Pre-Surgical Trial of Letrozole in Post-Menopausal Patients With Operable Hormone-Sensitive Breast Cancer
Phase: PHASE2
Conditions: Breast Cancer Female
Sponsor: University of Texas Southwestern Medical Center

Cover page
Title: Pre-Surgical Trial of Letrozole in Post-Menopausal Patients with
Operable Hormone-Sensitive Breast Cancer
NCT number: NCT03747042
Document date: 08/16/2021

STU2018-0015, Unni, FormA3-ResearchProtocol-V5-08.16.2021, Mod_24, 12-20-21 (5) 
 
 
 
 
 
SCCC- 11118   
 
Pre-Surgical Trial of Letrozole in Post-Menopausal Patients with   
Operable Hormone-Sensitive Breast Cancer  
 
Principal Investigator:    
Nisha Unni, MD   
  
  
  
  
UT Southwestern Medical Center   
 
  
  
  
5323 Harry Hines Blvd.   
  
  
  
  
Dallas, Texas 75390-9179   
  
  
  
  
214-648-4180   
  
  
  
  
214-648-6918   
  
  
  
  
Nisha.Unni@utsouthwestern.edu   
 
  
  
CONFIDENTIAL   
This material is the property of the UTSW Simmons Comprehensive Cancer Center.    
Do not disclose or use except as authorized in writing by the study sponsor.   
  
Template Updated: 12/2012; 9/2013; 8/2014; 4/2015; 10/2015; 11/2016   
   
 
 
 
 
  
  
  
  
  
Biostatistician    
  
Hong Zhu   
  
  
  
  
UT Southwestern Medical Center   
 
  
  
  
5323 Harry Hines Blvd.   
  
  
  
  
Dallas, TX  75390   
  
  
  
  
Clinical Sciences   
  
  
  
  
214-648-7438   
  
  
  
  
Hong.Zhu@utsouthwestern.edu   
  
  
  
  
   
  
  
  
  
   
Funding Source:                            CPRIT Grant / Carlos L. Arteaga, MD   
 
Initial version:  
June 22, 2018 Version 1.0  
 
Amended:  
  
April 9, 2019                   Version 2.0 
                                                          December 20, 2019 
 Version 3.0  
 
 
June 2, 2020 
 
 Version 4.0   
 
 
August 16, 2021 
 
 Version 5.0  
 
 
 
 
 
 
 
 
UT Southwestern Medical Center (UTSW)   
Harold C. Simmons Comprehensive Cancer Center   

STU2018-0015, Unni, FormA3-ResearchProtocol-V5-08.16.2021, Mod_24, 12-20-21 (5) 
 
Attn: Clinical Research Office   
5323 Harry Hines Blvd. MC 9179   
Dallas, Texas 75390-9179  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
CONFIDENTIAL   
This material is the property of the UTSW Simmons Comprehensive Cancer Center.    
Do not disclose or use except as authorized in writing by the study sponsor.   
  
Template Updated: 12/2012; 9/2013; 8/2014; 4/2015; 10/2015; 11/2016   
   
 
 
 
 
 
 
     Signature Page  
 
Protocol Version August 16, 2021 Version 5.0 
SCCC-11118   
 
The signature below constitutes the approval of this protocol and the attachments, and provides the  
necessary assurances that this trial will be conducted according to all stipulations of the protocol,  
including all statements regarding confidentiality, and according to local legal and regulatory requirements  
and applicable U.S. federal regulations and ICH guidelines.   
 
Protocol Title:  
Pre-Surgical Trial of Letrozole in Post-Menopausal Patients with   
Operable Hormone-Sensitive Breast Cancer 
 
 
Principal Investigator (PI) Name:_____________________________   
 
 
PI Signature: _____________________________   
 
 
Date: ____________________   
 
 

STU2018-0015, Unni, FormA3-ResearchProtocol-V5-08.16.2021, Mod_24, 12-20-21 (5) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
CONFIDENTIAL   
This material is the property of the UTSW Simmons Comprehensive Cancer Center.    
Do not disclose or use except as authorized in writing by the study sponsor.   
  
Template Updated: 12/2012; 9/2013; 8/2014; 4/2015; 10/2015; 11/2016   
   
Study Number – SCCC-11118   
________________________________________________________________________   
                                       TABLE OF CONTENTS   
  
  
  
  
   
LIST OF ABBREVIATIONS   
STUDY SCHEMA   
STUDY SUMMARY   
1.0       Introduction   
2.0 
Objectives   
3.0 
Eligibility Criteria   
4.0  
Ineligibility Criteria   
5.0 
Registration Procedures   
6.0  
Study Schedule of Events   
7.0 
Treatment Plan   
7.1 
Letrozole Treatment   
7.2 
Tissue Specimen Acquisition   
7.3 
Definitive surgery   
7.4  
Pathology   
7.5  
Further Testing   
7.6 
Blood specimen   
8.0 
Adverse Event Reporting   
8.1 
Adverse Event Monitoring   
8.2. 
Steps to determine If an Adverse Event Requires Expedited Reporting   
9.0 
Dose Modifications and Management of Toxicity  
10.0 
Drug Information   
11.0 
Removal of Patients from Protocol Therapy   
12.0 
Statistical Considerations   
13.0 
Data Safety    
14.0 
Monitoring and Quality Assurance   
15.0 
Changes in Protocol   
16.0 
Protocol Deviations   
17.0 
Data Management   
18.0 
Data Verification   
19.0 
Study Documentation   
20.0 
Records Retention   

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21.0 
Confidentiality    
22.0 
References   
 
Appendix I: Lab Manual   
 
 
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Absolute Lymphocyte Count   
American Society of Clinical Oncology   
AST  
Aspartate Aminotransferase   
BUN  
Blood Urea Nitrogen   
CBC  
Complete Blood Count   
CMP  
Comprehensive Metabolic Panel   
CR  
Complete Response   
CT  
Computed Tomography   
CTCAE  
Common Terminology Criteria for Adverse Events  
DLT  
Dose Limiting Toxicity   
DOT  
Disease Oriented Team   
DSMB  
Data and Safety Monitoring Board   
ECOG  
Eastern Cooperative Oncology Group   
FDA  
Food and Drug Administration   
GCP  
Good Clinical Practice   
H&P  
History & Physical Exam   
HRPP  
Human Research Protections Program   
IDE  
Investigational Device Exemption   
IHC  
Immunohistochemistry   
IND  
Investigational New Drug   
IV (or iv)  
Intravenously   
MRI  
Magnetic Resonance Imaging   
MTD  
Maximum Tolerated Dose   
NCI  
National Cancer Institute   
ORR  
Overall Response Rate   
OS  
Overall Survival   
PBMCs  
Peripheral Blood Mononuclear Cells   
pCR  
Pathologic Complete Response   
PD  
Progressive Disease   
PET  
Positron Emission Tomography   
PFS  
Progression Free Survival   
p.o.  
peros/by mouth/orally   
PR  
Partial Response   
RCB  
Residual Cancer Burden   
RECIST  
Response Evaluation Criteria in Solid Tumors   
SAE  
Serious Adverse Event   
SCCC  
Simmons Comprehensive Cancer Center   
SD  
Stable Disease   
SGOT  
Serum Glutamic Oxaloacetic Transaminase   
SPGT  
Serum Glutamic Pyruvic Transaminase   
 
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Study Number – SCCC-11118   
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WBC  
White Blood Cells   
LIST OF ABBREVIATIONS AE  Adverse 
Event   
ALT  
Alanine Aminotransferase   
ALC   
ASCO   

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STUDY SCHEMA   
 

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ER+/HER2-Negative Operable Breast Cancer, Stage I, II or III   
  in Post-menopausal Women    
   
Paraffin block (preferably) or unstained slides from pre-treatment core biopsies for    
IHC-based assays (Ki67, ER, PR, HER2)    
 
   
 
Letrozole: 2.5 mg PO QD for 7-30 days   
(last dose on the day before surgery)   
 
   
Surgery   
(Standard of Care: Mastectomy vs Lumpectomy)   
Post-treatment frozen AND formalin fixed core biopsies (or surgical resection tissue when   
intraoperative cores not feasible) for IHC-based assays (Ki67, ER, PR, HER2), and molecular  
analysis    
 
  
 
 
 
 
Follow up to assess disease status approximately every 6 months (+/- 1 month) for up to 10   
years   
 
 
 
 
 
 
 
 
 
 
 
 
 
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STUDY SUMMARY   
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
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Study Number – SCCC-11118   
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Title   
Pre-Surgical Trial of Letrozole in Post-Menopausal Patients with Operable   
Hormone-Sensitive Breast Cancer   
Short Title   
Letrozole in Post-Menopausal Patients with Operable Hormone-Sensitive   
Breast Cancer   
Protocol Number   
  SCCC-11118 
Phase   
NA    
Methodology   
Exploratory investigational   
Study Duration   
15 years   
Study Center(s)   
UT Southwestern Medical Center and Parkland Health and Hospital System   
To determine gene expression and/ or mutational or proteomic signatures in   
breast tumors that continue to exhibit high proliferation (i.e., Ki67) upon   
hormone deprivation (with letrozole).   
Number of Subjects   
300   
Diagnosis and Main   
Inclusion Criteria   
Study Product(s), Dose,   
Route, Regimen   
Letrozole 2.5mg once daily.   
Duration of administration   
7-30 days, last dose on the day prior to surgical resection   
Reference therapy   
N/A   
Statistical Methodology   
For gene expression level and/or a mutational or proteomic signature   
parameters, we will use ANOVA to evaluate differences among three or more   
groups and unpaired two-sample t-test for differences between two groups   
stratified based on post-treatment Ki67 index. If underlying assumptions of   
these methods (e.g., normality and homoscedasticity) are not satisfied, we will  
use the nonparametric Kruskal-Wallis ANOVA for three or more group   
comparisons and the nonparametric Wilcoxon rank sum test for two group   
comparisons. Multiple comparisons will be adjusted. P <0.05 will be considered  
statistically significant. We will split the data into training and validation   
datasets, and compute the prediction error to assess the model accuracy with   
cross validation.    
Objectives   
Post-menopausal, ER-positive, HER2-negative stage I, II or III invasive  
mammary carcinoma   

