Low-Dose Tamoxifen for Radiation-Induced Breast Cancer Risk Reduction: A Phase IIB Randomized Placebo-Controlled Trial
Phase: PHASE2
Conditions: Breast Cancer
Sponsor: University of Alabama at Birmingham

PROTOCOL TITLE:   
Low-Dose Tamoxifen for Radiation-Induced Breast Cancer Risk 
Reduction: A Phase IIB Randomized Placebo-Controlled Trial 
 
 
CLINICALTRIALS.GOV ID: NCT01196936 
 
PROTOCOL VERSION:  
10/26/2015 

Low-dose Tamoxifen For Radiation-Induced 
Breast Cancer Risk Reduction:  
A Phase IIB Randomized Placebo-Controlled Trial 
 
Protocol Version: 10/26/2015 
 
Principal Investigator:  
 
Smita Bhatia, MD, MPH  
Co-Investigators: 
 
 
  
F. Lennie Wong, PhD  
Lusine Tumyan, MD 
Consultants: 
Laura Kruper, MD 
Courtney Vito, MD 
 
Participating Sites: 
 
University of Alabama at Birmingham 
Children’s of Alabama 
 (coordinating center) 
Smita Bhatia, MD, MPH 
Amitkumar Mehta, MD 
Heidi Umphrey, MD, MS, FCAP 
Wendy Landier, PhD, CRNP 
Purnima Singh, PhD 
Kandice Barnett, RN 
 
City of Hope National Medical Center 
Saro Armenian, DO 
James Waisman, MD 
Karla Wilson, RN, MSN, FNP-C, CPON  
 
St. Jude Children’s Research Hospital 
Melissa Hudson, MD 
Leslie Robinson, PhD 
Tommie J. Brace, CRN 
Ruth Eliason, CRN 
 
University of Michigan 
Breast & Ovarian Risk Program and 
C.S. Mott Children’s Hospital 
Sofia D. Merajver, MD, PhD 
Marcia Leonard, RN, NP 
Margaret Miodonski, CRN 
 
University of Chicago 
Tara Henderson, MD 
Elizabeth Drumm, FNP 
 
Mayo Clinic 
Sandhya Pruthi, MD 
Amy Degnim, MD 
 
 
 
Wake Forest,  
School of Medicine 
Susan Melin, MD 
 
University of Minnesota 
Anne Blaes, MD 
Karim Sadak, MD 
Mukta Arora, MD 
Karen Pennington, PA 
 
Seattle Cancer Care Alliance,  
University of Washington, and  
Fred Hutchinson Cancer Research Center 
K. Scott Baker, MD 
Julie Gralow, MD 
Larissa Korde, MD 
 
Dana Farber Cancer Institute 
Lisa Diller, MD 
Judy Garber, MD 
Andrea Ng, MD 
Ann H. Partridge, MD, MPH 
 
MD Anderson 
Therese B Bevers, MD 
Diane M Weber, CRN 
Valerie O Sepeda, CRN 
 
University of Kansas Hospital 
Breast Cancer Prevention Center Research 
Laboratory 
Carol J. Fabian, MD 
Trina Metheny, BHS, CT 
Carola M. Zalles, MD 
 
University of Colorado,  
Anschutz Medical Campus 
Linda Overholser, MD 
Virginia F. Borges, MD 
 
Princess Margaret Hospital and 
Hospital for Sick Kids 
David Hodgson, MD 
Pat Merante, CRN 
 
 

 
Protocol Version: 10/26/15 
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TABLE OF CONTENTS 
Page No. 
 
SCHEMA 
 
 
3 
1.0 
OBJECTIVES  
4 
2.0 
BACKGROUND AND HYPOTHESES 
4 
3.0 
DRUG INFORMATION: TAMOXIFEN 
15 
4.0 
RECRUITMENT STRATEGY 
19 
5.0 
SUBJECT ELIGIBILITY 
20 
6.0 
DOSE SCHEDULE AND RULES FOR TREATMENT DISCONTINUATION 
23 
7.0 
TREATMENT PLAN  
26 
8.0 
TOXICITIES TO BE MONITORED AND DOSAGE MODIFICATIONS 
26 
9.0 
STUDY PARAMETERS AND CALENDAR 
32 
10.0 
CRITERIA FOR EVALUATION AND ENDPOINT DEFINITIONS 
33 
11.0 
SPECIAL INSTRUCTIONS 
34 
12.0 
STATISTICAL CONSIDERATIONS 
46 
13.0 
REGISTRATION GUIDELINES 
55 
14.0 
RECORDS TO BE KEPT AND DATA SUBMISSION SCHEDULE 
56 
15.0 
MINORITIES AND WOMEN STATEMENT 
57 
16.0 
ETHICAL AND REGULATORY CONSIDERATIONS 
58 
17.0 
REFERENCES 
59 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
Protocol Version: 10/26/15 
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Schema 
 
 
230 patients will be randomized 1:1 to take a tablet containing 5mg tamoxifen (TAM) versus a 
placebo of equal size and shape daily for two years. The primary endpoint is mammographic 
density, which will be measured at baseline and on an annual basis (at Year 01 and 02). Blood 
and urine collections will also be scheduled annually for measurement of circulating markers of 
efficacy and safety. Normal breast tissue will be sampled before and after the two-year 
treatment course for measurement of tissue-based efficacy markers (breast tissue sampling by 
RPFNA no longer collected from patients).  
 
 

 
Protocol Version: 10/26/15 
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1.0 
OBJECTIVES 
 
1.1 
To determine the impact of a two-year course of low-dose tamoxifen administered at 
5mg per day on surrogate endpoint biomarkers of breast cancer (BC) risk, including:  
 
mammographic breast density (MBD), an established radiographic biomarker of BC 
risk,  
 
cytomorphology and proliferative index, tissue biomarkers closely linked to BC risk, 
and 
 
sex steroid hormones and insulin growth factors, circulating biomarkers of BC risk. 
1.2 
To establish safety and tolerability of this low-dose tamoxifen regimen, assessing both 
objective measures (lipid profiles, clotting factors and bone metabolism markers) and 
patient-reported outcomes. 
1.3 
To examine the modifying effect of demographic, clinical, and molecular characteristics 
on the risk: benefit ratio from this two-year low dose tamoxifen intervention. 
1.4 
To explore the relationship between this low-dose tamoxifen regimen and clinical 
measures of efficacy (new breast cancer and DCIS diagnoses) and toxicity 
(thromboembolic events, reports of hot flashes and gynecological symptoms, liver 
function abnormalities, and other cancer diagnoses). 
 
 
2.0 
BACKGROUND AND HYPOTHESES 
 
2.1 
BC risk in radiation exposed childhood, 
adolescent, 
and 
young 
adult 
cancer 
(CAYAC) survivors. Radiation therapy (RT) is 
a cornerstone of treatment for several cancers 
that 
commonly 
occur 
during 
childhood, 
adolescence, and young adulthood. These 
include Hodgkin lymphoma (HL), non-Hodgkin 
lymphoma 
(NHL), 
Ewing 
sarcoma 
(ES), 
rhabdomyosarcoma, 
and 
Wilms 
tumor. 
Occasionally other lymphomas and sarcomas 
are treated with RT as well. RT for HL and NHL 
with bulky mediastinal involvement typically 
involves the chest. Among sarcomas of the 
upper extremities treated with RT, the radiation 
port may include the axilla, from which there 
may be scatter to the chest wall. While RT for Wilms tumor typically involves the 
abdomen, occasionally the radiation RT port may cross the diaphragm. RT is also 
delivered as part of the total body irradiation (TBI) conditional regimen for hematopoietic 
cell transplant (HCT), commonly used in the treatment of acute leukemia and as salvage 
therapy for germ cell tumors and other solid cancers that may occur in children, 
adolescents, and young adults. 
 
