to be collected 
Identifiers and information to be collected for this study:   
  Name 
 
Medical record number 
  Date of birth 
  Race and Ethnicity 
  Address 
  Telephone and fax numbers 
  E-mail address 
  Pathology accession numbers  
 
Health Insurance Information 
  Tumor characteristics 
  Medical history including dates of treatments and procedures 
  Health information generated from participation in the study 
Identifiers and information to be kept for the sample bank for future research: 
  Tumor characteristics 
  Age at diagnosis 
  Race & Ethnicity 
  
Health information generated from participation in the study including 
information related to treatment and study follow-up 
 
 
 


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29 
 
10. STUDY CALENDAR 
 
Patients will have EGFR testing done on initial tumor biopsy/excision done for standard of care.  
If EGFR testing meets criteria for eligibility for dasatinib treatment, remaining of pre-study 
screening tests will then be completed and the on-study calendar below will be followed.  Pre-
study evaluations are to be conducted within 2 weeks prior to start of protocol therapy (day 1).  
A standard of care visit can be used for a pre-study evaluation if within this time window.   In the 
event that the patient’s condition is deteriorating, laboratory evaluations should be repeated 
within 48 hours prior to initiation of the therapy. For patients with negative nEGFR testing, they 
will proceed off study to standard of care treatment.  
 
 
 
nEGFR 
screen 
Pre-
Study 
Screeng 
 
 
Days 1-6 
One – Three  days 
prior to surgery  
(Days 7- 10) or can 
be same day as 
biopsy (Days 8-11)  
 
Day of Breast 
Surgery or 
breast tumor 
biopsy (if 
planning 
neoadjuvant 
chemotherapy) 
(Day 8- 11) 
Day of breast 
surgery for 
neoadjuvant 
chemotherapy 
treated 
nEGFR+ 
group 
Follow-upd 
Dasatinib (if 
nEGFR+) 
 
 
X-----------------------X 
 
 
 
Informed Consent 
X 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
Medical history 
X 
X 
 
 
 
 
 
Concurrent meds 
 
X------------------------------------------------------------X 
 
 
Physical exam 
 
X 
 
X 
 
 
X 
Vital signs 
 
X 
 
X 
 
 
X 
Height 
 
X 
 
 
 
 
 
Weight 
 
X 
 
 
 
 
X 
Performance status 
 
X 
 
X 
 
 
X 
CBC w/diff, plts 
 
X 
 
 
X 
 
 
 
Serum chemistrya 
 
X 
 
 
X 
 
 
 
EKG (as clinically  
indicated) 
 
X 
 
X 
 
 
 
Adverse event 
evaluatione 
 
X-----------------------------------------------------------------------------X 
B-HCG 
 
Xb 
 
 
 
 
 
Radiologic testing 
(per SOC)c 
 
X-----------------------------------------------------------------------------X 
Tumor tissue 
collection for 
nEGFR testing and 
X 
 
 
 
 
X 
X 
 


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30 
inclusion in TMAf  
 
 
a. 
Albumin, alkaline phosphatase, total bilirubin, bicarbonate, BUN, calcium, chloride, creatinine, magnesium, potassium, total 
protein, SGOT [AST], SGPT [ALT], sodium. 
b. 
Serum pregnancy test (women of childbearing potential). 
c. 
Radiologic testing at any time up prior to surgery per SOC 
d. 
Long term clinical follow up for interval treatment and disease status every 4-6 months for 2 years after breast surgery.   
e. 
AE evaluation only required in those who received dasatinib.  Required 30 (+/- 14) days after surgery date (if neoadjuvant 
chemotherapy not given) or through completion of cycle 1 for patients who receive neoadjuvant chemotherapy.   
f. 
After initial nEGFR testing, archived tumor tissue will be collected for all patients for inclusion in the TMA.  For those 
proceeding to surgery, this will include the initial breast biopsy and the surgery specimen.  For those receiving neoadjuvant 
chemotherapy, this will include the initial breast biopsy, a research biopsy the day after dasatinib and prior to neoadjuvant 
chemotherapy (if nEGFR+ and treated with dasatinib) and from the final breast surgery specimen (if residual tumor).    
g. 
Pre-study evaluations, including history and physical, are to be done within 2 weeks prior to start of protocol therapy (Day 1) 
h. 
In the event that the patient’s condition is deteriorating, laboratory evaluations should be repeated within 48 hours prior to 
initiation of the therapy, per MD discretion. 
 