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1.0  
Introduction   
An estimated 175,000 new cases of breast cancer will be diagnosed this year in the United States  
and, despite adjuvant therapy with cytotoxic chemotherapy; many of these women will die of breast  
cancer.  The heterogeneity of these tumors results in a highly varied response to current treatments.  
The developments of rationally-designed agents which target molecules differentially expressed in  
tumor have provided new prospects for the treatment breast cancer. These agents are in general  
tumor selective and therefore more clinically effective than chemotherapeutic drugs. Given that  
75% of invasive breast cancers are either estrogen receptor (ER) or progesterone receptor (PR)  
positive, the most frequently utilized targeted therapies are those that modulate estrogen levels  
and/or ER function.   
     Estrogen receptor in breast cancer: Estrogen plays a critical role in the development and  
progression of breast cancers. The estrogen receptor (ER), a member of the nuclear receptor  
superfamily of transcription factors, mediates most of the actions of estrogen1. There are two ERs,  ER 
and ER. These are products of different genes and have similar but not identical structure.  ER is 
the most studied in breast cancer. Less is known about ER and some data suggest it may  opposing 
functions to those of ER 2,3 Estrogen signaling through ER occurs through distinct  pathways. 
The classical mechanism of ER action involves estrogen binding to ER in the nucleus  after which 
receptors dimerize and bind to specific estrogen-responsive elements (EREs) in the  promoters of 
target genes4. A second non-classical mechanism involves ER-mediated gene  expression 
without direct binding to DNA but through modulation of protein-protein interactions  with other 
DNA-binding transcription factors5. Estrogen can also exert non-genomic actions which  are too rapid 
to be accounted by gene transcription and RNA and protein synthesis. Reported  nongenomic 
effects of 17-estradiol include direct or indirect activation of adenylate cyclase and  production of 
cyclic AMP, mitogen activated protein kinase (MAPK), AKT, c-Src, Shc, the  regulatory subunit 
of phosphoinositide 3-kinase (PI3K) p85, insulin-like growth factor I receptor  (IGF-IR), epidermal 
growth factor receptors (EGFR), HER2 (ErbB2), and endothelial nitric oxide  synthase (eNOS), 
among others6. Finally, amplified growth factor receptor signaling can post- translationally modify 
ERs and co-activators resulting in estrogen-independent transcriptional  activity or ERs at EREs7.    
   Estrogen Modulation:  The standard treatment for ER+ breast cancer includes therapies  
designed to block ER function. Selective ER modulators (SERMs), such as tamoxifen, block ligand  
binding to ER and partially block its activity. Ovarian ablation and luteinizing hormone-releasing  
hormone (LHRH) reduce the level of estrogen, thus inhibiting ligand-induced ER activation.  
Aromatase inhibitors (anastrozole, letrozole, and exemestane) do the same by blocking the main  
enzyme involved in the production of estrogen in post-menopausal women.  Pure antiestrogens,  
such as fulvestrant bind ER and induce receptor degradation8. Several studies have suggested  
modest superiority of aromatase inhibitors over tamoxifen9-14.   
Aromatase Inhibitors: Estrogen deprivation is a commonly utilized approach against hormone- 
dependent breast cancer. Aromatase inhibitors block the peripheral and intra-tumoral enzyme, p450  
aromatase, which catalyzes the final step in estrogen synthesis15. These agents have the advantage  of 
potentially blocking both genomic and non-genomic ER functions1. Today, third-generation  
aromatase inhibitors, such as anastrozole (Arimidex), letrozole (Femara), and exemestane   
 
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(Aromasin) are currently the treatment of choice as first-line therapy of hormone-dependent  
metastatic disease in postmenopausal women and have been shown to be superior to tamoxifen in  this 
setting 10. In a pivotal double-blind phase III study, post-menopausal women with hormone  receptor-
positive or hormone receptor-unknown locally advanced or metastatic breast cancer  received 
treatment with either letrozole or tamoxifen as first-line therapy. Letrozole not only  showed 

STU2018-0015, Unni, FormA3-ResearchProtocol-V5-08.16.2021, Mod_24, 12-20-21 (5) 
 
superior median time to progression compared to tamoxifen, but also superior time to  treatment 
failure, overall response rate, and overall clinical benefit. There was a trend toward  improved 
overall survival in the letrozole arm, but this was not significant. This study documented  the superiority 
of letrozole over tamoxifen in first-line endocrine therapy in postmenopausal women  with advanced 
breast cancer and was one of several trials demonstrating superiority of the  aromatase inhibitors 
over tamoxifen in the first-line setting 9.    
The ATAC Study is a randomized trial compared anastrozole to tamoxifen and the combination of  
both as adjuvant therapy in over 9,000 post-menopausal women with early, ER-positive cancers.  
After a median follow-up of 33 months, there were fewer recurrences and fewer new primary breast  
cancers as well as a statistically better disease-free survival in the anastrozole arm compared to the  
other two treatments11, 12. The MA.17 international Intergroup trial, initiated by the National Cancer  
Institute of Canada-Clinical Trials Group, was a double-blind, randomized, placebo-controlled trial  
performed in over 5,100 postmenopausal women with ER-positive or unknown primary breast  
cancer. It randomized patients who were disease-free after 5 years of adjuvant treatment with  
tamoxifen to an additional 5 years of treatment with letrozole or placebo. The planned duration of  
treatment for patients in the study was 5 years, but the trial was terminated early because of an  
interim analysis showing a favorable letrozole effect on time without recurrence or contralateral  
breast cancer13. An updated analysis was published more recently14. After a median follow-up of 30  
months, women in the letrozole arm had statistically significantly better disease-free survival and  
distant disease-free survival than women in the placebo arm. Overall survival was the same in both  
arms. However, among lymph node–positive patients, overall survival was statistically significantly  
improved with letrozole14. Based on these studies, aromatase inhibitors have become the standard  of 
care in post-menopausal women with hormone-dependent breast cancer.   
Antiestrogen Resistance: While these therapies are effective initially in many patients, the  
overwhelming majority of ER+ breast cancers eventually become resistant to antiestrogens,  
ultimately resulting in disease progression and mortality. Several resistance mechanisms have been  
reported. These include the rare loss of ER by tumors as well as the increasingly frequent selection  of 
cells with ESR1 activating mutations mainly within the ligand-binding domain (LBD). Ligand- 
independent ER-mediated transcription, and perturbation of the interaction ER and coactivators and  
corepressors of transcription1 has been of particular interest. Alterations in the intracellular  
pharmacology including membrane-initiated steroid signaling (non-genomic) where cellular ER  
signaling occurs via direct interaction with growth factor signal transduction pathways. These can  be 
activated by estrogen or SERMs such as Tamoxifen16. Activation of surface tyrosine kinase  
receptors such as IGF-IR, EGFR, and HER2 as well as kinases such as c-Src, PI3 kinase, MAPK,  and 
AKT result in activation of ER or its coactivator proteins.  This crosstalk between the ER  receptor 
signaling pathway and growth factor receptor pathways can result in genomic activation of  ER despite 
the presence of antiestrogens 17,18. SERMs may repress or activate transcription of  estrogen-target 
genes depending on the absolute and relative levels of ER coregulator proteins 19, 20.   
 
 
 
 
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Upon the emergence of resistance, the agonistic effects of SERMs like tamoxifen can predominate,  
leading to breast tumor growth. However, very little clinical data exist to confirm these mechanisms  
in primary tumors.   
Study Rationale:  Pre-surgical and neoadjuvant studies with anti-cancer therapies have suggested  that 
treatment-induced cellular or molecular endpoints in tumor tissues can correlate with clinical  
outcome. For example, treatment-induced tumor cell apoptosis, as measured by cleaved caspase-3  

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IHC, one week after administration of the HER2 antibody trastuzumab correlates with clinical  
response of HER2-overexpressing breast cancers 21. The neoadjuvant IMPACT trial compared the  
anastrozole, tamoxifen, and the combination. Drug-induced inhibition of tumor cell proliferation at  2 
weeks, as measured by Ki67, was better in patients treated with anastrozole vs. the other two arms  22, 
23. This result parallels the result of the large ATAC trial where relapse-free survival was better in   
patients treated with adjuvant anastrozole compared to the other two arms 24, 25. Taken together,  
these data suggest that the ‘intervened’ Ki67 index in ER+ surgically-removed breast cancers  
treated for a short time with tamoxifen or an aromatase inhibitor can be used to identify cancers that  
are highly sensitive to antiestrogens vs. those that are resistant or destined to recur faster (Fig 1). If  so, 
tumors segregated as a function of a low or high Ki67 after a short pre-surgical period will  harbor 
different somatic alterations, gene expression and/or protein signatures. In addition, we  
hypothesize that tumors with a high Ki67 after hormonal therapy will harbor genomic alterations,  
molecules or ‘pathways’ that are causally associated with antiestrogen resistance and/or a  
biomarker of such phenotype.    
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
We propose a short pre-surgical non-therapeutic trial involving postmenopausal women with newly  
diagnosed ER+, HER2-Negative operable breast cancers.  After undergoing a core needle biopsy   
 
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for tissue acquisition, study participants will take a 7 to 30-day course of letrozole in  accordance with 
standard of care.  They will then undergo definitive surgical resection of their  primary tumor 
(mastectomy vs lumpectomy) as per standard of care guidelines. We should note that  in all these 
patients, the standard of care will be at least 5 years of post-operative adjuvant letrozole  – or another 
aromatase inhibitor. Pre and post treatment tumor tissue will be processed for routine  histopathologic 
analysis and study analysis with:    
1) Cell proliferation assay: Ki67 IHC.    
Figure 1. Clinical Trial Schema. Diagram
shows the percentage of Ki67+ cells in
pre- and post-anastrozole (2-week) tumor
sections 
from 
biopsies 
of 
individual
patients enrolled in the IMPACT trial
(data provided by Mitch Dowsett). In this
trial, patients continued therapy for a total  of
3 months after a biopsy done at 2  weeks.
In this application, we propose that  letrozole
will induce similar changes in  tumor
proliferation 
to 
those 
induced 
by
anastrozole after a 2-week pre-surgical
‘window’. We hypothesize that 1) the
Ki67 index measured in the surgical
specimen will allow the segregation of
highly 
hormone-dependent 
tumors 
vs.
those that are not, and 2) tumors with high 
Ki67 in the surgical specimen will harbor
genomic 
alterations, 
molecules 
or
‘pathways’ that are causally associated
with 
antiestrogen 
resistance 
and/or 
a
biomarker of such phenotype.   