Over the past 20 years, the incidence of childhood cancer has increased modestly from 
11.5 cases per 100,000 children in 1975 to 14.8 per 100,000 in 2004 [3]. During this 
same time, however, death rates dramatically declined for most childhood cancers. For 
instance, 5-year survival rates for all childhood cancers combined increased from 58% in 

 
Protocol Version: 10/26/15 
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1975–77 to 80% percent in 1996–2003 [3]. This trend has been attributed to therapeutic 
advances, including the use of RT.  
 
For example, until the late 1980s, standard treatment of pediatric patients with HL 
involvement of mediastinal lymph nodes (constituting three-quarters of patients) included 
RT at a dose of 40 to 45 Gy radiation to the “mantle” field, which encompasses the 
primary lymph node regions of the neck, supraclavicular, infraclavicular, axillary, and 
mediastinal areas (Figure 1) [2]. With this treatment, nearly 90% of patients diagnosed 
with HL, and treated with mantle radiation prior to age twenty survived beyond five years 
[3]. After the first late effects of HL therapy, such as growth deformities, began to be 
recognized, the field and dose of RT was reduced and radiation was combined with 
chemotherapy [4]. The combined modality approach has maintained high cure rates, 
such that late effects like second malignant neoplasm (SMNs) and cardiovascular 
disease now account for more deaths than those attributed to progressive HL [5]. In fact, 
of approximately 120,000 HL survivors in the United States, SMNs are the leading cause 
of death, with BC the most frequent solid tumor occurring in females treated for HL [6].  
Magnitude of risk of BC after radiation exposure:  Reported standardized incidence 
ratios (SIR) or relative risks (RR) for BC in this population range from 11.6 to 136.0, with 
the highest risk estimates generally from relatively smaller single institution studies [7-9]. 
From the single institution study with the largest number of women exposed to chest RT 
prior to 21 years of age (N=307), Wolden et al reported a RR of 26.2 [10]. A population-
based study by Metayer et al reported a RR of 11.6 derived from 2,737 women who 
were diagnosed with HL prior to 21 years of age identified from nine U.S. NCI SEER 
registries and selected European countries [11]. Within this cohort, 62 girls and young 
women diagnosed between ages 10 and 21 subsequently developed BC. Another 
population-based study, the British Childhood Cancer Survivor Study (British CCSS), 
which included 16 cases of BC among 383 women who were treated for HL prior to 15 
years of age [12], reported a similar SIR of 13.3. In contrast, in an international 
population-based study, Maule et al reported a SIR of 20.9 and an absolute excess risk 
of 40.6 based on 6 cases of BC that developed in 422 women with a prior diagnosis of 
HL occurring before age 15 [13]. Bhatia et al from the multi-institutional Late Effects 
Study Group (LESG) reported an excess risk of 55.5 based on 29 cases of BC among 
480 women diagnosed with HL prior to age 17 [14]. On the other hand, Kenney and 
colleagues reported an SIR of 24.7 in women who were treated for a pediatric cancer 
with chest RT prior to age 21 from the 26-institution U.S. Childhood Cancer Survivor 
Study (CCSS) [15]. With 95 cases of BC among 6,068 female pediatric cancer survivors, 
this study had a significantly larger number of women with BC, and thus may provide a 
more robust estimate. In addition, the risk estimate from the CCSS study included all 
pediatric cancers prior to BC, with a consequent heterogeneity of radiotherapy doses 
and fields, while the other risk estimates were based only on HL survivors. Thus, it is 
clear that the risk of BC is elevated in all radiation-exposed pediatric cancer survivors.  
Long latency between radiation exposure and diagnosis of BC:  The risk of BC does not 
begin to increase until approximately 8 years after chest RT, and the median age at BC 
diagnosis is 32 to 35 years, based on the LESG and CCSS studies [14, 15]. The British 
CCSS reported that by twenty-five years of follow-up, the cumulative risk of BC was 
12.2%.  
Risk of BC with attained age: Both the LESG and CCSS studies reported cumulative 
incidence of BC to be 13.9% and 12.9% by age 40 and 20% and 22% by age 45, 
respectively. Cumulative incidence rates reported by Travis et al ranged from 4.1% to 
39.6% by age 30, depending on the age at diagnosis and amount of radiation received 

 
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[16]. For perspective, among BRCA1 or BRCA2 gene mutation carriers, the estimated 
cumulative incidence of BC at age 30 ranges from 1-5% and at age 40 ranges from 10-
19% [17-20]. Thus, the risk for developing BC among HL survivors appears to exceed 
that of BRCA mutation carriers, especially before age 30 (4-40% in HL survivors 
compared to 1-5% in BRCA carriers). For survivors of other cancers, the risk appears to 
be elevated approximately 2-fold, similar to the degree of elevated risk for which 
standard-dose tamoxifen is typically offered. Importantly, in none of the studies of 
women treated with chest RT was a plateau of BC risk with increasing interval of follow-
up observed (Figure 7, page 14).  
Risk modification by radiation dose, age, endogenous hormones, and family history:  
Bhatia et al demonstrated preliminary evidence for an increasing risk of BC with a 
radiation dose of 26 Gy or more [14]. This has been confirmed by the CCSS, where a 
clear dose-response relationship is evident between the dose of RT to the chest and the 
risk of subsequent BC down to exposures of 15 Gy (Figure 2) [15]. Using a case-control 
design, Travis et al confirmed that BC risk was related to the dose of RT [21]. Following 
23.1-37.1 Gy and 37.2-61.3 Gy radiation delivered to the site where a secondary BC 
later appeared, the RR was 8.5 and 10.5, respectively. In Figure 3, Travis et al clearly 
demonstrates that BC risk increases with dose of radiation and latency from exposure to 
radiation. A novel finding in this study was a protective effect of exposure to an alkylating 
agent (AA) or radiation to the ovaries with respect to the development of BC following 
chest RT (Figure 3). While still having an increased BC risk compared to population 
rates, those who received chest RT plus an AA or pelvic RT had a significantly lower BC  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
Protocol Version: 10/26/15 
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risk than women treated with chest RT alone. Hill et al reported that parity appeared to 
contribute to BC risk among women who did not receive ovarian-damaging therapy [22]. 
The finding of a reduced risk in women with pelvic RT was confirmed in the CCSS 
(Figure 4) [15]. The findings across these studies highlight that endogenous estrogen 
may modify BC risk in this population. Although both the CCSS study and the report by 
Travis et al found that a family history (FH) of breast or ovarian cancer was associated 
with an increased risk of secondary BC among CAYAC survivors exposed to chest RT 
(Figure 4) [15, 22], multiple regression models suggested that additional risk of BC after 
chest RT is unlikely to be larger among women with a FH of breast or ovarian cancer. 
Lastly, age at radiation exposure also is an important risk factor, with chest RT 
exposures prior to age 30 impacting most significantly on BC risk (Figure 5).  
A synergistic effect between radiation and estrogen exposures was, in fact, originally 
described in a rat mammary carcinoma model in the 1970s [23]. This effect was related 
to hormonal dose [24], radiation dose [25], and age at radiation exposure [26]. All of 
these laboratory observations are consistent with the epidemiologic observations among 
radiation exposed CAYAC survivors described above. Therefore, estrogen appears to 
play an important role in the etiology of radiation-induced BCs, and an estrogen-blocking 
intervention is expected to prevent radiation-induced BCs.  
2.2 
Current options for BC risk reduction.  In other populations with an elevated risk for 
developing BC, such as women with a family history of BC, those with reproductive risk 
factors associated with BC, women with a personal history of premalignant breast 
disease, or those who are BRCA carriers, risk reduction strategies have included surgical 
and pharmacologic interventions [27].  
Surgical interventions:  Prophylactic bilateral mastectomy is associated with a 90-95% 
reduction in risk in women with familial BC [28]. This risk reduction measure has been 
the intervention of choice in women at high risk for BC who also have high cancer worry; 
however, not all women feel comfortable with this surgical option [29]. Alternatively, a 
50-70% reduction in BC risk has been observed with bilateral oophorectomy, in addition 
to a 80-90% reduction in ovarian cancer risk [30, 31]. Prophylactic bilateral salpingo-
oophorectomy (BSO) is a standard recommendation for women with a germline BRCA1 
or BRCA2 mutation after they have completed childbearing primarily because of the 
associated elevated ovarian cancer risk. However, due to increased risks of 
osteoporosis, dyslipidemia and cardiovascular disease resulting from early menopause, 
BSO is not a procedure that is recommended for BC risk reduction in other patient 
populations in which an elevated BC risk is not associated with an elevated ovarian 
cancer risk (eg. carriers of germline mutations in PTEN or TP53) [32]. 
Pharmacologic interventions:  Tamoxifen, a synthetic selective estrogen receptor 
modifier (SERM), was demonstrated to decrease the risk for BC by approximately 50% 
in women at moderately increased risk for developing BC as defined by the Gail model, 
a validated statistical tool which estimates the risk of developing BC by age 90 and 
within 5 years based on the following risk factors: current age, age at menarche, age at 
first birth, number of first degree relatives with BC, and number of prior breast biopsies 
[33, 34]. This finding led to the approval of tamoxifen for BC risk reduction by the Food 
and Drug Administration (FDA) in 1998. Raloxifene, another agent in the SERM class 
originally developed for the treatment of postmenopausal osteoporosis, was also noted 
to be associated with dramatic reductions in breast cancer risk [35, 36]. The NSABP P-2 
Study of Tamoxifen and Raloxifene (STAR) trial compared the relative efficacy of the two 
SERMs, tamoxifen and raloxifene, for BC prevention in postmenopausal women at 