 
11. MEASUREMENT OF EFFECT 
 
11.1 
Antitumor Effect – Solid Tumors 
 
Although significant tumor response is not anticipated with 7-10 days of dasatinib, for the 
purposes of this study, patients will be evaluated for pathologic evidence of response at the time 
of breast surgery.  Evidence of response will be described using tumor cell viability.  
 
11.1.1 Definitions 
 
Evaluable for toxicity.  All patients will be evaluable for toxicity from the time of their 
first treatment with dasatinib 
 
Evaluable for primary endpoint:  All patients who complete at least 7 days of the 
dasatinib with a dose of this medication taken within the 48 hours prior to breast surgery 
and who have adequate breast tissue for nuclear EGFR testing from surgery specimen. 
 
12. DATA SAFETY AND MONITORING PLAN 
 
Oversight And Monitoring Plan 
The UWCCC Data and Safety Monitoring Committee (DSMC) is responsible for the 
regular review and monitoring of all ongoing clinical research in the UWCCC.  A 
summary of DSMC activities are as follows: 
• Reviews all clinical trials conducted at the UWCCC for subject safety, protocol 
compliance, and data integrity. 
• Reviews all Serious Adverse Events (SAE) requiring expedited reporting, as defined 
in the protocol, for all clinical trials conducted at the UWCCC, and studies conducted 
at external sites for which UWCCC acts as an oversight body. 
• Reviews all reports generated through the UWCCC DSMS elements (Internal Audits, 
Quality Assurance Reviews, Response Reviews, Compliance Reviews, and Protocol 
Summary Reports) described in Section II of this document. 


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31 
• Notifies the protocol Principal Investigator of DSMC decisions and, if applicable, any 
requirements for corrective action related to data or safety issues. 
• Notifies the CRC of DSMC decisions and any correspondence from the DSMC to the 
protocol Principal Investigator. 
• Works in conjunction with the UW Health Sciences IRB in the review of relevant 
safety information as well as protocol deviations, non-compliance, and unanticipated 
problems reported by the UWCCC research staff.  
• Ensures that notification is of SAEs requiring expedited reporting is provided to 
external sites participating in multi-institutional clinical trials coordinated by the 
UWCCC. 
 
Monitoring and Reporting Guidelines 
 
UWCCC quality assurance and monitoring activities are determined by study sponsorship and 
risk level of the protocol as determined by the PRMC.  This study falls into the category for: 
Intermediate Monitoring 
Protocols subject to intermediate monitoring generally include UW Institutional Phase I/II and 
Phase II Trials. These protocols undergo review of subject safety at regularly scheduled DOWG 
meetings where the results of each subject’s treatment are discussed and the discussion is 
documented in the DOWG meeting minutes. The discussion includes the number of subjects 
enrolled, significant toxicities, dose adjustments, and responses observed. Protocol Summary 
Reports are submitted on a semi-annual basis by the study team for review by the DSMC. 
 
The UWCCC Data and Safety Monitoring Committee has approved the request to provide 
oversight for the conduction of this study at University of Illinois at Chicago.  All instances of 
SAEs, unanticipated problems and non-compliance will be reported to UWCCC DSMC and UW 
Health Sciences IRB via notification to Dr. Wisinski and Tammy Koehn.  Deviations will be 
entered into Oncore directly by the UIC study team. 
 