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In addition, post treatment tumor tissue will be processed for study analysis including:   
1) DNA and RNA sequencing of tumor tissue   
We hypothesize that:   
 1) The Ki67 index measured in the surgical specimen will allow the segregation of highly  
hormone-dependent tumors vs. those that are not. Molecular profiling of those tumors will identify  
genomic alterations and/or gene expression/protein alterations causally associated with endocrine  
resistance.   
1.1  Letrozole   
Femara (Letrozole) is a non-steroidal aromatase inhibitor (inhibitor of estrogen synthesis). It is an   
FDA approved agent from Novartis for the treatment of ER+ post-menopausal breast cancer.    
1.11 Clinical Study Results   
In a double-blind, phase III study, 907 post-menopausal women with hormone receptor-positive or   
hormone receptor-unknown locally advanced or metastatic breast cancer received treatment with  
either letrozole or tamoxifen as first-line therapy. Letrozole showed superior median time to  
progression (TTP) compared to tamoxifen (9.4 months vs. 6.0 months) 9. This trial was one of  
several supporting use of aromatase inhibitors over tamoxifen in post-menopausal patients in the  
metastatic setting.   
The MA.17 International Intergroup Trial, initiated by the NCI of Canada-Clinical Trials Group,  
was a double-blind, randomized, placebo-controlled trial performed in over 5,100 postmenopausal  
women with ER+ or unknown primary breast cancer. This trial randomized patients who were  
disease-free after 5 years of adjuvant treatment with tamoxifen to an additional 5 years of treatment  
with letrozole or placebo. The planned duration of treatment for patients in the study was 5 years,  but 
the trial was terminated early because of an interim analysis showing a favorable letrozole effect  on 
time without recurrence or contralateral breast cancer13. An updated analysis was published  more 
recently14. After a median follow-up of 30 months, women in the letrozole arm had  statistically 
significantly better disease-free survival and distant disease-free survival than women in  the placebo 
arm. Overall survival was the same in both arms. However, among lymph node–positive  patients, 
overall survival was statistically significantly improved with letrozole. Letrozole is FDA  approved 
for endocrine treatment of hormone-sensitive breast cancers in the adjuvant and  metastatic 
settings.    
Side effects encountered include hot flashes, anorexia, arthralgia, myalgia, and alopecia, which  
were all statistically significantly more common in those receiving letrozole; vaginal bleeding was  
statistically significantly more common in those receiving placebo. More patients receiving  
letrozole had a fracture, a new diagnosis of osteoporosis or cardiovascular disease on study, but   
 
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only the incidence of self-reported new osteoporosis was statistically significantly different between  the 
two arms. Other side effects associated with the use of letrozole include, but are not limited to,  nausea 
and vomiting, fatigue, headache, dizziness, swelling of the hands, feet, or lower legs,  
constipation, diarrhea, abdominal pain, and/or cough.   
2.0         Objectives  
2.0  
         Primary Objective  
To determine that in breast tumors that continue to exhibit high proliferation (i.e., Ki67) upon 
hormone deprivation (with letrozole), their gene expression and/or a mutational or proteomic 
signatures will harbor molecules or ‘pathways’ that are biomarkers of resistance to endocrine 
therapy or a cause of it.  
The ultimate goal of these aims is to identify clinically-targetable pathways which can  

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be exploited to enhance responses and survival in patients with ER+ breast cancer. This  
study will also provide a platform of ER+ tumors with pharmacodynamic information  
following estrogen suppression in vivo (Ki67) that can be used to segregate tumors into  
‘responders’ and ‘not responders’ where we can focus the unbiased discovery of  
mechanisms of resistance to endocrine therapy  
 
3.0 
Eligibility Criteria  
3.0 
Eligibility waivers are not permitted. Subjects must meet all of the inclusion and  
exclusion criteria to be registered to the study. Study treatment may not begin until  
a subject is registered.  
3.1 
Patients must provide informed written consent  
3.2 
ECOG performance status 0-2.  
3.3 
Clinical stage operable I, II or III invasive mammary carcinoma, which is ER-  
positive by IHC and HER2-negative by Herceptest (0 or 1+) or not amplified by  
in situ hybridization as per routine clinical testing  
 3.3.1 Patients who have measurable residual tumor at the primary site  
 3.3.2 Patients who will undergo surgical treatment with either segmental resection  
or total mastectomy  
3.4 
Measurable tumor  
Measurable disease: a mass that can be reproducibly measured by physical exam,   
mammogram or ultrasound and is at least 1 cm in size  
3.5 
Post-menopausal female subjects ≥18 years of age, as defined by any of the  
following:  
• 
Subjects at least 55 years of age;  
• 
Subjects under 55 years of age and amenorrhoeic for at least 12 months or 
follicle-stimulating hormone (FSH) values ≥40 IU/L and estradiol levels ≤ 
pg/mL (140 pmol/L) or in postmenopausal ranges per local or institutional 
reference ranges;  
• 
Prior bilateral oophorectomy or prior radiation castration with amenorrhea for at   
least 6 months. 
 
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 (There is no upper age limit for enrollment to this study)  
3.6 
No prior chemotherapy for this primary breast cancer.  
3.7 
Patients with a prior history of contralateral breast cancer are eligible if they have   
no evidence of recurrence of their initial primary breast cancer.  
3.8 
Women may have been taking tamoxifen or raloxifene as a preventive agent prior to   
study entry but must have discontinued the drug for at least 14 days prior to study   
enrollment.  
3.9 
Subjects must have ended hormone replacement therapy (HRT) (e.g., conjugated   
estrogens tablets, USP, [Premarin]), at least 7 days prior to receiving the first dose   
of randomized therapy.  
3.10 
Patients must have adequate hepatic and renal function. All tests must be obtained   
less than 4 weeks from study entry.  This includes:  
Creatinine < 2 X upper limits of normal  

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Bilirubin, SGOT, SGPT <1.5X upper limits of normal  
3.11 
Able to swallow and retain oral medication  
 
4.0 
Exclusion Criteria   
Patients with locally advanced disease who are candidates for other preoperative   
chemotherapy at the time of initial evaluation. This may include patients with   
locally advanced disease such as:    
4.0.1 Inflammatory breast cancer (T4d)   
4.0.2 Fixed axillary lymph node metastases (N2)   
4.0.3 Metastasis to ipsilateral internal mammary node (N3)   
4.1 
Locally recurrent breast cancer   
4.2 
Evidence of distant metastatic disease (i.e. lung, liver, bone, brain, etc.)   
4.3 
Serious medical illness that in the judgment of the treating physician places the   
patient at high risk of operative mortality.   
4.4 
Severe uncontrolled malabsorption condition or disease (i.e. grade II/III diarrhea,   
severe malnutrition, short gut syndrome)   
4.5 
Dementia, altered mental status, or any psychiatric condition that would prohibit the   
understanding or rendering of informed consent.   
4.6 
Use of an investigational drug within 30 days or 5 half-lives, whichever is longer,   
preceding the first dose of letrozole.   
 
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
5.0    Registration Procedures   
All subjects must be registered with the research office before enrollment to study. Prior to  
registration, eligibility criteria must be confirmed with the Study Coordinator. All subjects will  be 
registered in the Velos Clinical Management System.     
 
Upon confirmation of eligibility and enrollment as per the afore-mentioned instructions, the   
subject will be assigned a secondary number in the order of enrollment. All the subject numbers  
will end in “LETROZOLE”. For example, subject 001-LETROZOLE will become 001-01-  
LETROZOLE upon enrollment. If subject 002-LETROZOLE screen fails, and subject 003-  
LETROZOLE is the next subject enrolled, subject 003-LETROZOLE will become 003-02-  
LETROZOLE and so-on.    
 
Each newly consented subject should be numbered using the schema provided above. Upon    
Registration in Velos, the study coordinator will assign the additional registration code according  
to the numbering schema outlined above and maintain the subject’s status throughout the life of  
the study.   
 
The numbering schema should clearly identify the sequential number of the subject enrolled as  
well as the status of the subjects enrolled so that the number of subjects consented versus the   
number of subjects actually enrolled may be easily identified in the Velos system.     
 
To be eligible for registration to the study, the participant must meet each inclusion and  
exclusion criterion listed in the eligibility checklist.    

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Note: All study documents should be in hand 24-48 hours prior to the patient’s anticipated start  
date.    
Upon satisfactory review of eligibility documents submitted, the Principal Investigator will approve  
enrollment. Once registration/enrollment is confirmed, proceed with protocol procedures.   
Issues that would cause treatment delays should be discussed with the Protocol Chair. Any requests  
for eligibility exceptions and/or deviations must be approved in writing by the Principal  
Investigator, the Data and Safety Monitoring Committee, and the UT Southwestern IRB.    
As is generally accepted, standard of care procedures performed prior to consent, but within the  
protocol defined screening window for each assessment, can be used for study purposes. All  
research-only procedures must be performed after patient consent.   
 