 
Protocol Version: 10/26/15 
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moderately elevated risk for BC and found that raloxifene was equivalently efficacious 
but was associated with a better side effect profile [37]. However, raloxifene has only 
been tested in postmenopausal women, and is only FDA approved for either 
osteoporosis or BC prevention in women who have reached menopause.  
Aromatase inhibitors (AI) are more efficacious and are better tolerated than tamoxifen in 
the treatment of postmenopausal hormone receptor positive BC. In the adjuvant setting, 
a reduction in the incidence of contralateral BC was observed for all three currently 
marketed AIs, anastrozole, letrozole, and exemestane [38-40], suggesting that AIs may 
also be useful for BC chemoprevention. Clinical trials are now underway to test this 
hypothesis; however, their efficacy is limited to postmenopausal women, since negative 
feedback to the hypothalamus and the pituitary further drives, rather than suppresses, 
ovarian estrogen production in premenopausal women.  
Therefore, the only FDA-approved BC chemopreventive option in premenopausal 
women is tamoxifen. However, tamoxifen administered at a 20 mg daily dose is 
associated with uterine malignancies, stroke and venous thromboembolism. Although 
these adverse events occurred in less than 1% of the nearly 13,000 women studied, 
concerns about these side effects have been a deterrent to the widespread use of 
tamoxifen at a standard dose of 20 mg for BC chemoprevention. In addition, tamoxifen 
at this dose results in vasomotor and gynecological symptoms in approximately two-
thirds of users, which can interfere with patient compliance with their use [33].  
These concerns led some investigators in Italy to evaluate lower doses of tamoxifen in a 
four-arm 
double-blind 
placebo-controlled 
randomized 
trial 
of 
210 
healthy 
postmenopausal women, hoping to find a dose that would be better tolerated yet still 
efficacious for breast cancer risk reduction [41]. Participants were randomized to one of 
four treatment arms: 1) tamoxifen at 1 mg/day and a weekly placebo, 2) a daily placebo 
and tamoxifen at 10 mg/week, 3) tamoxifen at 5 mg/day and a weekly placebo, or 4) 
both daily and weekly placebos for 12 months. Plasma insulin-like growth factor 1 (IGF-
I), which is positively correlated with BC risk, was the primary endpoint, evaluated before 
and after the 12-month treatment. Secondary endpoints included MBD, IGF binding 
protein-3 (IGFBP-3), fibrinogen, antithrombin-III (AT3), C-telopeptide (CTX), and 
endometrial proliferation assessed by Pipelle biopsy. MBD is positively correlated with 
BC risk, while IGFBP-3 is negatively correlated with BC risk. Along with IGF-1, they 
served as surrogate endpoints of efficacy. Fibrinogen and AT3 are markers of clotting 
risk, CTX, which is positively correlated with bone resorptive activity, is a marker for 
osteoporosis risk, and endometrial proliferation is a marker of uterine cancer risk. 
Collectively, these were safety endpoints in the trial. The investigators reported a 
significant decrease in IGF-I in all tamoxifen arms relative to placebo (p = 0.005), with 
the greatest change observed in the 5 mg/day treatment arm (p = 0.019). Low-dose 
tamoxifen also lowered MBD and increased IGFBP-3, with optimal effects observed in 
the 5 mg/day arm. These findings all consistently suggest that 5 mg/day dosing is the 
most effective dose schedule of low-dose tamoxifen. No significant change was 
observed with CTX, suggesting that low-dose tamoxifen does not adversely affect bone 
health. Interestingly, a significant decrease in AT3 (p = 0.006), particularly with the 
5 mg/day dose schedule, was also seen, suggesting that low-dose tamoxifen may be 
associated with a lower clotting risk than even placebo. While there was an association 
between low-dose tamoxifen and endometrial thickness, no association was found with 
endometrial proliferation measured with Ki67 expression. In fact, Ki67 was lowest in 
patients receiving the 5 mg/day dose, suggesting that low-dose tamoxifen at that dose 
would likely not be associated with an increased risk of endometrial carcinoma as is 
observed with standard dose tamoxifen. Also, menopausal symptoms were not 