REVIEW AND OVERSIGHT REQUIREMENTS   
 
a) Serious Adverse Event – Reported Within 24 Hours 
Serious Adverse Events requiring reporting within 24 hours (as described in the protocol) must 
also be reported to the Data and Safety Monitoring Committee (DSMC) Chair via an email to 
saenotify@uwcarbone.wisc.edu within one business day.  The OnCore SAE Details Report must 
be submitted along with other report materials as appropriate (NCI AdEERS form or FDA 
Medwatch Form #3500 and/or any other documentation available at that time of initial 
reporting). The DSMC Chair will review the information and determine if immediate action is 
required. Within 10 working days, all available subsequent SAE documentation must be 
submitted electronically along with a 24 hour follow-up SAE Details Report and a completed 
UWCCC SAE Routing Form to saenotify@uwcarbone.wisc.edu. All information is entered and 
tracked in the UWCCC database. 
The Principal Investigator notifies all investigators involved with the study at the UWCCC, the 
IRB, the sponsor, and the funding agency and provides documentation of these notifications to 


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32 
the DSMC. 
If the SAE occurs on a clinical trial in which the UW PI serves as the sponsor-investigator, the PI 
reviews the event to determine whether the SAE requires reporting to the FDA and other 
participating investigators 
For a multiple-institutional clinical trial the PI is responsible for ensuring SAEs are reported to 
the FDA as well as to all participating investigators. 
 
See Section below for detailed instructions on SAE reporting. 
b) Serious Adverse Event – Reported within 10 Days 
Serious Averse Events requiring reporting within 10 days (as described in the protocol) must also 
be reported to the Data and Safety Monitoring Committee (DSMC) Chair via an email to 
saenotify@uwcarbone.wisc.edu.  The OnCore SAE Details Report must be submitted along with 
other report materials as appropriate (NCI AdEERS form or FDA Medwatch Form #3500 and/or 
any other documentation available at that time of initial reporting). The DSMC Chair will review 
the information and determine if further action is required. All information is entered and tracked 
in the UWCCC database. 
The Principal Investigator notifies all investigators involved with the study at the UWCCC, the 
IRB, the sponsor, and the funding agency and provides documentation of these notifications to 
the DSMC. 
If the SAE occurs on a clinical trial in which the UW PI serves as the sponsor-investigator, the PI 
reviews the event to determine whether the SAE requires reporting to the FDA and other 
participating investigators.   
For a multi-institutional clinical trial the PI is responsible for ensuring SAEs are reported to the 
FDA as well as to all participating investigators. 
See Section below for detailed instructions on SAE reporting. 
c) 
Sponsor-Investigator Responsibilities for SAE Review 
In the event the UWCCC Principal Investigator is acting as the Sponsor-Investigator (i.e., the PI 
holds the IND), the PI assumes responsibilities of the study sponsor in accordance with FDA 21 
CFR 312.32.  In this capacity, the UWCCC PI reviews all reports of serious adverse events 
occurring on the study at the UWCCC and participating external sites and makes a determination 
of 1) suspectedness (i.e., whether there is a reasonable possibility that the drug caused the AE); 
and 2) unexpectedness (the event is not listed in the Investigator’s Brochure or is not listed at 
the specificity or severity that has been observed) in the context of this study.  SAE with 
suspected causality to study drug and deemed unexpected are reported as IND Safety Reports by 
the UWCCC PI to the FDA, all participating investigators on the study, and the external global 
sponsor (if applicable) within 15 calendar days. All fatal or life-threatening SAE that are 
unexpected and have suspected causality to the study drug will be reported by the UWCCC PI to 
the FDA, all participating investigators on the study, and the external global sponsor (if 
applicable) within 7 calendar days. 
 
Refer to Section below for UWCCC PI instructions for reporting to the FDA. 