 
 
 
 
 
 
 
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
 
 
 
 
6.0 Study Schedule of Events   
 

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1. Performed as routine medical assessment if indicated   
2. Comprehensive Metabolic Panel: Creatinine, glucose, total bilirubin, alkaline phosphatase, and  
SGOT/SGPT within 4 weeks.    
3. We will need access to archived tumor sections from the diagnostic paraffin block (or 10  
unstained slides) from the initial diagnostic core biopsy at some point during the study. These will  be 
requested when appropriate and sent to:   
Sunati Sahoo, M.D., Department of Pathology/UTSW, 5323 Harry Hines Blvd., Dallas, TX 75390- 
9072, Telephone: (214) 633-6361.   
 
4. A tumor sample will be obtained from the surgical specimen at time of surgical resection, and  
flash frozen whenever possible. A companion formalin fixed paraffin embedded tissue sample   
should also be prepared. Both samples should be delivered (snap frozen tissue in liquid nitrogen)  
directly to: Saurabh Mendiratta, 6001 Harry Hines Blvd. NB5.112. Dallas, TX- 75390-8807   
Phone: 214-648-6167.   
 
 
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
5. One 10-ml purple top tube should be collected from all enrolled patients. This blood may be  
collected at any time during or after the study; however, ideally the specimen should be collected at  
the time of enrollment.    
6. Will be given to patients along with medication and will be completed and returned at the post- 
operative visit.   
7. Enrolled patients will be followed approximately every 6 months for up to 10 years.  Follow up  
information may be collected from the patient’s medical record if they are still coming to the  
respective clinic or they will be contacted by telephone. If during this follow up period, the patient  
experiences a recurrence of disease and undergoes a standard of care biopsy, a sample of this tumor  
Tests   
       Pre-Study   
At the time of Surger y        Post - Op   
History & Physical   
X1   
   
X¹   
Performance Status   
X1   
   
X¹   
CMP 2   
X1   
   
   
Tumor Specimen   
X1,3,    
          X1, 4   
X1, 4   
(fixed, paraffin   
embedded)    
Prescription Given  
Letrozole 2.5mg   
X   
   
   
Pill Diary   
X6   
   
X6    
Follow Up   
   
   
X7   
Peripheral Blood   
Collection (Purple   
top) 10ml   
X1,3,   
          X1,4  
      X1, 4   
(frozen section)  
  
         X1, 4  
X1, 4   
X5  
  
   

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tissue may be obtained for additional research testing, pending there is leftover tumor tissue  
available and the patient agrees to provide the sample.       
    
    
7.0 Treatment Plan   
 
7.1  Letrozole Treatment   
Patients will receive a prescription for letrozole to be taken orally at a dose of 2.5 mg/day for 7-30   
days to allow for variations in surgical scheduling. Patients are to undergo surgical resection of their  
tumor the day after the last dose of letrozole.    
 
7.2   Tissue Specimen Acquisition   
Post-treatment tumor biopsies (a goal of 1-6 cores) will be obtained following 7-30 days of   
letrozole treatment on the day of surgery. Frozen AND formalin fixed core biopsies will be  
obtained whenever possible. The tissue will be used for study-specific assays as well as routine  
histopathology. Post-treatment core needle biopsy tissue will be obtained, whenever possible, by  
the surgeon intraoperatively at the time of routine surgical resection. A formalin-fixed paraffin  
embedded tumor block from the patient’s surgical resection is also requested. This block will be  
used for a subset of the correlative studies if no tumor is obtained by the post treatment core  
biopsies. This block will be returned to pathology promptly after the correlative studies are  
completed.    
*If post-treatment needle core biopsies cannot be performed, fresh tissue may be procured from the  
surgical specimen according to institutional guidelines. Tissue must be procured as rapidly as  
possible after the specimen is removed from the patient, preferably with in 5-10 minutes. This  
option is suboptimal and discouraged as tissue degradation will affect the study specific assays. We  
realize; however, this may be unavoidable in some cases.   
 
Instructions on fresh tissue specimen handling:   
According to the following schedule, core biopsy specimens should be immediately flash   
frozen in liquid nitrogen* OR placed in 10% neutral buffered formalin immediately   
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
following removal from the patient following procedures established by the Simmons  
Cancer Center Tissue Core.    
 
• 
For the formalin-fixed cores, an approximately 20:1 volume of formalin 
to tissue should   
be used.  Core biopsies placed in 10% neutral buffered formalin must fix 
at room   
temperature for at least 6 h and no more than 24 h.  Core biopsies must 
be transferred   
from formalin to 70% ethanol after 6-24 h to prevent over fixation and 
placed in the   
refrigerator.   
 
 

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• 
Label ALL cryovials before freezing (see specific instructions #1 below).    
• 
Send all samples to Saurabh Mendiratta at the address mentioned earlier.    
 
 
 
 
 
 
 
 
 
 
 
 
Tissue specimen labeling and documentation:   
 
1.  Label each cryovial and formalin-filled collection container with the patient name, Medical  
Record #, Patient study ID #, location of biopsy, collection date and time of collection using an  
ethanol resistant marker.  Each set of patient samples should be accompanied by a Tissue   
Registration form.     
 
7.3        Definitive Surgery   
Surgical treatment (total mastectomy or segmental resection with lymph node evaluation if   
clinically indicated) will occur the day after completion of therapy.    
The primary lesion obtained at the time of the definitive surgical procedure (partial or total  
mastectomy) will be sent for standard of care histopathologic analysis; wherever possible intra- 
opearative cores from the central portion of the tumor will be obtained for study-specific assays. All  
specimens will be handled according to established institutional guidelines to maintain the accuracy  of 
the analysis of tumor size and margin status.    
 
7.4        Pathology   
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
Following standard of care histopathologic analysis, additional paraffin-embedded sections will be  
submitted at a later time to the Simmons Cancer Tissue Core to determine tumor proliferation with  
Ki67 (MIB1Ab, Dako Cytomation) IHC. These tests are of no clinical utility and will be done for  
research purposes only.   
 
7.5   Further Testing   
Frozen cores and/or peels from the formalin-fixed paraffin-embedded (FFPE) tumor blocks from   
the surgical specimen will be selected and/or macrodissected – when needed – based on ≥20%   
tumor cellularity as assessed by Dr. Sahoo, expert breast pathologist in the trial. These will be   
sent to Dr. Carlos Arteaga’s laboratory for further testing.   
 
 
  
7.6 Blood and Negative Tissue Sample:     
# cores in   
10% NBF   
1   
1   
0   
2   
2   
0   
3   
2   
1   
4   
3   
1   
5   
3   
2   
6   
4   
2   
# Cores  
Obtained   
# cores flash   
frozen   

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One 10 ml purple top tube should be collected from all enrolled patients.  This blood may be   
collected at any time during or after the study; however, ideally the specimen should be collected  
before the patient receives any subsequent therapy.  This blood is requested for the purpose of   
obtaining a buffy coat specimen to be banked for future nucleic acid extraction.  This specimen   
will serve as a highly pure source of patient germline DNA for the correlative studies for this   
protocol.     
                                                                                                                                                              
The blood samples will be logged in, barcoded and processed by a dedicated research team   
member within 6 hours of collection.    
 
All specimens will be stored in a locked freezer and subsequently distributed in a de-identified   
manner as needed for the correlative studies. Only the principal investigators and their designees  
will know the identity of these participants.    
 
  
7.7 Follow Up Information    
Patients that complete the course of letrozole and have their surgery will be followed for disease  
recurrence.  This follow up will be approximately every six months from the patient’s surgery date  for 
up to ten years.  The patient’s disease status will be collected from their medical record, if  
available.  In the event that follow up information is not available in the patient’s medical record, a  
member of the research team will contact the patient.      
 
8.0  Adverse Event Reporting   
  
8.1 
Adverse Event Monitoring   
Adverse event data collection and reporting, which are required as part of every clinical trial, are  
done to ensure the safety of subjects enrolled in the studies as well as those who will enroll in   
future studies using similar agents. Adverse events are reported in a routine manner at scheduled   
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
times during a trial. Additionally, certain adverse events must be reported in an expedited  
manner to allow for optimal monitoring of subject safety and care.    
 
All subjects experiencing an adverse event, regardless of its relationship to study therapy, will be  
monitored until:    
• 
the adverse event resolves or the symptoms or signs that constitute the adverse 
event   
return to baseline; any abnormal laboratory values have returned to baseline;    
• 
there is a satisfactory explanation other than the study therapy for the changes   
• 
observed; or  death.   
 