 
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significantly worsened by low-dose tamoxifen. In summary, low-dose tamoxifen, 
especially when taken at 5 mg/day is associated with a decreased risk of BC and without 
the typical adverse effects associated with standard dose tamoxifen.  
A follow-up trial randomized 235 premenopausal women with intraepithelial neoplasia 
(IEN) or 5-year Gail risk >1.3% to tamoxifen 5 mg daily versus fenretinide 200 mg daily 
in a 2x2 factorial fashion for two years. As with the previous study, a two-year course of 
5 mg daily of tamoxifen was associated with a significant decrease in MBD and IGF-1 
[42, 43]. Unlike the first study, the second study included long-term follow-up for 
outcomes, and after a median of 38 months, 24 women developed BC [42]. Trends 
toward lower annual BC rates were observed in the tamoxifen and fenretinide single 
agent arms, compared to placebo (2.0, 2.7, and 5.4 per 100, respectively) [42]. Based 
on the findings in both studies, we believe that a 5mg daily dose of tamoxifen will be a 
better tolerated, efficacious chemopreventive option for a mixed population of pre- and 
post-menopausal high-risk women.  
A preclinical tamoxifen chemoprevention trial resulted in both decreased incidence and 
prolonged latency of radiation-induced mammary carcinomas [44]. In this study, female 
Sprague-Dawley rats were exposed to 300, 500, or 900 cGy total body irradiation (TBI) 
vs sham and 30 days later half of animals in each radiation treatment group received 
tamoxifen. Tumor formation after 12 months of follow-up was completely inhibited, and 
tumor incidence after 24 months follow-up was reduced by 50% in all TBI groups 
combined. The tamoxifen intervention was most efficacious among animals who 
received 300 or 500 cGy TBI. 
Thus, there is strong preclinical, epidemiologic, and clinical trial evidence supporting our 
choice tamoxifen to reduce BC risk among young predominantly premenopausal female 
radiation-exposed HL survivors.  
2.3 
Clinical characteristics of radiation-associated BC. Little information exists regarding 
clinical characteristics and outcomes of women diagnosed with BC after radiation 
exposure. In the only peer-reviewed published study evaluating BC outcomes after RT for 
prior lymphoma – a single institution experience described by Sanna et al [45], BCs in 53 
women with either a prior diagnosis of HL or NHL treated at the European Institute of 
Oncology were compared to sporadic BC cases treated at the same institution matched 
for age, year of diagnosis, stage, and hormone receptor status. The cases were not 
significantly different in terms of biologic features (eg. Her-2 expression), but there was a 
trend toward more lobular histology, higher grade, and higher Ki67 proliferation index in 
the lymphoma-associated breast cancers. Interestingly, Sanna and colleagues reported 
that in their patient sample, both 5-year disease free survival (54% versus 91% in 
controls) and overall survival (87% versus 99%) were statistically significantly decreased 
(p < 0.0001 and p = 0.03, respectively) for BC diagnosed after lymphoma. Although no 
significant differences in medical and surgical therapy could be detected overall, 
anthracycline use was significantly lower among women with node-positive disease 
(p = 0.03). In addition, significant differences in RT among the lymphoma survivors were 
observed, with over a third receiving intraoperative electron beam radiotherapy, 
compared to only 10% of controls (p = 0.0001). Such treatment differences are likely due 
to concerns for toxicity due to prior treatment exposures, and probably underscore the 
differences in outcome, since histologic differences observed were minor. Regardless, 
the finding of worse clinical outcomes from secondary BC in lymphoma survivors 
provides a strong rationale to support the importance of finding ways to prevent this 

 
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disease in women who have survived HL, who are at enormously high risk for developing 
BC with limited ability to tolerate full treatment for BC, if it occurs.  
Because the control group for the patient cohort described by Sanna and colleagues 
was matched for hormone receptor status, nothing can be said about ER expression in 
HL-associated BC compared to sporadic BC. Only three studies have reported hormone 
receptor expression and are listed in Table 1. In all three studies, ER expression data 
was available on less than half of BC cases. Nevertheless, the two larger studies, 
including the CCSS study and one case series from Stanford, report a similar proportion 
of ER-expressing breast tumors among cases of secondary BC after HL compared to 
what would be expected with sporadic BC [10, 15]. Furthermore, several animal models 
have demonstrated no reduction in ER expression among radiation-induced rodent 
mammary tumors [46, 47].  
In addition, as reviewed above, it has been observed that CAYAC survivors who had 
received chest RT plus an alkylating agent or radiation to the ovaries, both of which are 
associated with premature menopause, had a significantly lower BC risk than CAYAC  
survivors treated with chest RT without these exposures [15, 16]. Similarly, surgical 
menopause after BSO has been shown to decrease sporadic and BRCA-associated BC 
[30, 31]. Thus, it is expected that a hormonal intervention would be appropriate for 
chemoprevention of radiation-associated BC, just as it is for sporadic and BRCA-
associated BC [48].  
Table 1: Breast Cancer After Hodgkin’s Disease – Tumor Characteristics 
Patient 
Population 
Female 
BC 
cases 
Average 
age at 
BC dx 
Histology 
Grade 
Stage 
ER/PR Reference
MSKCC, Breast 
Service records  
1969-1991 
45 
43 
IDC 31 (69%),     
ILC 2 (9%),       
mixed 1 (2%),     
colloid 2 (9%), 
medullary 1 (2%) 
DCIS 7 (16%),     
LCIS 1 (2%) 
Of n=31: 
Grade 1=1 (3%), 
Grade 2=21 (68%),
Grade 3=9 (29%). 
NR 
Of n=18: 
8 (44%) 
positive
[49] 
Stanford Hodgkin 
Disease radiation 
oncology clinical 
database  
1960-1997 
71 
42.6 
IDC 60 (85%),     
ILC 2 (3%);       
DCIS 8 (11%),     
LCIS 1 (1%) 
Of n=50: 
Grade 1=9 (13%), 
Grade 2=32 (45%),
Grade 3=19 (27%) 
Of n=56 invasive:  
31 (55%) stage I,  
18 (32%) stage II, 
5 (9%) stage III, & 
2 (4%) stage IV. 
Of n=30: 
19 (63%) 
positive
[50] 
26-center CCSS  
1970-1986 
111 
35 
IDC 77 (86%),     
ILC 4 (4%),       
mixed 3 (3%), 
phylloides 2 (2%), 
angiosarcoma 1 
(1%), fibrosarcoma 
1 (1%) 
NR 
Of n=66 invasive:  
30 (45%) Stage I;  
27 (41%) Stage II; 
4 (6%) Stage III; & 
5 (8%) Stage IV 
Of n=37: 
28 (76%) 
positive
  [15] 
 

 
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2.4 
Biomarkers of BC risk 
reduction. 
 
Because 
Phase 
III 
prevention 
trials require very large 
numbers of participants 
in 
order 
to 
show 
significant differences in 
clinical endpoints, many 
chemoprevention 
trials 
rely 
on 
biomarker 
endpoints [51]. There 
are several promising 
markers 
of 
BC 
risk. 
Mammographic 
breast 
density 
(MBD), 
proliferative 
index 
measured 
by 
Ki67 
expression, and intra-
epithelial 
neoplasia 
(IEN, includes lobular carcinoma in situ (LCIS), atypia and hyperplasia without atypia) are 
all considered biologically plausible surrogate endpoint biomarkers which both have a 
strong statistical association with BC and have been shown to be modulated by BC 
prevention intervention strategies [52]. Of these, the marker most easily and reliably 
measured is the radiographic marker, mammographic breast density (MBD).   
Mammographic breast density. On mammographic images, fibroglandular elements of 
the breast appear white, or dense, on a dark background of fatty tissue (Figure 6). 
Multiple epidemiologic studies have demonstrated an association between increased 
MBD and increased BC risk [53-60]. A meta-analysis reported that high MBD was 
associated with a nearly 4-fold increased risk in BC [61]. The earliest studies of MBD 
used a semi-quantitative scale, and reported an increased risk of BC in individuals with 
the highest MBD. Specifically, Boyd et al found that women with dense breast tissue in 
> 75% of their breast had an odds ratio (OR) of up to 6.0 (95% confidence interval: 2.5-
14.1) [62]. However, when three independent radiologists performed the readings, the 
ORs were 6.0, 3.7, and 2.8 for each of the readers. This suggests that MBD is a strong 
risk factor for BC, but that the semi-quantitative measurement has a large degree of 
variation between readers. Such variations led to the development of more quantitative 
measurements of MBD. Three separate groups have developed techniques to assess 
MBD using digitized mammograms [63-65]. These techniques allow for quantitative 
measurements, allowing for greater inter-reader reliability. Large cohort studies nested 
within the BC Detection and Demonstration Project and the Canadian National Breast 
Screening Study using these techniques demonstrate a clear positive linear relationship 
between MBD and BC risk (Table 2) [54, 55].  
 