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33 
d) Study Progress Review 
Protocol Summary Reports (PSR) are required to be submitted to the DSMC in the timeframe 
determined by the risk level of the study (quarterly; semi-annually; or annually). The PSR 
provides a cumulative report of SAEs, as well as instances of non-compliance, protocol 
deviations, and unanticipated problems, toxicities and responses that have occurred on the 
protocol in the timeframe specified.  PSRs for those protocols scheduled for review are reviewed 
at each DSMC meeting. 
Protocol Summary Reports enable DSMC committee members to assess whether significant 
benefits or risks are occurring that would warrant study suspension or closure. This information 
is evaluated by the DSMC in conjunction with other reports of quality assurance activities (e.g., 
reports from Internal Audits, Quality Assurance Reviews, etc.) occurring since the prior review 
of the protocol by the DSMC.  Additionally, the DSMC requires the study team to submit 
external DSMB or DSMC reports, external monitoring findings for industry-sponsored studies, 
and any other pertinent study-related information.  
In the event that there is significant risk warranting study suspension or closure, the DSMC will 
notify the PI of the DSMC findings and ensure the appropriate action is taken for the protocol 
(e.g., suspension or closure).  The DSMC ensures that the PI reports any temporary or permanent 
suspension of a clinical trial to the sponsor (e.g., NCI Program Director, Industry Sponsor 
Medical Monitor, Cooperative Group Study Chair, etc.) and other appropriate agencies. DSMC 
findings and requirements for follow-up action are submitted to the CRC.   
I. EXPEDITED REPORTING OF SERIOUS ADVERSE EVENTS 
 
Depending on the nature, severity, and attribution of the serious adverse event an SAE 
report will be phoned in, submitted in writing, or both according to Table below. All 
serious adverse events must also be reported to the UWCCC Data and Safety Monitoring 
Committee Chair.  All serious adverse events must also be reported to the UW IRB (if 
applicable), and any sponsor/funding agency not already included in the list. 
 
Determine the reporting time line for the SAE in question by using the following table.  
Then refer to sections A and B below if the SAE occurred at the UWCCC.   
 
Expedited Reporting Requirements for Adverse Events that occur on Studies within 30 Days of 
the Last Administration of the investigational Agent/Intervention 
 
FDA Reporting Requirements for Serious Adverse Events (21 CRF Part 312) 
NOTE: Investigators MUST immediately report to the UWCCC and any other parties outlined in the protocol 
ANY Serious Adverse Events, whether or not they are considered related to the investigational 
agent(s)/intervention (21 CFR 312.64). 
An adverse event is considered serious if it results in ANY of the following outcomes: 
1) Death. 
2) A life-threatening adverse event. 
3) An adverse event that results in inpatient hospitalization or prolongation of existing hospitalization 
for ≥ 24 hours. 
4) A persistent or significant incapacity or substantial disruption of the ability to conduct normal life 
functions. 
5) A congenital anomaly/birth defect. 


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34 
6) Important Medical Events (IME) that may not result in death, be life threatening, or require 
hospitalization may be considered serious when, based upon 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 (FDA, 21 CFR 312.32; ICH E2A and ICH E6). 
ALL SERIOUS adverse events that meet the above criteria* MUST be immediately reported to the 
UWCCC within the timeframes detailed in the table below: 
Hospitalization 
Grade 1 
Timeframes 
Grade 2 
Timeframes 
Grade 3 
Timeframes 
Grade 4 & 5 
Timeframes 
Resulting in 
hospitalization  ≥ 
24 hrs 
10 Calendar Days 
24-Hour; 5 
Calendar Days 
Not resulting in 
Hospitalization ≥ 
24 hrs 
Not required 
10 Calendar Days 
Expedited AE reporting timelines are defined as: 
• 
24-Hour; 5 Calendar Days – The AE must initially be reported within 24 hours of learning of the 
AE, followed by a complete expedited report within 5 calendar days of the initial 24-hour report. 
• 
10 Calendar Days – A complete expedited report on the AE must be submitted within 10 
calendar days of learning of the AE 
1 Serious adverse events that occur more than 30 days after the last administration of investigational 
agent/intervention and have an attribution of possible, probable, or definite require reporting as follows: 
 