[Inserted from Adverse Event Definitions and Reporting Section – Appendix IB SCCC  
DSMC Plan]   
 
 
  
8.1.1    
Definition    
The study will use the CTCAE version 5.0 for reporting. Adverse Events will be reported as   

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indicated by the appropriate following table (see below).      
An adverse event is defined as any untoward or unfavorable medical occurrence in a human   
research study participant, including any abnormal sign (for example, abnormal physical exam or  
laboratory finding), symptom, clinical event, or disease, temporarily associated with the subject’s  
participation in the research, whether or not it is considered related to the subject’s participation   
in the research.   
Adverse events encompass clinical, physical and psychological harms. Adverse events occur   
most commonly in the context of biomedical research, although on occasion, they can occur in   
the context of social and behavioral research. Adverse events may be expected or unexpected.   
Adverse event & Serious adverse event will only be collected till the last day of Letrozole dose   
taken.   
Severity    
Adverse events will be graded by a numerical score according to the defined NCI Common   
Terminology Criteria for Adverse Events (NCI CTCAE) and version number specified in the   
protocol. Adverse events not specifically defined in the NCI CTCAE will be scored on the   
Adverse Event log according to the general guidelines provided by the NCI CTCAE and as   
outlined below.     
• 
Grade 1: Mild    
• 
Grade 2: Moderate   
• 
Grade 3: Severe or medically significant but not immediately life threatening   
Grade 4: Life threatening consequences   
• 
Grade 5: Death related to the adverse event   
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
 
Serious Adverse Events    
ICH Guideline E2A and the UTSW IRB define serious adverse events as those events, occurring  
at any dose, which meets any of the following criteria:    
• 
Results in death     
• 
Immediately life-threatening    
• 
Results in inpatient hospitalization or prolongation of existing hospitalization    
• 
Results in persistent or significant disability/incapacity    
• 
Results in a congenital anomaly/birth defect    
• 
Based upon appropriate medical judgment, may jeopardize the subject’s health and may   
require medical or surgical intervention to prevent one of the other outcomes listed in this   
definition.   
Note: A “Serious adverse event” is by definition an event that meets any of the above criteria.   
Serious adverse events may or may not be related to the research project. A serious adverse event  
determination does not require the event to be related to the research. That is, both events   
completely unrelated to the condition under study and events that are expected in the context of   
the condition under study may be serious adverse events, independent of relatedness to the study  
itself. As examples, a car accident requiring overnight hospitalization would be a serious adverse  
event for any research participant; likewise, in a study investigating end-stage cancer care, any   
hospitalization or death which occurs during the protocol-specified period of monitoring for   

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adverse and serious adverse events would be a serious adverse event, even if the event observed   
is a primary clinical endpoint of the study.   
  
8.1.2 Unanticipated Problems Involving Risks to Subjects or Others (UPIRSOs):   
The term “unanticipated problem” is found, but not defined in the regulations for the Protection   
of Human Subjects at 45 CFR 46, and the FDA regulations at 21 CFR 56. Guidance from the   
regulatory agencies considers unanticipated problems to include any incident, experience, or   
outcome that meets each of the following criteria:   
• 
Unexpected (in terms of nature, severity or frequency) AND   
• 
Definitely or probably related to participation in the research AND    
• 
Serious or a possible unexpected problem in that the research places subjects or others at   
greater risk of harm than was previously known or recognized. Note: Any serious adverse   
event would always suggest a greater risk of harm.   
Follow-up    
All adverse events will be followed up according to good medical practices.    
    
  
8.1.3    
Reporting   
The UTSW IRB requires reporting of all UPIRSOs according to the guidance below. For   
participating centers other than UTSW, local IRB guidance should be followed for local   
reporting of serious adverse events. All SAEs occurring during the protocol-specified monitoring   
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
period should be submitted to the UTSW study team within 2 business days of the center  
learning of the event.   
 
UPIRSOs occurring on the study require expedited reporting, and are submitted to the UTSW  
IRB through the UTSW eIRB by the UTSW study team and to the SCCC DSMC Coordinator.   
Hardcopies or electronic versions of the eIRB report; FDA Form #3500A forms, or other sponsor  
forms, if applicable; and/or any other supporting documentation available should be submitted to  the 
UTSW study team and will be forwarded to the DSMC Coordinator.  The DSMC  
Coordinator forwards the information onto the DSMC Chairman who determines if immediate  
action is required.  Follow-up eIRB reports, and all subsequent SAE documentation that is  
available are also submitted to the DSMC Chair who determines if further action is required.  
(See Appendix IV of the SCCC DSMC Plan for a template Serious Adverse Event Form which  
may be utilized when a sponsor form is unavailable and SAE submission to the eIRB is not  
required).   
    
All serious adverse events which occur on research subjects on protocols for which the SCCC is   
the DSMC of record require reporting to the DSMC regardless of whether IRB reporting is   
required. Hardcopies or electronic versions of the FDA Form #3500A forms, or other sponsor   
forms, if applicable; and/or any other supporting documentation available should be forwarded to  
the DSMC Coordinator.     
  
   
If the event occurs on a multi-institutional clinical trial coordinated by the UTSW Simmons   
Cancer Center, the DOT Manager or lead coordinator ensures that all participating sites are   
notified of the event and resulting action, according to FDA guidance for expedited reporting.   
DSMC Chairperson reviews all serious adverse events upon receipt from the DSMC   

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Coordinator.  The DSMC Chairperson determines whether action is required and either takes   
action immediately, convenes a special DSMC session (physical or electronic), or defers the   
action until a regularly scheduled DSMC meeting.    
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
UTSW Institutional Review Board (IRB)   
Submit via eIRB with a copy of the final sponsor report as attached supporting   
documentation   
 
1.  
SAEs   
Serious adverse events (SAEs) for studies where the SCCC DSMC is the DSMC of record  
require reporting to the DSMC coordinator within 2 working days of PI awareness, or as   
described in the protocol.     
 
2. 
Unanticipated Problems Involving Risks to Subjects or Others (UPIRSOs)   
Local Serious Adverse Event UPIRSOs require reporting to the UTSW IRB within 48 hours of  PI 
awareness of the event (life threatening or fatal events experienced by subjects enrolled by the  
investigator(s) under UTSW IRB jurisdiction).     
Local UPIRSOs (non-serious events experienced by subjects enrolled by the investigator(s)   
under UTSW IRB jurisdiction) require reporting to the UTSW IRB within 5 business days of PI  
awareness of the event.   
External UPIRSOs including those that occur as non-local events require reporting to the UTSW  
IRB within 10 working days of PI awareness of the event.   
For further guidance for Investigators regarding safety reporting requirements for INDs and   
BA/BE studies, refer to FDA Draft Guidance document:   
http://www.fda.gov/downloads/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/U 
CM227351.pdf   
Telephone reports to:   
Investigator: Nisha Unni     
Telephone Number: 214-648-4180   
UTSW SCC Data Safety Monitoring Committee Coordinator (if fax report is not   
available) within 1 working day to 214-648-7097.   
Written reports to:   
Investigator: Nisha Unni   
Email: nisha.unni@utsouthwestern.edu   
UTSW SCC Data Safety Monitoring Committee Coordinator  
Email: SCCDSMC@utsouthwestern.edu   
Fax: 214-648-5949 or deliver to BLB.306   

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8.2 
Steps to Determine If an Adverse Event Requires Expedited Reporting   
Step 1: Identify the type of adverse event using the NCI Common Terminology Criteria for  
Adverse Events (CTCAE v5).    
Step 2: Grade the adverse event using the NCI CTCAE v5.   
 
Step 3: Determine whether the adverse event is related to the protocol therapy    
Attribution categories are as follows:   
- 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.   
- Unrelated – The AE is clearly NOT related to the study treatment.   
Note: This includes all events that occur within 30 days of the last dose of protocol treatment.   
Any event that occurs more than 30 days after the last dose of treatment and is attributed   
(possibly, probably, or definitely) to the agent(s) must also be reported accordingly.   
 
 
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________________________________________________________________________   
Step 4: Determine the prior experience of the adverse event.    
Expected events are those that have been previously identified as resulting from administration  
of the agent. An adverse event is considered unexpected, for expedited reporting purposes only,  
when either the type of event or the severity of the event is not listed in:   
- The current known adverse events listed in the Agent Information Section of this   
protocol;   
- The drug package insert;   
- The current investigator’s Brochure  
  
9.0       Dose Modifications and Management of Toxicity   
Toxicity grading is based on the NCI-CTC Version 5.0. All NCI CTCAE grade 3 or 4 toxicities will  
result in Letrozole interruption.   
9.1  Dose Reductions for Letrozole   
There will be no dose reductions of letrozole. If a subject is required to stop treatment with letrozole   
for more than 3 days, the Principal Investigator must be consulted. Letrozole must be taken  
continuously for a minimum of 7-30 days.   
9.2  Other   
Other serious adverse events or grade 3 or 4 toxicity considered to be related to letrozole should be   
managed with symptomatic support and letrozole discontinued.   
 
10.     Drug Information   
Other Names: Femara   
Classification: Non-steroidal aromatase inhibitor (inhibitor of estrogen synthesis)   
Mode of Action: Aromatase inhibitors inhibit the peripheral and intra-tumoral enzyme p450  
aromatase, which catalyzes the final step in estrogen synthesis. Depriving ER of its ligand is an  
effective endocrine therapy. These agents have the advantage of potentially blocking genomic and  non-
genomic ER functions.   
Storage and Stability: Letrozole is stored at room temperature.   
Dose:  2.5 mg by mouth once a day for 7 – 30 days  

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Route of Administration:  Oral. Tablets should be taken preferably in the morning with up to 200  ml 
of water and should be taken 1 hour before or 2 hours after meals. Patients who are unable to  
swallow tablets may dissolve the tablets in distilled water for administration.   
Incompatibilities:  None known.   
Availability: Patients will be given a prescription for letrozole 2.5 mg to be taken by mouth daily    
Interactions: Treatment of women with letrozole significantly lowers serum estrone, estradiol and  
estrone sulfate and has not been shown to significantly affect adrenal corticosteroid synthesis,  
aldosterone synthesis, or synthesis of thyroid hormones. Clinical interaction studies with cimetidine  
and warfarin indicated that the co-administration of letrozole with these drugs does not result in  
clinically- significant drug interactions.   
 
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Study Number – SCCC-11118   
________________________________________________________________________   
 
11.0    Removal of Patients from Protocol Therapy   
            11.1 Disease Progression   
Contra-lateral breast cancer is not considered disease progression. If the patient progresses during  
therapy with letrozole, she should proceed to surgery and be taken off letrozole and treated at the  
discretion of the treating physician. To our knowledge, this has never been reported to occur after  such 
short treatment with an aromatase inhibitor like letrozole.   
 