 
Protocol Version: 10/26/15 
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Table 2: Association between breast cancer risk and percentage of breast 
area containing mammographic densities 
Breast Cancer Detection and Demonstration Project  
a screening study of >280,000 followed 1973-1980 
% Densities 
Cases (n) 
Controls (n) 
Unadjusted OR 
(95% CI) 
Adjusted OR 
(95% CI) 
Cumulative 
lifetime BC 
risk 
0 
141 
309 
1.0 
1.0 
4.1% 
1-24 
445 
632 
1.6 (1.3-2.1) 
1.6 (1.2-9.0) 
7.0% 
25-49 
490 
489 
2.5 (2.0-3.2) 
2.5 (1.9-3.2) 
10.3% 
50-74 
576 
554 
2.9 *2.2-3.6) 
2.8 (2.1-3.6) 
15.6% 
75+ 
194 
136 
4.5 (3.3-6.3) 
4.4 (3.1-6.1) 
17.7% 
 
1,846 
2,120 
Byrne, JNCI, 1995
More recently, Ursin et al at University of Oslo have successfully adapted their method 
to work with digital mammography, which is currently more widely used than traditional 
film mammography, and avoids measurement error that can be introduced by variability 
in the acquisition and subsequent digitization of mammographic films [65, 66]. With the 
ability to better quantify MBD using standard digital mammograms, investigators can 
now use it as a modifiable biomarker of BC risk [67, 68].  
Endogenous or exogenous estrogen, which is associated with stimulation of normal and 
premalignant breast epithelia, is associated with increases in MBD [69], and estrogen 
withdrawal is associated with a decline in MBD [70]. The degree of modulation of MBD 
parallels the magnitude of risk modification as well. In the Postmenopausal 
Estrogen/Progestin Interventions (PEPI) Trial, MBD was increased by combined 
estrogen (E) and progesterone (P) hormone replacement therapy (HRT), but not by 
estrogen alone [71]. Similarly, the Women’s Health Initiative (WHI) found that the use of 
E+P HRT was associated with a 25%  increased incidence of BC, while HRT with E 
alone did not have any significant impact on BC incidence [72, 73]. Also of note, the 
effect of E+P on MBD could be observed as early as after one year of treatment, 
preceding its effect on BC incidence after five years of use [74] Based on these findings, 
estrogen blockade would be predicted to suppress MBD. Indeed, several reports have 
found that the SERMs, tamoxifen and raloxifene, both of which have been associated 
with decreased BC incidence in large clinical trials [33, 36], are both associated with 
decreases in MBD after 1-2 years of treatment, preceding effects on clinical endpoints 
seen after five years of treatment [75-79].  
Recently, a MBD reduction of 10% or more 12-18 months after initiating therapy was 
shown to predict who achieved clinical benefit from standard-dose tamoxifen [80]. While 
that study used subjective MBD readings in which MBD was scored in 5% increments, 
the Ursin method, a computer-assisted method that reports MBD on a continuous scale, 
was shown to be very sensitive to small changes in the PEPI trial [71]. 
Thus, biomarker observations within clinical trials suggest that two years is a sufficient 
duration to detect significant differences in MBD as a primary endpoint, that the MBD 
biomarker endpoint will predict who will have clinical benefit from tamoxifen BC risk 
reduction, and that the Madena method will sensitively detect MBD changes in response 
to tamoxifen. 

 
Protocol Version: 10/26/15 
13
 
Cytomorphology 
and 
proliferation. 
According 
to 
well-established 
models 
of 
carcinogenesis, the transition of normal cells to invasive cancer involves the following 
progressive intermediate morphologic steps: from typical hyperplasia, to atypical 
hyperplasia, and then carcinoma-in-situ [81]. Each of these premalignant states are 
associated with increased BC risk, as well as lower category, or non-IEN, benign breast 
disease [82]. In a study of normal and premalignant breast lesions, the percentage of 
Ki67-expressing cells was significantly increased in all of the premalignant lesions in 
comparison to normal lobules, with the following trend observed: the highest expression 
was seen in DCIS, followed by LCIS and atypical hyperplasia [83]. Among women with 
typical hyperplasia, patients with benign breast lesions that progressed to BC had a 
significantly higher proliferative index, measured by Ki67 expression in their lesions, 
compared to patients who did not [84]. Cytomorphology may also be assessed from fine 
needle aspiration samples using the 24-point Masood scoring classification (Table 3) 
[85]. Cytomorphology classified in this manner has also been shown to be significantly 
associated with Ki67 expression [86]. Control of cellular proliferation is important for 
cancer prevention since active proliferation has an integral role in carcinogenesis, 
including the processes of initiation and promotion [87].  
Although no data exists of the impact of tamoxifen on Ki67 in the prevention setting, a 
decrease in Ki67 index has been shown to correlate with a clinical response to 
neoadjuvant therapy with tamoxifen [88]. In fact, early changes in Ki67 index during 
neoadjuvant therapy with another hormonal agent, letrozole, has been noted to precede 
tumor response [89, 90]. According to Ki67 reproducibility studies, a > 36% change in 
Ki67 index after 2-3 weeks of therapy is necessary to predict a statistically significant 
improvement in response to primary chemo-endocrine therapy at 3 months [91]. In the 
prevention setting, letrozole has been reported to significantly decrease Ki67 index 

 
Protocol Version: 10/26/15 
14
determined from fine needle aspirations when women with atypia were treated for 6 
months [92]. As far as breast pathology itself is concerned, tamoxifen has been 
associated with a decrease in DCIS, LCIS, atypical hyperplasia, and other benign breast 
disease [33, 93].  
Several methods are available for sampling breast epithelial cells: random periareolar 
fine needle aspiration (RPFNA), which has been used extensively by Dr. Carol Fabian's 
group, core needle biopsy, used in several of our previous biomarker modulation BC 
prevention studies, and ductal lavage [94, 95]. Each of these techniques is well tolerated 
by healthy women, and each has unique advantages and disadvantages [96]. Both 
RPFNA and core biopsy have the highest epithelial yields, sufficient for multiple 
biomarker assays, but RPFNA is slightly less invasive. In addition, RPFNA allows 
cytomorphology evaluation using the standardized continuous-scale Masood scoring 
method described above, which is highly reliable and has been shown to reflect changes 
in response to hormonal chemopreventive interventions [92].  
Circulating biomarkers of BC risk - Dose-response relationships have been noted 
between tamoxifen and multiple serum biomarkers of BC risk. These include decreased 
IGF-1 and increased IGFBP-3, as discussed earlier, as well as increased sex hormone-
binding globulin (SHBG) levels [97, 98]. While these circulating biomarkers are 
correlated to BC risk, cytomorphology and proliferative Ki67 index are the only direct 
measures of breast tissue response to BC risk-reducing interventions. Sampling of 
normal breast epithelia to assess both biomarkers is feasible through a well tolerated 
method, RPFNA (please see discussion above). Nevertheless, this procedure is not 
considered standard of care for radiation exposed female CAYAC survivors, while 
annual screening mammography is. In addition, while MBD is evaluable on all digital 
mammograms, some RPFNA samples will not yield sufficient cells for Ki67 assessment. 
Therefore, we have chosen MBD to be the primary endpoint in the proposed 
chemoprevention trial.  
2.5 
Significance 
In summary, evidence from several high quality studies indicates that women treated 
with chest RT by the age of 40 years have a very high risk of developing BC [7-16]. The 
risk begins to increase within 8 years following RT, and a substantial proportion of these 
women will be diagnosed with their BC before the age of 40. Indeed, by the age of 40-45 
years, about 20% of women treated with moderate to high dose chest RT for childhood 
HL will be diagnosed with BC, 
mirroring the magnitude of BC risk 
seen in women who carry germline 
mutations in the BRCA1 or BRCA2 
genes (see Figure 7). Options to 
reduce 
BC 
risk 
include 
prophylactic 
mastectomy 
or 
oophorectomy, but not all women 
are 
interested 
in 
invasive 
procedures. 
Non-surgical 
interventions include a variety of 
drugs which interfere with estrogen 
signaling to the breast; however, 
the only one which is FDA 
approved for young women who 