Expedited 24-hour notification followed by complete report within 5 calendar days for: 
• 
All Grade 4 and Grade 5 AEs 
Expedited 10 calendar day reports for: 
• 
Grade 2 adverse events resulting in hospitalization or prolongation of hospitalization 
• 
Grade 3 events 
 
2 For studies using PET or SPECT IND agents, the AE reporting period is limited to 10 radioactive half-
lives, rounded UP to the nearest whole day, after the agent/intervention was last administered.  
Footnote “1” above applies after this reporting period.  
 
A.        SAE Requiring [24] Hour Reporting Occurs at UWCCC: 
 
1. Report to the UWCCC: 
 
Reference the SAE SOP (Standard Operating Procedure) and the SAE Reporting 
Workflow for DOWGs on the UWCCC website (http://www.uwccc.wisc.edu) for 
specific instructions on how and what to report to the UWCCC for [24] hour initial 
and follow-up reports.  A follow-up report is required to be submitted within 10 
days of the initial [24] hour report.   
 
For this protocol, the following UWCCC entities are required to be notified: 
 
a) saenotify@uwcarbone.wisc.edu 
b) Kari B. Wisinski, MD (kbwisinski@medicine.wisc.edu) 
c) Tammy Koehn (tmkoehn@medicine.wisc.edu) 


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d) Any other appropriate parties listed on the SAE Routing Form (for follow-up 
reports only) 
 
2. Report to the IRB: 
 
       
 
Consult the UW-IRB website for reporting guidelines.   
      
   If the SAE Requiring 24 Hour Reporting occurs at UIC: 
       The UW PI is responsible for the submission of the SAE to the UW Health Sciences IRB             
       for any site for which the UW serves as the IRB of record.  The outside site should enter the   
       initial information and upload any supporting documentation into Oncore and send an email     
       (within 24 hours) with the subject line of “UW15114 24 Hour SAE Notification” to:  
a) Dr. Kari Wisinski (kbwisinski@medicine.wisc.edu) 
b) Tammy Koehn (tmkoehn@medicine.wisc.edu) 
 
Upon receipt of the outside SAE report, it will be reviewed by the PI and the Breast DOT 
will be responsible for reporting it to UW Health Sciences IRB and UWCCC DSMC via 
reporting outlines for SAEs requiring 24 hour reporting at UWCCC. 
 
A follow up report should be entered into Oncore within 10 days and email notification 
sent to:  
c) Dr. Kari Wisinski (kbwisinski@medicine.wisc.edu) 
d) Tammy Koehn (tmkoehn@medicine.wisc.edu) 
 
 
B.        SAE Requiring [10] Day Reporting Occurs at UWCCC:  
 
1. Report to the UWCCC: 
 
Reference the SAE SOP and the SAE Reporting Workflow for DOWGs on the 
UWCCC website (http://www.uwccc.wisc.edu) for specific instructions on how 
and what to report to the UWCCC for [10] day reports.   
 
For this protocol, the following entities are required to be notified: 
 
a) saenotify@uwcarbone.wisc.edu 
b) Any appropriate parties listed on SAE Routing Form 
 
2. Report to the IRB: 
 
       
 
Consult the UW-IRB website for reporting guidelines. 
 
   If the SAE Requiring 10 day reporting occurs at UIC: 
       The UW PI is responsible for the submission of the SAE to the UW Health Sciences IRB             
       for any site for which the UW serves as the IRB of record.  The outside site should enter the   
       initial information and upload any supporting documentation into Oncore and send an email     


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36 
       with the subject line of “UW15114 10 Day SAE Notification” to:  
e) Dr. Kari Wisinski @ kbwisinski@medicine.wisc.edu 
f) Tammy Koehn @ tmkoehn@medicine.wisc.edu 
Upon receipt of the outside SAE report, it will be reviewed by the PI and the Breast DOT 
will be responsible for reporting it to UW Health Sciences IRB and UWCCC DSMC via 
reporting outlines for SAEs requiring 10 day reporting at UWCCC. 
 