11.2 Extraordinary Medical Circumstances:     
If at any time the constraints of this protocol are detrimental to the patient’s health, letrozole should   
be discontinued. In this event:   
• 
Notify the principal investigator in writing   
• 
Document reason(s) for treatment termination on the flow sheets   
 
11.3 If patient wishes to be taken off study   
11.4 If the patient does not recover from toxicity in a timely fashion as outlined in section    
11.5 Patient non-compliance with protocol   
.   
12.0 
Statistical Considerations   
The main objective of this Aim is to use the Ki67 index measured 7 – 30days after therapy with  
letrozole to segregate hormone receptor-positive breast cancers that are highly hormone-dependent  vs. 
those that are not. For this analysis, we had access to the raw Ki67 data from both the IMPACT  trial36 
(provided by Mitch Dowsett, Royal Marsden Hospital, London) and from the P024 trial22  (provided 
by Matt Ellis). With these data, we initially calculated that with 125 ER+ patients, 33%  or 42 of them 
will exhibit a ≤1% Ki67 after 2 weeks of treatment with the aromatase inhibitor. This  will provide at 
least 90% power to detect a 2-fold difference in gene expression level between ≤1%  Ki67 (n=42) and 
>1% Ki67 (n=83) based on a conservative assumption that the standard deviation  is 0.7 (base-two 
logarithmic scale) and the acceptable number of false positives is 0.01. In addition,  40 more patients 
will be enrolled to test the accuracy of the statistical model that will be developed  based on the initial 
125 patients. Therefore, a total sample size of 165 will not only yield an  excellent power but 
also have the ability to examine the model accuracy.   
Dr. Carlos L. Arteaga, conducted a similar study that so far has enrolled  over 200 patients34. 
Participation in this study has not resulted in any side effects and/or delays in  surgery. Preliminary 

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analysis of the molecular data is already identifying novel potential  mechanism of resistance 
to estrogen deprivation in ER+ breast cancer. We anticipate further  discoveries with expansion 
of this trial as well as a source of tumors that we can use for validation  of the initial findings.    
Estimated Annual Accrual:  Combined, UTSW Simmons Cancer Center and Parkland Hospital  
oncology clinic sees approximately 300-350 hormone-receptor positive breast cancer patients/year.  
This represents approximately 10% increase since 2004. We anticipate an annual growth rate of  
approximately 5%/year over the next 3 years. Eighty percent are patients with stage I or II breast   
 
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Study Number – SCCC-11118   
________________________________________________________________________   
cancers. We estimate that 1-2 patients per week will be potential candidates for this trial. Patient  
accrual will be monitored closely by the Disease Oriented Team (DOT).   
For gene expression level and/or a mutational or proteomic signature parameters, we will use   
ANOVA to evaluate differences among three or more groups and unpaired two-sample t-test for   
differences between two groups stratified based on post-treatment Ki67 index. If underlying   
assumptions of these methods (e.g., normality and homoscedasticity) are not satisfied, we will   
use the nonparametric Kruskal-Wallis ANOVA for three or more group comparisons and the   
nonparametric Wilcoxon rank sum test for two group comparisons. Multiple comparisons will be   
adjusted. P <0.05 will be considered statistically significant. We will also split the data into   
training and validation datasets, and compute the prediction error to assess the model accuracy   
with cross validation.    
 
 
13.0     Data Safety    
This study will not provide direct clinical benefit to patient participants. Its purpose is to establish   
molecular profiles that will stratify ER positive breast cancers according to antiestrogen resistance   
and risk of recurrence. The data generated by this study will help the design of future clinical trials  
and may offer benefit to patients enrolled in those controlled trials. This is explained in the consent  
form. We should note that in all the patients enrolled in this study, the FDA-approved standard of   
care will be at least 5 years of post-operative adjuvant letrozole – or another aromatase inhibitor.   
Risks to subjects will be minimized by screening to assure appropriate selection of participants,  
strict inclusion and exclusion criteria, close monitoring, and safety monitoring and reporting.  The  
procedures are consistent with sound research design.   
 
We are proposing to use well-tolerated doses of letrozole in a very short time interval. Therefore, it  is 
almost unheard of that we will encounter unacceptable toxicity from administration of this drug.  We 
should emphasize that, based on current standards of care, all patients enrolled in this trial, will  meet 
criteria for adjuvant letrozole for 5 years.    
After signing consent, all patients must pass the screening procedure (detailed in the eligibility   
criteria in the protocol) to be eligible for the study. Every effort will be made to instruct the   
patient of the potential side effects of the treatment and of measures to mitigate them.   
Appropriate dose modifications of standard and study drugs secondary to toxicities are detailed   
in the protocol. The importance of notifying the principal investigator, the research nurse, or   
treating physician of ANY adverse event will be stressed. All serious adverse events will be   
promptly reported to all participating sites IRB and sponsoring institutions as outlined in the   
protocol.    
The Principal Investigator or designee will inform the IRB of any serious adverse event in   
accordance with the institution’s IRB policy.  The investigator is responsible for the detection   

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and documentation of events meeting the criteria and definition of an AE or SAE, as provided in   
this protocol.  During the study when there is a safety evaluation, the investigator or site staff   
will be responsible for detecting, documenting, and report AEs and SAEs, as detailed in the   
protocol.  Furthermore, the investigator will be required to provide periodic safety updates on the   
conduct of the study at his/her site and notification of study closure to the participating sites IRB.    
 
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Study Number – SCCC-11118   
________________________________________________________________________   
This study will be conducted in accordance with "good clinical practice" (GCP) and all  
applicable regulatory requirements, including, where applicable, the 1996 version of the  
Declaration of Helsinki. The investigator is responsible for ensuring that this protocol, the site’s  
informed consent document, and any other information that will be presented to potential  
subjects (e.g., advertisements or information that supports or supplements the informed consent)  are 
reviewed and approved by the appropriate IEC/IRB.  The investigator agrees to allow the  
IEC/IRB direct access to all relevant documents. The IEC/IRB must be constituted in accordance  
with all applicable regulatory requirements.     
 
14.0 Monitoring and Quality Assurance   
Trial monitoring will be conducted no less than annually and refers to a regular interval review  
of trial related activity and documentation performed by the DOT, which includes but is not   
limited to accuracy of case report forms, protocol compliance, timeless and accuracy of Velos  
entries and AE/SAE management and reporting. Documentation of trial monitoring will be   
maintained along with other protocol related documents and will be reviewed during internal   
audit.     
 
The UTSW Simmons Cancer Center (SCC) Data Safety Monitoring Committee (DSMC) is   
responsible for monitoring data quality and patient safety for all UTSW SCC clinical trials.  As  
part of that responsibility, the DSMC reviews all local serious adverse events and unanticipated  
problems in real time as they are reported and reviews adverse events on a quarterly basis.  The  
quality assurance activity for the Clinical Research Office provides for periodic auditing of   
clinical research documents to ensure data integrity and regulatory compliance.  A copy of the  
DSMC plan is available upon request.    
 
The SCC DSMC meets quarterly and conducts annual comprehensive reviews of ongoing   
clinical trials, for which it serves as the DSMC of record. The QAC works as part of the DSMC   
to conduct regular audits based on the level of risk. Audit findings are reviewed at the next   
available DSMC meeting.  In this way, frequency of DSMC monitoring is dependent upon the   
level of risk.  Risk level is determined by the DSMC Chairman and a number of factors such as   
the phase of the study; the type of investigational agent, device or intervention being studied; and  
monitoring required to ensure the safety of study subjects based on the associated risks of the   
study. Protocol-specific DSMC plans must be consistent with these principles.    
 
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Study Number – SCCC-11118   
________________________________________________________________________   
15.0   Changes in Protocol    
Any change to the informed consent document must be reviewed and approved by the Principal 

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Investigator before being submitted to the Institutional Review  Board/Independent Ethics 
Committee at participating institutions. Amendments should not be  implemented until all 
necessary approvals have been obtained, except when necessary to  eliminate an immediate 
hazard to study subjects.   
 
16.0    Protocol Deviations   
The Principal Investigator is responsible for implementing and maintaining quality  assurance and 
quality control to ensure that studies are conducted according to the protocol,  GCP, and all 
applicable regulatory requirements. A protocol deviation is any noncompliance  with the protocol. 
Noncompliance can be on the part of the study participant, the Investigator, or  the study site staff. 
Deviations to the protocol are not permitted except when necessary to  eliminate an immediate 
hazard to study subjects.   
 
17.0    Data Management   
 
REDCap is the UTSW SCCC institutional choice for the electronic data capture of case report  
forms for this and all SCCC Investigator Initiated Trials. REDCap will be used for electronic  
case report forms in accordance with Simmons Comprehensive Cancer Center requirements.   
 
Trial monitoring will be conducted no less than annually and refers to a regular interval review  
of trial related activity and documentation performed by the DOT, which includes but is not   
limited to accuracy of case report forms, protocol compliance, timeless and accuracy of Velos  
entries and AE/SAE management and reporting. Documentation of trial monitoring will be   
maintained along with other protocol related documents and will be reviewed during internal   
audit.    
 
The UTSW Simmons Comprehensive Cancer Center (SCCC) Data Safety Monitoring 
Committee (DSMC) is responsible for monitoring data quality and patient safety for all UTSW 
SCCC clinical trials.  As part of that responsibility, the DSMC reviews all serious adverse 
events and UPIRSOs in real time as they are reported and reviews adverse events on a quarterly 
basis.  The quality assurance activity for the Clinical Research Office provides for periodic 
auditing of clinical research documents to ensure data integrity and regulatory compliance.  A 
copy of the DSMC plan is available upon request.  
 