 
Protocol Version: 10/26/15 
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have not yet reached menopause is tamoxifen. Tamoxifen taken at 20 mg/day for five 
years is associated with a 50% reduction in BC risk [33]. However, when taken at that 
dose, tamoxifen is associated with rare but concerning side effects that interfere with 
patient compliance with the medication. On the other hand, tamoxifen taken at 5 mg/day 
is associated with a decreased risk of BC without the typical adverse effects associated 
with standard dose tamoxifen [41]. Findings across several studies suggest that a 
hormonal intervention would be appropriate for chemoprevention of radiation-associated 
BC [15, 16, 50]. Because definitive Phase III BC prevention trials require tens of 
thousands of participants to show significant differences in clinical endpoints, many 
chemoprevention trials make use of biomarker surrogate endpoints [51]. MBD, Ki67 
proliferative index, and evidence of IEN on cytomorphology are all considered 
biologically plausible surrogate endpoint biomarkers with both a strong statistical 
association with BC and evidence of modulation by BC prevention intervention 
strategies. Of these, the marker most easily and reliably measured is MBD, which we 
have chosen to be our primary endpoint, although we will also measure the other two as 
secondary endpoints. Tamoxifen has been shown to decrease both MBD and the 
incidence of IEN in the prevention setting in an epidemiologically defined BC risk 
population [33]. We now aim to demonstrate a similar effect in female CAYAC survivors 
at high risk for developing BC due to chest RT exposure at a young age. Considering 
that SMNs are the leading cause of death in CAYAC survivors, with BC the most 
frequent secondary solid tumor [6], we are compelled to develop targeted intervention 
studies for those at highest risk. Thus, we propose the first BC chemoprevention trial for 
female CAYAC survivors who received chest RT as part of their treatment regimen by 
the age of 40. 
2.6 
Hypotheses 
We hypothesize that low-dose tamoxifen delivered at a dose of 5 mg per day for two 
years will be an efficacious and a safe option for BC risk reduction in young women at an 
extremely high risk for developing BC due to exposure to chest RT during childhood or 
early adulthood.  We also hypothesize that there exists a subgroup within the patient 
population who will derive the most benefit from taking low-dose tamoxifen for BC 
prevention with the least risk. 
  
 
3.0 
DRUG INFORMATION: TAMOXIFEN 
 
3.1 
Chemistry 
The chemical name for tamoxifen citrate is (Z)2-[4-(1,2-diphenyl-1-butenyl) phenoxy]-N, 
N-dimethylethanamine 2-hydroxy-1,2,3- propane-tricarboxylate (1:1). The structural and 
empirical formulas are: (C32H37NO8). Tamoxifen citrate has a molecular weight of 
563.62, the pKa is 8.85, and the equilibrium solubility in water at 37°C is 0.5 mg/mL and 
0.2 mg/mL in 0.02 N HCl at 37°C. Commercially available tamoxifen tablets also contain 
the following inactive ingredients: anhydrous lactose, colloidal silicon dioxide, 
crospovidone, magnesium stearate, microcrystalline cellulose and sodium lauryl sulfate. 
 
3.2 
Mechanism of Action 
Tamoxifen is a nonsteroidal agent that has demonstrated potent antiestrogenic 
properties in animal test systems. The antiestrogenic effects are thought to be related to 
its ability to compete with estrogen for binding sites in target tissues such as breast. 

 
Protocol Version: 10/26/15 
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3.3 
Human Toxicity 
Toxicity data from prevention trials in healthy women are reviewed in detail here [33, 41].  
Standard dose tamoxifen: The Breast Cancer Prevention Trial (BCPT, NSABP P-1) was 
a double-blind randomized placebo-controlled trial with a primary objective to determine 
whether 5 years of tamoxifen therapy at 20 mg per day would reduce the incidence of 
invasive breast cancer in women at high risk for the disease. Secondary objectives 
included an evaluation of the incidence of ischemic heart disease; the effects on the 
incidence of bone fractures; and other events that had been associated with the use of 
tamoxifen in breast cancer therapeutic trials, including: endometrial cancer, pulmonary 
embolus, deep vein thrombosis, stroke, and cataract formation and surgery.  
In the BCPT, NSABPP-1 trial, 13,388 women of at least 35 years of age, who had a 5-
year risk for developing breast cancer of 1.67% or more estimated by the Gail model, 
were randomized to receive either tamoxifen 20 mg per day or placebo for five years. 
After a median follow-up of 54.6 months, the incidence of invasive breast cancer was 
reduced by 49% among women assigned to tamoxifen compared to placebo 
(p<0.00001; relative risk (RR)=0.51, 95% CI: 0.39-0.66). In addition, a significant 
decrease in the incidence of ductal carcinoma in situ (DCIS) was seen (p<0.002; RR = 
0.50, 95% CI 0.33-0.77). There was no statistically significant difference in the number of 
myocardial infarctions, severe angina, or acute ischemic cardiac events between the two 
groups. With regards to skeletal fractures, several beneficial nonsignificant trends were 
noted: there was a 45% reduction in fractures of the hip (RR = 0.55; 95% CI 0.25–1.15), 
a 39% reduction in wrist fractures (RR = 0.61; 95% CI 0.29–1.23), and a 26% reduction 
in fractures of the spine (RR = 0.74; 95% CI 0.41–1.32). 
The risks associated with 5 years of 20 mg per day of tamoxifen in this trial include 
endometrial cancer, deep venous thrombosis (DVT), pulmonary embolus (PE), stroke, 
cataract formation and cataract surgery, as shown in the table below, with statistical 
comparisons between treatment groups immediately following:   
 
Participants who received tamoxifen in this trial had a 2.53 times greater risk of 
developing an invasive endometrial cancer (95% CI 1.35–4.97) than did women who 
received placebo. Of note, the increased risk was predominantly in women 50 years of 
age or older. The RR of women aged 49 years or younger was 1.21 (95% CI 0.41– 
3.60), whereas it was 4.01 (95% CI 1.70–10.90) in women aged 50 years or older.  
Pulmonary emboli (“blood clot in the lungs” in the table above) were rare, fewer than 1 
case per 1000, but were observed in almost three times as many women in the 