A follow up report should be entered into Oncore within 10 days and email notification 
sent to:  
g) Dr. Kari Wisinski (kbwisinski@medicine.wisc.edu) 
h) Tammy Koehn (tmkoehn@medicine.wisc.edu) 
 
C.        Other Reporting Requirements 
 
Reporting to the FDA 
Serious Adverse Events occurring on studies on which a UW PI is acting as sponsor-
investigator must be reported to the FDA within the appropriate time frame.  Mandatory 
and voluntary reporting guidelines and instructions are outlined on the FDA website: 
http://www.fda.gov/Safety/MedWatch/HowToReport/default.htm 
 
Data Management:  Study personnel are responsible for entering case report forms 
directly into the OnCore database in accordance with the UWCCC data management 
policy. Data must be submitted within 2 weeks of a subject visit and will be monitored 
monthly by the Breast DOWG Cancer Research office at the UWCCC. 
 
Outside sites (UIC) will complete data entry case report forms and submit them to 
UWCCC for data entry.  Data must be submitted within 2 weeks of a subject visit.  Data 
will be verified via submission of de-identified source documentation from UIC to 
UWCCC.  Remote auditing will occur a minimum of once per year. 
 
13. STATISTICAL CONSIDERATIONS 
 
The primary endpoint is the increase in plasma membrane EGFR expression as measured by 
VECTRA.  An increase of at least 25% from baseline to post-dasatinib treatment will be 
considered significant.  We hypothesize that more than 50% of patients will have such increase.  
If there are fewer than 20% who reach this increase, then this would not be considered an 
effective approach.  With a type I error set at 5% and power at 80%, 19 patients will be needed to 
evaluate the primary endpoint.  Analysis will be conducted using exact binomial confidence 
intervals.  It is expected that 20% of all consented TNBC cases will have nEGFR, thus the total 
patient population would be estimated to be approximately 95, with 19 patients treated with 
dasatinib for the primary endpoint.  The secondary endpoint of safety and tolerability of 
dasatinib in patients with operable TNBC will be based on NCIC Adverse Events Version 4.0 
and will be assessed by frequency tables.  In addition, the secondary endpoint of pCR will be 
defined as ypT0 ypNO and assessed by the investigator.  The point estimate of the pCR endpoint 
and its exact 95% confidence intervals (CI) will be calculated.  In evaluating pCR, subjects with 


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37 
missing data will be considered non-responders.   Breast cancer recurrence rates and patterns of 
metastatic recurrence in TNBC by nEGFR status will be described in tabular formats and 
compared by Chi-Square or Fisher’s exact test.  Also, potential intracellular mechanisms which 
impact dasatinib effect will be correlated with cellular localization of EGFR in operable TNBC 
to explore the effects.  Descriptive statistics will be given. Formal statistical tests will be 
performed based on the outcome types and no multiple comparison adjustment will be made due 
to the exploratory nature of the analyses. 
 
With an estimated accrual of one nEGFR positive TNBC patient proceeding to primary breast 
surgery or neoadjuvant chemotherapy every 1-2 months, we anticipate study completion within 
36-48 months 
 
 
 


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38 
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American Cancer Society, Cancer Facts & Figures 2015.,  
 
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Cortazar P, Zhang L, Untch M, et al: Pathological complete response and 
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Cappuzzo F, Ciuleanu T, Stelmakh L, et al: Erlotinib as maintenance 
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Hatakeyama H, Cheng H, Wirth P, et al: Regulation of heparin-binding 
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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. 
 