 
The SCCC DSMC meets quarterly and conducts annual comprehensive reviews of ongoing 
clinical trials, for which it serves as the DSMC of record. The Quality Assurance Coordinator 
(QAC) works as part of the DSMC to conduct regular audits based on the level of risk. Audit 
findings are reviewed at the next available DSMC meeting.  In this way, frequency of DSMC 
monitoring is dependent upon the level of risk.  Risk level is determined by the DSMC 
Chairman and a number of factors such as the phase of the study; the type of investigational 
agent, device or intervention being studied; and monitoring required to ensure the safety of 
study subjects based on the associated risks of the study. Protocol-specific DSMC plans must be 
consistent with these principles 
 
18.0      Data Verification   
The Principal Investigator is responsible for the conduct of the clinical trial at the site in  
accordance with Title 21 of the Code of Federal Regulations and/or the Declaration of Helsinki.  The 
Principal Investigator is responsible for personally overseeing the treatment of all study  patients. 

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The Principal Investigator must assure that all study site personnel, including sub- investigators 
and other study staff members, adhere to the study protocol and all FDA/GCP/NCI  regulations and 
guidelines regarding clinical trials both during and after study completion.   
The Principal Investigator at each institution or site will be responsible for assuring that all the  
required data will be collected and entered onto the Case Report Forms. Periodically, monitoring  
visits may be conducted and the Principal Investigator will provide access to his/her original   
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
records to permit verification of proper entry of data. At the completion of the study, all case  
report forms will be reviewed by the Principal Investigator and will require his/her final  
signature to verify the accuracy of the data.   
 
Data will be collected via paper CRFs and entered into the Redcap database. The study team will  
check data accuracy by performing source data verification. Source data verification is a direct  
comparison of the entries made on the CRFs against the appropriate source documentation.  
Discrepancies in the data will be brought to the attention of the Investigator and/or the  
Investigator’s staff.  Any necessary corrections will be made directly to the CRFs or via queries  by 
the Investigator and/or the Investigator’s staff.   
 
19.0 Adherence to the Protocol 
Except for an emergency situation, in which proper care for the protection, safety, and well-being of 
the study subject requires alternative treatment, the study shall be conducted exactly as described in 
the approved protocol.  
Exceptions (also called single-subject exceptions or single-subject waivers): include any departure 
from IRB-approved research that is not due to an emergency and is: 
• intentional on part of the investigator; or 
• in the investigator’s control; or 
• not intended as a systemic change (e.g., single-subject exceptions to eligibility 
[inclusion/exclusion] criteria) 
 Reporting requirement*: Exceptions are non-emergency deviations that require 
prospective IRB approval before being implemented. Call the IRB if your request is 
urgent. If IRB approval is not obtained beforehand, this constitutes a major deviation. For 
eligibility waivers, studies which utilize the SCCC-DSMC as the DSMC of record must 
also obtain approval from the DSMC prior to submitting to IRB for approval.  
Emergency Deviations: include any departure from IRB-approved research that is necessary to:  
• avoid immediate apparent harm, or 
• protect the life or physical well-being of subjects or others 
Reporting requirement*: Emergency deviations must be promptly reported to the IRB 
within 5 working days of occurrence. 
  

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Serious Noncompliance (formerly called major deviations or violations): include any departure 
from IRB-approved research that: 
• 
Increase risk of harm to subjects; and/or 
• 
 
adversely affects the rights, safety, or welfare of subjects (any of which may also be an 
unanticipated problem); and/or  
• 
adversely affects the integrity of the data and research (i.e., substantially compromises the 
integrity, reliability, or validity of the research) 
Reporting requirement*: Serious Noncompliance must be promptly reported tothe IRB 
within 5 working days of discovery. 
 
Continuing Noncompliance: includes a pattern of repeated    noncompliance (in  or more protocols 
simultaneously, or over a period of time) which continues after initial discovery, including 
inadequate efforts to take or implement corrective or preventive action within a reasonable 
time frame. 
Reporting requirement*: Continuing Noncompliance must be promptly reported to  
the IRB within 5 working days of discovery. 
 
Noncompliance (that is neither serious nor continuing; formerly called minor deviations) any 
departure from IRB-approved research that: 
 
• Does not meet the definition of serious noncompliance or continuing noncompliance 
Reporting requirement*: Noncompliance that is neither serious nor continuing  should be 
tracked and summarized the next IRB continuing review, or the notice of study closure- 
whichever comes first.. 
 
*Reporting Requirements reflect UTSW HRPP/IRB guidelines; participating sites should follow the 
reporting guidelines for their IRB of record 
 
20.0    Study Documentation    
Pre-Study Documentation   
o Prior to initiating the trial, the study team will collect all relevant regulatory 
documents,  including but not limited to the following documents:   
o A current curriculum vitae of the Principal Investigator and each sub-investigator listed on   
the FDA Form 1572   
o A copy of the current medical licenses of the Principal Investigator and all sub-investigators   
listed on the FDA 1572   
o A copy of the Protocol Acceptance Page signed and dated by the Principal Investigator   
o A letter from the IRB stipulating approval of the protocol, the informed consent document   
and any other material provided to potential trial participants with information about the trial   
(e.g. advertisements)   
o A copy of the informed consent document approved by both the sponsor and local IRB   
o The current IRB Federalwide Assurance Letter   
o Current laboratory certification for the reference laboratory and curriculum vitae of the   
laboratory director   
o A list of current laboratory normal values for the reference laboratory   
o A copy of the signed Delegation of Authority Log   
o Protocol training documentation for study staff    
o The drug destruction SOP    

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o A copy of the fully executed contract must be on record    
 
 
Required regulatory documents include, but are not limited to the following: local IRB approvals  
(i.e., protocol, consent form, amendments, patient brochures, recruitment material, etc.), each  
participant’s informed consent, enrollment form, eligibility checklist and tissue block  
registration, summary of unanticipated problems or protocol deviations, and documentation of  
expertise of the investigators.    
 
 
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
 
21.0     Records Retention   
Study documentation includes all Case Report Forms, data correction forms or queries, source  
documents, Sponsor-Investigator correspondence, monitoring logs/letters, and regulatory  
documents (e.g., protocol and amendments, IRB correspondence and approval, signed patient  
consent forms).    
 
Source documents include all recordings of observations or notations of clinical activities and all  
reports and records necessary for the evaluation and reconstruction of the clinical research study.   
 
Government agency regulations and directives require that the study investigator retain all study  
documentation pertaining to the conduct of a clinical trial. In the case of a study with a drug  
seeking regulatory approval and marketing, these documents shall be retained for at least two  
years after the last approval of marketing application in an International Conference on  
Harmonization (ICH) region. In all other cases, study documents should be kept on file until  
three years after the completion and final study report of this investigational study.   
Following closure of the study, the site will maintain a copy of all site study records in a safe and  
secure location. Records may be destroyed according to SCCC and institutional policies.   
 
22.0      Confidentiality   
Names and records of patients taking part in this study will remain confidential. Members of the  
research team will comply with appropriate HIPAA guidelines. Records that contain patient  
identifiers will be available only to the physicians, nurses, and other health care personnel directly  
involved in the management of the patient. Data that include a unique patient identifier code, but  
not patient name or other information that could jeopardize patient privacy, will be available to  
research personnel involved in the study. Data will be stored in secured location in a password  
protected data base.   
 
The investigator agrees to keep all information provided by this study in strict confidence and to  
request similar confidentiality from her staff and the IRB/IEC/REB. Study documents provided  
(protocols, investigators' brochures, CRFs and other material) will be stored appropriately to ensure  
their confidentiality.    
 

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Patient medical information obtained by this study is confidential, and disclosure to third parties  
other than those noted below is prohibited. Upon the patient’s permission, medical information may  
be given to his or her personal physician or other appropriate medical personnel responsible for his  or 
her welfare.  This medical information must be made available to the IRB and DSMC, upon  
request, for source verification of study documentation.  Data generated by this study must be  
available for inspection upon request by representatives of the U.S. FDA, local health authorities,  
Study Chair, principal Investigator and their authorized representative(s), collaborators and  
licensees, and the IRB for each study site, if appropriate.   
 
 
23.0 
Obligations of Investigators 
The Principal Investigator is responsible for the conduct of the clinical trial at the site in accordance 
with Title 21 of the Code of Federal Regulations and/or the Declaration of Helsinki. The Principal 
Investigator is responsible for personally overseeing the treatment of all study patients. The Principal 
Investigator must assure that all study site personnel, including sub-investigators and other study staff 
members, adhere to the study protocol and all FDA/GCP/NCI regulations and guidelines regarding 
clinical trials both during and after study completion. 
 