 
Protocol Version: 10/26/15 
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tamoxifen group as in the placebo group (total number of cases per treatment group: 18 
versus 6, respectively; RR = 3.01; 95% CI 1.15–9.27). When the incidence of pulmonary 
embolism was related to the age of participants, it appeared that most of the risk was 
confined to the older subgroup. Among women aged 49 years or younger, only one 
event occurred in the placebo group and two events occurred in the tamoxifen group 
(RR = 2.03; 95% CI 0.11–119.62), in contrast to those aged 50 years or older, in whom 5 
events occurred in the placebo group and 16 in the tamoxifen group (RR = 3.19; 95% CI 
1.12–11.15). 
More women who received tamoxifen developed deep venous thrombosis (“blood clot in 
the veins” in the table above) than did women who received placebo (35 versus 22 
cases, respectively; RR = 1.60; 95% CI 0.91–2.86). Again, the excess risk appeared to 
be greater among women aged 50 years or older. For women aged 49 years or younger, 
the number of cases was 8 in the placebo group versus 11 in the tamoxifen group (RR = 
1.39; 95% CI 0.51–3.99). In women 50 years of age or older, the number of cases was 
14 versus 24, with an RR of 1.71 (95% CI 0.85–3.58). 
The rate of cataract development among women who were cataract-free at the time of 
randomization was 21.7 per 1000 women in the placebo group and 24.8 per 1000 
women in the tamoxifen group (the rates in the table above represent total number of 
cases per treatment group). This represents an RR of 1.14, with confidence intervals 
that indicate marginal statistical significance (95% CI 1.01–1.29). There was also a 
difference by treatment group with respect to cataract surgery. In the placebo and 
tamoxifen groups, the rates of developing cataracts and undergoing cataract surgery 
were 3.00 and 4.72 per 1000 women, respectively (RR = 1.57; 95% CI 1.16–2.14). 
In breast cancer treatment trials, tamoxifen at 20 mg per day has been associated with 
changes in liver enzyme levels, and on rare occasions, a spectrum of more severe liver 
abnormalities including fatty liver, cholestasis, hepatitis and hepatic necrosis. In most 
reported cases the relationship to the drug was uncertain. 
Common yet less serious side effects associated with standard dose tamoxifen include 
hot flashes, vaginal discharge, and irregular menstrual bleeding. On the NSABP P-1 
trial, hot flashes occurred in 68% of women on placebo and in 80% of women on 
tamoxifen. Vaginal discharge occurred in 35% and 55% of women on placebo and 
tamoxifen, respectively. There was no difference in the incidence of vaginal bleeding 
between treatment arms.  
During postmarketing surveillance, there have been some reports of occasional 
headaches or skin rashes on tamoxifen at 20 mg/day. T4 elevations were reported for a 
few postmenopausal patients which may be explained by increases in thyroid-binding 
globulin, but these laboratory findings were not accompanied by clinical hyperthyroidism. 
 
Low dose tamoxifen: As reviewed in the background section, Decensi et al performed a 
trial of several lower doses of tamoxifen in healthy postmenopausal women. Findings 
suggested that 5 mg per day dosing is the most effective dose schedule of low-dose 
tamoxifen compared to placebo. No significant change was observed with C-
telopeptides, suggesting that low-dose tamoxifen does not adversely affect bone health. 
A significant decrease in antithrombin-III (p = 0.006) suggested that low-dose tamoxifen 
may still be associated with a higher clotting risk compared with placebo; but, this may 
be improved compared with standard-dose tamoxifen. While there was an association 
between low-dose tamoxifen and endometrial thickness, no association was found with 
endometrial proliferation measured with Ki67 expression. In fact, Ki67 was lowest in 
patients receiving the 5 mg/day dose, suggesting that low-dose tamoxifen at that dose 

 
Protocol Version: 10/26/15 
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would likely not be associated with an increased risk of endometrial carcinoma as is 
observed with standard dose tamoxifen. In addition, menopausal symptoms were not 
significantly worsened by low-dose tamoxifen. A follow-up study of premenopausal 
women found similar results.  
 
In summary, because the dose that we will be administering will be 5 mg per day rather 
than 20 mg per day, we expect no significant increase in these adverse events in the 
treatment group compared to the placebo group. In addition, the median age of the 
patient cohort from which trial participants will be drawn is 36 years old, while the 
adverse effects associated with the 20 mg per day of tamoxifen were only significantly 
elevated above placebo among women who were 50 years old or older at the time of 
treatment initiation. However, precautions will be taken, nevertheless (see Section 8.2 
for guidelines regarding adverse event reporting and for data safety monitoring). In 
addition, women with childbearing potential will be warned that they should avoid 
pregnancy while taking study medication and for two months after discontinuation.  
 
3.4  
Pharmaceutical Data  
Absorption and Distribution: Following a single oral dose of 20 mg tamoxifen (standard-
dose tamoxifen), an average peak plasma concentration of 40 ng/mL (range 35 to 
45 ng/mL) occurred approximately 5 hours after dosing. The decline in plasma 
concentrations of tamoxifen is biphasic with a terminal elimination half-life of about 5 to 
7 days. The average peak plasma concentration of N-desmethyl-tamoxifen is 15 ng/mL 
(range 10 to 20 ng/mL). The average steady-state plasma concentrations of tamoxifen 
and N-desmethyl-tamoxifen after administration of 20 mg tamoxifen once daily for 
3 months are 122 ng/mL (range 71-183 ng/mL) and 353 ng/mL (range 152-706 ng/mL), 
respectively. Chronic administration of 10 mg tamoxifen given twice daily for 3 months to 
patients results in average steady-state plasma concentrations of 120 ng/mL (range 67-
183 ng/mL) for tamoxifen and 336 ng/mL (range 148-654 ng/mL) for N-desmethyl 
tamoxifen. After initiation of therapy, steady state concentrations for tamoxifen are 
achieved in about 4 weeks and steady-state concentrations for N-desmethyl tamoxifen 
are achieved in about 8 weeks, suggesting a half-life of approximately 14 days for this 
metabolite. In a steady-state, crossover study of 10 mg tamoxifen tablets given twice a 
day vs. a 20 mg tamoxifen tablet given once daily, the two schedules were found to be 
bioequivalent. 
Metabolism: Tamoxifen is extensively metabolized after oral administration. N-desmethyl 
tamoxifen is the major metabolite found in patients' plasma. The biological activity of N-
desmethyl tamoxifen appears to be similar to that of tamoxifen. 4-Hydroxytamoxifen and 
a side chain primary alcohol derivative of tamoxifen have been identified as minor 
metabolites in plasma. Tamoxifen is a substrate of cytochrome P-450 3A, 2C9 and 2D6, 
and an inhibitor of P-glycoprotein.  
Excretion: Studies in women receiving 20 mg of 14C-tamoxifen have shown that 
approximately 65% of the administered dose was excreted from the body over a period 
of 2 weeks with fecal excretion as the primary route of elimination. The drug is excreted 
mainly as polar conjugates, with unchanged drug and unconjugated metabolites 
accounting for less than 30% of the total fecal radioactivity. 
Pregnancy: Category D. Tamoxifen may cause fetal harm when administered to a 
pregnant woman. Women should be advised not to become pregnant while taking 
tamoxifen or within 2 months of discontinuing tamoxifen and should use barrier or other 

 
Protocol Version: 10/26/15 
19
nonhormonal contraceptive measures if sexually active. Tamoxifen does not cause 
infertility, even in the presence of menstrual irregularity. 
 
3.5  
Supplier 
Tamoxifen is commercially available in 10 mg and 20 mg tablets. For this trial, generic 
5mg capsules and matching placebo capsules will be manufactured by Sharp Clinical 
Services. 
Tamoxifen and matching placebo will be manufactured as a capsule for oral 
administration. Each 5 mg tamoxifen capsule contains 7.6 mg of tamoxifen citrate, which 
is equivalent to 5 mg of tamoxifen, as well as anhydrous lactose, colloidal silicon dioxide, 
crospovidone, magnesium stearate, microcrystalline cellulose and sodium lauryl sulfate, 
as inert excipients. Matching placebo capsules will contain anhydrous lactose.  
For this study, blistered cards will contain a month’s supply of tamoxifen 5mg or 
matching placebo capsules.  
 
3.6   
Storage and Stability 
Tamoxifen 5 mg capsules and matching placebo will be shipped at room temperature. 
The capsules should be stored at a controlled room temperature of 20-25°C (68-77°F) in 
the supply kits containing the blister cards. They will be stored and dispensed in the 
original containers.  
 