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Appendix B:   Medications That May Cause QTc Prolongation 
 
Drugs that are generally 
accepted to have a risk of 
causing Torsades de Pointes 
Drugs that in some reports have 
been associated with Torsades de 
Pointes and/or QTc prolongation but 
at this time lack substantial evidence 
for causing Torsades de Pointes 
Drugs that, in some reports, have been weakly 
associated with Torsades de Pointes and/or QTc 
prolongation but that are unlikely to be a risk for 
Torsades de Pointes when used in usual 
recommended dosages and in subjects without other 
risk factors (e.g., concomitant QTc prolonging 
drugs, bradycardia, electrolyte disturbances, 
congenital long QTc syndrome, concomitant drugs 
that inhibit metabolism) 
Generic/Brand Name 
Generic/Brand Name 
Generic/Brand Name 
Amiodarone /Cordarone® 
Alfuzosin /Uroxatral® 
Amitriptyline /Elavil® 
Amiodarone /Pacerone® 
Amantadine /Symmetrel® 
Ciprofloxacin /Cipro® 
Arsenic trioxide /Trisenox® 
Atazanavir /Reyataz® 
Citalopram /Celexa® 
Astemizole /Hismanal® 
Azithromycin /Zithromax® 
Clomipramine /Anafranil® 
Bepridil /Vascor® 
Chloral hydrate /Noctec® 
Desipramine /Pertofrane® 
Chloroquine /Aralen® 
Clozapine /Clozaril® 
Diphenhydramine /Benadryl® 
Chlorpromazine /Thorazine® 
Dolasetron /Anzemet® 
Diphenhydramine /Nytol® 
Cisapride /Propulsid® 
Dronedarone /Multaq® 
Doxepin /Sinequan® 
Clarithromycin /Biaxin® 
Felbamate /Felbatrol® 
Fluconazole /Diflucan® 
Disopyramide /Norpace® 
Flecainide /Tambocor® 
Fluoxetine /Sarafem® 
Dofetilide /Tikosyn® 
Foscarnet /Foscavir® 
Fluoxetine /Prozac® 
Domperidone /Motilium® 
Fosphenytoin /Cerebyx® 
Galantamine /Reminyl® 
Droperidol /Inapsine® 
Gatifloxacin /Tequin® 
Imipramine /Norfranil® 
Erythromycin /Erythrocin® 
Gemifloxacin /Factive® 
Itraconazole /Sporanox® 
Erythromycin /E.E.S.® 
Granisetron /Kytril® 
Ketoconazole /Nizoral® 
Halofantrine /Halfan® 
Indapamide /Lozol® 
Mexiletine /Mexitil® 
Haloperidol /Haldol® 
Isradipine /Dynacirc® 
Nortriptyline /Pamelor® 
Ibutilide /Corvert® 
Lapatinib /Tykerb® 
Paroxetine /Paxil® 
Levomethadyl /Orlaam® 
Lapatinib /Tyverb® 
Protriptyline /Vivactil® 
Mesoridazine /Serentil® 
Levofloxacin /Levaquin® 
Sertraline /Zoloft® 
Methadone /Dolophine® 
Lithium /Lithobid® 
Solifenacin /VESIcare® 
Methadone /Methadose® 
Lithium /Eskalith® 
Trimethoprim-Sulfa /Sulfa® 
Pentamidine /Pentam® 
Moexipril/HCTZ /Uniretic® 
Trimethoprim-Sulfa /Bactrim® 
Pentamidine /NebuPent® 
Moxifloxacin /Avelox® 
Trimipramine /Surmontil® 
Pimozide /Orap® 
Nicardipine /Cardene® 
 