 
 
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
24.0 References   
1. Tsai, M.J. & O'Malley, B.W. Molecular mechanisms of action of steroid/thyroid receptor   
superfamily members. Annu Rev Biochem 63, 451-86 (1994).   
2. Liu, M.M., Albanese, C., Anderson C.M. et al. Opposing actions of estrogen receptors    
and on cyclin D1 gene expression. J. Biol Chem. 277,24353-24360 (2002).   
3. Pettersson, K., Delauney, F., Gustafsson, J.A. Estrogen receptor b acts as a dominant regulator   
of estrogen signaling. Oncogene 19, 4970-4978 (2000).   
4. Nilsson, S. et al. Mechanisms of estrogen action. Physiol Rev 81, 1535-1565 (2001).   
5. O'Lone, R., Frith, M.C., Karlsson, E.K. & Hansen, U. Genomic targets of nuclear estrogen   
receptors. Mol Endocrinol 18, 1859-75 (2004).   
6. Losel, R. & Wehling, M. Nongenomic actions of steroid hormones. Nat Rev Mol Cell Biol 4,   
46-56 (2003).   
7. Bjornstrom, L. & Sjoberg, M. Mechanisms of estrogen receptor signaling: convergence of   
genomic and nongenomic actions on target genes. Mol Endocrinol 19, 833-42 (2005).   
8. Osborne, C.K. & Schiff, R. Estrogen-receptor biology: continuing progress and therapeutic   
implications. J Clin Oncol 23, 1616-22 (2005).   
9. Mouridsen H, Gershanovich M, Sun Y, et al. Superior efficacy of letrozole versus tamoxifen   
as first-line therapy for postmenopausal women with advanced breast cancer: results of a   
phase III study of the International Letrozole Breast Cancer Group. J Clin Oncol 19(10):2596-  
606 (2001).   
10. Come SE, Buzdar AU, Ingle JN, et al. Proceedings of the Fifth International Conference on   
Recent Advances and Future Directions in Endocrine Therapy for Breast Cancer: conference   
summary statement. Clin Cancer Res 12(3 Pt 2):997s-1000s (2006).   
11. Baum M, Budzar AU, Cuzick J, et al. Anastrozole alone or in combination with tamoxifen   
versus tamoxifen alone for adjuvant treatment of postmenopausal women with early breast   

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cancer: first results of the ATAC randomized trial. Lancet 359(9324):2131-9 (2002).    
12. Howell A., Cuzick J, Baum M, et al. Results of the ATAC (Arimidex, Tamoxifen, Alone or in   
Combination) trial after completion of 5 years' adjuvant treatment for breast cancer.   
Lancet;365(9453):60-62 (2005).   
13. Goss PE, Ingle JN, Martino S, et al. A randomized trial of letrozole in postmenopausal women   
after five years of tamoxifen therapy for early-stage breast cancer. N Engl J Med   
349(19):1793-802 (2003).   
14. Goss PE, Ingle JN, Martino S, et al. Randomized trial of letrozole following tamoxifen as   
extended adjuvant therapy in receptor-positive breast cancer: updated findings from NCIC   
CTG MA.17. J Natl Cancer Inst 97(17):1262-71 (2005).   
15. Cole P.A., Robinson, C.H. .Mechanism and inhibition of cytochrome p450 aromatase. J Med   
Chem 33; 2933-2944 (1990).   
16. Nemere I, Pietras RJ, Blackmore PF membrane receptors for steroid hormones, signal   
transduction and physiologic significance J Chem Biochem88:438-445 (2003).   
17. Massarweh S, Schiff, R Resistance to endocrine therapy in breast cancer: exploiting estrogen   
receptor/growth factor signaling crosstalk. Endocr Relat Cancer. 13: S15-S24 (2006).    
18. Schiff, R, Massarweh, S, Shou, J, Osborne CK. Breast cancer endocrine resistance: how   
growth factor signaling and estrogen receptor coregulators modulate response. Clin Cancer   
Res 83: 447S-454s, (2003).   
 
 
 
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Study Number – SCCC-11118   
________________________________________________________________________   
19. Smith CL, Nawaz Z, O’Malley BW, et al. Coactivator and corepressor activation of the   
agonist/antagonist activity of the mixed antiestrogen trans-4-hydroxy-tamoxifen. Mol   
Endocrinol. 11:657-666. (1997).   
20. Lavinsky RM, Jepsen K, Heinzel T et al.  Diverse signaling pathways modulate nuclear   
receptor recruitment of N-CoR and SMRT complexes. Proc Natl Acad  Sci. USA 95:2920-  
2925 (1998)   
21. Kurokawa, H. et al. Inhibition of HER2/neu (erbB-2) and mitogen-activated protein kinases   
enhances tamoxifen action against HER2-overexpressing, tamoxifen- resistant breast cancer   
cells. Cancer Res 60, 5887-94. (2000).   
22. Mitch Dowsett, Ian E. Smith, Steve R. Ebbs, et al  Short-term changes in Ki-67 during   
neoadjuvant treatment of primary breast cancer with anastrozole or tamoxifen alone or   
combined correlate with recurrence-free survival.  Clin Can Res 11: 951s-958s, (2005)   
23. Dowsett, M., et al. Biomarker changes during neoadjuvant anastrozole, tamoxifen, or the   
combination: influence of hormonal status and HER-2 in breast cancer--a study from the   
IMPACT trialists. J Clin Oncol 23, 2477-92 (2005).   
24. Baum M, Buzdar A, Cuzick J, et al. Anastrozole alone or in combination with tamoxifen   
versus tamoxifen alone for adjuvant treatment of postmenopausal women with early-stage   
breast cancer: results of the ATAC (Arimidex, Tamoxifen Alone or in Combination) trial   
efficacy and safety update analyses. javascript:AL_get(this, 'jour', 'Cancer.'); Cancer   
98(9):1802-1810 2003.    
25.  Howell A, Cuzick J, Baum M, et al. Results of the ATAC (Arimidex, Tamoxifen, Alone or in   
Combination) trial after completion of 5 years' adjuvant treatment for breast cancer. Lancet   
365 (9453):60-2. (2005).   
26. Cronin, M. et al. Measurement of gene expression in archival paraffin-embedded tissues:     
development and performance of a 92-gene reverse transcriptase-polymerase chain reaction   
assay. Am J Pathol 164, 35-42 (2004).   

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27. Paik, S. et al. A multigene assay to predict recurrence of tamoxifen-treated, node-negative   
breast cancer. N Engl J Med 351, 2817-26 (2004).   
28. Paik, S. et al. Gene expression and benefit of chemotherapy in women with node-negative,   
estrogen receptor-positive breast cancer. J Clin Oncol 24, 3726-34 (2006).   
29. Habel L.A., Shak S., Jacobs K. et al. A population based study of tumor gene expression and   
risk of breast caner death among lymph node negative patients. Breast Cancer Res 8 (3): R25   
(2006).   
30. Cronin, M., Sangli, C. Liu M.L. et al. Analytical validation of the Oncotype DX genomic   
diagnostic test for recurrence prognosis and therapeutic response prediction in node-negative,   
estrogen receptor-positive breast cancer. Clin Chem 53(6):1084-91 (2007).   
31. Fan, C., Oh D.S., Wessels L., et al. Concordance among gene-expression-based predictors for   
breast cancer. N Engl J Med 355, 560-9 (2006).   
32. Ellis, M.J., Tao, Y., Bhatnagar, A.S., et al. Cell cycle complete response after neoadjuvant   
letrozole predicts superior relapse-free and overall survival: Long-term follow-up of the   
letrozole P024 study. Proc. Am. Soc. Clin. Oncol. 25 (18S) (2007).   
33. Mackay, A. Urruticoechea A, Dixon JM, et al. Molecular response to aromatase inhibitor   
treatment in primary breast cancer. Breast Cancer Res 9, R37 (2007).   
34. Giltnane JM, Hutchinson KE, Stricker TP, Formisano L, Young CD, Estrada MV, Nixon   
MJ, Du L, Sánchez V, Gonzalez-Ericsson P, Kuba MG, Sanders ME, Mu XJ, Van Allen   
EM, Wagle N, Mayer IA, Abramson VG, Gómez H, Rizzo M, Toy W, Chandarlapaty S,   
 
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Study Number – SCCC-11118   
________________________________________________________________________   
Mayer EL, Christiansen J, Murphy D, Fitzgerald K, Wang K, Ross JS, Miller VA, Stephens  
PJ, Yelensky R, Garraway L, Meszoely I, Balko JM, Arteaga CL. Genomic profiling of  
ER+ breast cancers after short term estrogen suppression reveals alterations associated with  
endocrine resistance. Sci. Transl. Med. 2017 Aug 9; 9(402). pii: eaai7993. doi:  
10.1126/scitranslmed.aai7993. PMID 28794284   
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Study Number – SCCC-11118   
________________________________________________________________________   
 
Appendix I: Lab Manual   
BLOOD SAMPLES:    
1. 1 EDTA tube (10ml purple top tube)   
Collection: Blood sample will be collected by the CRO staff.   
Processing: Farjana Fattah lab group will process these samples for buffy coat. Processed within  
6 hours of collection. SOP for buffy coat processing:   
o 
Centrifuge the tube at 1100-1300g for 10 minutes at 4 degrees Celsius.   
o 
Discard the plasma layer.   
o 
Collect buffy coat into one labelled 2mL cryo tube and -80 degrees Celsius storage   
Storage: Farjana’s team will store these samples after processing and Saurabh Mendiratta will  
coordinate and collect from them from time to time.   
 
TISSUE (Punch Biopsies):   
1. 1-6 core biopsy OR 10 unstained slides   
Collection: Cheryl Lewis group will collect the tissue sample.   
o 
First four (1-4) cores would be completely flash frozen (no FFPE).   
o 
If there is a fifth core, it should be formalin fixed.   
Sixth core, if 
available, should be flash frozen.   
Instructions on fresh tissue specimen handling:   
o 
According to the following schedule, core biopsy specimens should be immediately flash   
frozen in liquid nitrogen* OR placed in 10% neutral buffered formalin immediately   
following removal from the patient following procedures established by the Simmons   
Cancer Center Tissue Core.    
o 
For the formalin-fixed cores, an approximately 20:1 volume of formalin to tissue should be   
used.  Core biopsies placed in 10% neutral buffered formalin must fix at room temperature   
for at least 6 h and no more than 24 h.  Core biopsies must be transferred from formalin to   
70% ethanol after 6-24 h to prevent over fixation and placed in the refrigerator.   
o 
Label ALL cryovials before freezing. Label each cryovial and formalin-filled collection   
container with the patient name, Medical Record #, Patient study ID #, location of biopsy,   
collection date and time of collection using an ethanol resistant marker.  Each set of   
patient samples should be accompanied by a Tissue Registration form.    

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Storage:   
Dr. Lewis group will store them and once these boxes will be full, all samples will be sent to   
Saurabh Mendiratta, 6001 Harry Hines Blvd. NB5.112. Dallas, TX- 75390-8807 Phone: 214-648-6167.  
It will be stored in Dr. Arteaga lab freezer.   
 
 
 
 
 
 
 
 
 
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