Probucol /Lorelco® 
Nilotinib /Tasigna® 
 
Procainamide /Pronestyl® 
Octreotide /Sandostatin® 
 
Procainamide /Procan® 
Ofloxacin /Floxin® 
 
Quinidine /Cardioquin® 
Ondansetron /Zofran® 
 
Quinidine /Quinaglute® 
Oxytocin /Pitocin® 
 
Sotalol /Betapace® 
Paliperidone /Invega® 
 


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Drugs that are generally 
accepted to have a risk of 
causing Torsades de Pointes 
Drugs that in some reports have 
been associated with Torsades de 
Pointes and/or QTc prolongation but 
at this time lack substantial evidence 
for causing Torsades de Pointes 
Drugs that, in some reports, have been weakly 
associated with Torsades de Pointes and/or QTc 
prolongation but that are unlikely to be a risk for 
Torsades de Pointes when used in usual 
recommended dosages and in subjects without other 
risk factors (e.g., concomitant QTc prolonging 
drugs, bradycardia, electrolyte disturbances, 
congenital long QTc syndrome, concomitant drugs 
that inhibit metabolism) 
Generic/Brand Name 
Generic/Brand Name 
Generic/Brand Name 
Sparfloxacin /Zagam® 
Perflutren lipid microspheres 
/Definity® 
 
Terfenadine /Seldane® 
Quetiapine /Seroquel® 
 
Thioridazine /Mellaril® 
Ranolazine /Ranexa® 
 
 
Risperidone /Risperdal® 
 
 
Roxithromycin* /Rulide® 
 
 
Sertindole /Serlect® 
 
 
Sertindole /Serdolect® 
 
 
Sunitinib /Sutent® 
 
 
Tacrolimus /Prograf® 
 
 
Tamoxifen /Nolvadex® 
 
 
Telithromycin /Ketek® 
 
 
Tizanidine /Zanaflex® 
 
 
Vardenafil /Levitra® 
 
 
Venlafaxine /Effexor® 
 
 
Voriconazole /VFend® 
 
 
Ziprasidone /Geodon® 
 
 
 
 
 


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Appendix C 
MEDICATION DIARY for Dasatinib 
 
Patient Initials:___  ___  ___ 
   
 
 
 
 
 
This is a medication diary on which you are to record the dasatinib you take.  Enter the 
time when the medication is taken, as well as the dose of medication.  You will begin 
taking the dasatinib on Study Day 1 and continuing through Day 7-10, depending on 
when your breast surgery or biopsy date is scheduled.  The last dose of dasatinib should 
be the day prior to surgery or breast biopsy. 
 
• You should take 1 tablet (100mg) once daily with or without food  
• If antacids (ie. tums/Maalox) are needed, the antacid dose should be administered at 
least 2 hours prior to or after the dose of dasatanib.   
• Try to schedule dasatinib around the same time each day (preferably in the 
morning).   Daily dose may be taken 4 hours early or after the scheduled time. If 
you do not take a medication in the window of scheduled time, do not make it up 
later or the next day.   
• If the medication is not taken, please specify the reason.  If you vomit after taking 
the dose, please indicate on study diary, but do not repeat dose.   
• You are to return this diary and your medication bottle to your Research Staff.   
 
Study 
Day 
Date  
MM/DAY/YEAR 
 
Time Taken 
Dose (mg) 
Comments 
(Side effects, 
complaints, other 
medication) 
1* 
 
 
Dasatinib 
Dose 
Time:______ ________mg  
2 
 
 
Dasatinib 
Dose 
Time:______ ________mg  
3 
 
 
Dasatinib 
Dose 
Time:______ ________mg  
4 
 
 
Dasatinib 
Dose 
Time:______ ________mg  
5 
 
 
Dasatinib 
Dose 
Time:______ ________mg  
6 
 
 
Dasatinib  
Dose 
Time:______ ________mg  
7 
 
 
Dasatinib  
Dose 
Time:______ ________mg  
8 
 
Dasatinib  
Dose 
Time:______ ________mg  
9 
 
Dasatinib  
Dose 
Time:______ ________mg  
10 
 
Dasatinib  
Dose 
Time:______ ________mg