3.7 
Administration 
Trial participants will be instructed to take one capsule per day orally. They will be 
instructed to swallow the capsule whole, with water or another non-alcoholic liquid, 
preferably the same time each day. They may take the capsules with or without food. If 
they forget a dose, participants will be asked to take it when they remember, and then 
take the next dose as usual, but if they do not remember until their next dose, to not take 
extra capsules to make up the missed dose. 
For premenopausal subjects, they will be instructed to begin their study medication 
course while menstruating or after negative beta-HCG test as below. For premenopausal 
subjects who do not have regular menstrual periods or are between periods, a beta-
HCG level will be performed to confirm that the patient is not pregnant before starting 
study medication. Participants will be warned that they should avoid pregnancy while 
taking study medication and for two months after discontinuation. They will be advised to 
use barrier or nonhormonal contraceptive measures.  
 
 
4.0 
RECRUITMENT STRATEGY  
 
Via the Consortium for Pediatric Interventional Research (CPIR), a consortium of five 
institutions, including the City of Hope National Medical Center (COH), St. Jude 
Children’s Research Hospital (SJCRH), Emory University (EU), University of Michigan 
(UM), and Princess Margaret Hospital in Toronto (PMH), we have constructed a cohort 
of HL survivors who were exposed to chest RT during childhood or young adulthood, at 
least 5 years prior to study participation.  This cohort will form the initial basis from which 
accrual in this trial will be conducted. Other sites and additional CAYAC diagnoses 
treated with RT involving the chest may be added as needed to complete accrual for the 
trial.  

 
Protocol Version: 10/26/15 
20
 
Recruitment will be conducted both prospectively and retrospectively as described 
below.  Because this is a highly motivated patient population, at study activation we 
anticipated that it would be feasible to achieve a recruitment yield of 30%, and that the 
accrual goal of 230 would be achievable across the CPIR participating institutions 
described above. After the initial two years of accrual, we observed an average 
recruitment yield of only 14%. Therefore, new sites were recruited, and additional sites 
have been identified and will be recruited as needed to reach the accrual goal.   
4.1 
Retrospective recruitment   
Participants in any long-term cancer survivorship clinic, database or registry associated 
with a participating site, who are alive and eligible based on medical record review will 
be mailed a letter and study brochure describing the study. The letter lets them know 
that a representative from the study will be calling to tell them more about the study, 
unless they opt out (i.e. passive consent). Upon contact, the patient will be given another 
opportunity to opt out. Patients who agree to be prescreened will be interviewed per the 
Intake Form. Subjects who are screened by this method will form cohort 1.  
4.2 
Prospective recruitment   
Alternatively, the study will be introduced to eligible patients at their survivorship and/or 
high risk screening and/or prevention clinic visit and given a brochure describing the 
study to take home to read. The study clinical research associate (CRA) or clinical 
research nurse (CRN) will then follow up with a call the following week to assess interest 
and confirm eligibility. Subjects who are screened by this method will form cohort 2. 
Patients may also self-identify and call in to a local site via recruitment advertising on the 
internet or from a flyer placed in the community (such as community centers, libraries 
and churches). Self-identifying subjects will form cohort 3.  
4.3 
Informed consent process   
Eligible and interested individuals will be approached for informed consent.  One of the 
members of the Protocol Management Team (PMT) at each site will introduce the study 
to potentially eligible patients, either at a survivorship clinic visit or via a mailing as 
described. The PMT at each site includes the consortium PI, the co-investigator, and the 
clinical research nurse (CRN), and/or clinical research associate (CRA). All potential 
risks and benefits of the study, as well as alternatives, will be fully explained. Whenever 
possible, the informed consent process will be conducted in the patient’s preferred 
language. If that is not possible, a professional translator will be used. 
 
 
5.0  
ELIGIBILITY CRITERIA 
 
5.1 
Inclusion Criteria 
 
 
The following are requirements for entry onto this Phase IIb trial: 
 
 
Females, 25 years of age or older at the time of registration  
 
Exposure to RT delivered to the chest, axilla, and/or supraclavicular areas at a 
     cumulative dose of 12 Gy or more by age 40 years. In addition, patients who 
received total body irradiation by age 40 may be considered.  

 
Protocol Version: 10/26/15 
21
 
No evidence of active disease from their primary cancer for at least 2 continuous 
years prior to registration. The indication for RT is not specified but cannot be for 
primary breast cancer. Common examples of primary cancer diagnoses include, 
but are not limited to: lymphoma, leukemia, sarcoma, and Wilms tumor occurring 
in pediatric patients, and lymphoma, leukemia, and sarcoma occurring in young 
adults. Primary cancer therapy must have been completed at least 6 months 
prior to registration.  
 
Well-defined menopausal status falling into one of the following categories:  
o Premenopausal, defined as age at registration 45 years old or younger 
with regular monthly period for at least 6 consecutive months prior to 
registration.  
o Postmenopausal, defined as continuous absence of menstruation for 12 
months OR status-post bilateral oophorectomy OR follicle stimulating 
hormone (FSH) level in the postmenopausal range.  
 
5.2 
Exclusion Criteria 
 
 
 
Subjects will be excluded if any of the following characteristics are present:  
 
SMN other than those listed below diagnosed within 2 years of study entry. Patients 
with the listed indolent or pre-invasive neoplasms may be eligible if diagnosed within 
2 years and all treatment was completed at least 6 months prior to registration:  
o non-melanoma cancers of the skin 
o thyroid cancer 
o cervical cancer confined to the cervix or cervical intraepithelial neoplasia 
(CIN) 
o ductal carcinoma in situ (DCIS) or breast IEN (includes atypical hyperplasia 
and LCIS) 
o superficial or non-invasive transitional cell carcinoma of the bladder 
For women with a prior history of ductal carcinoma in situ (DCIS) or breast IEN, only 
one breast could have been involved and all therapy must have been completed at 
least 6 months prior to registration. In addition women with a prior history of invasive 
breast cancer may also be eligible, as long as only one breast was involved, they 
were diagnosed at least 2 years prior to study entry, and therapy was completed at 
least 6 months prior to study entry. 
 
Bilateral breast implants or status-post bilateral prophylactic mastectomy. 
 
Evidence of malignant breast disease on any form of breast imaging. The study only 
requires annual mammography; however, annual breast MRI is considered standard 
of care in this patient population (per COG or NCCN follow-up guidelines), and 
breast ultrasound may be indicated if a palpable lesion is detected on screening 
clinical breast exam.  Abnormal imaging may require additional radiographs and/or 
breast biopsy.  Patients who are found to have benign breast disease with or without 
atypia may continue on study as long as there is no evidence of malignancy. If there 
is evidence of malignancy, and only one breast is involved, they may be 
reapproached 6 months after completion of therapy for consideration of the trial.  
 
Baseline categorical mammographic density scored as BIRAD 1, or extremely fatty, 
in both breasts (see Figure 9).  If the patient has a prior history of IEN (DCIS, LCIS, 
or atypical hyperplasia), the contralateral breast must not have a mammographic 
density score of BIRAD 1.  This determination will be made at the local site.  

 
Protocol Version: 10/26/15 
22
 
 
Current or recent use (within 6 months of registration or baseline mammogram, 
whichever is first) of any of the following hormonal agents:   
o systemic hormone replacement therapy (includes oral or transdermal 
formulations). Vagifem and Estring, two formulations of locally applied 
vaginal estrogen associated with minimal systemic absorption, may be 
allowed. Other estrogen-containing vaginal creams, while not an 
exclusion, should be avoided whenever possible.  Patients with a history 
of hormone modifying herbal supplements (see section 11.14) are 
eligible, but patients will be asked to avoid their use after on study.  
o hormonal forms of contraception (includes oral, transdermal, implanted, 
and injectable formulations),  
o selective estrogen receptor modifiers (such as tamoxifen and raloxifene),  
o aromatase inhibitors,  
o GnRH analogs,  
o prolactin inhibitors, or 
o androgens or antiandrogens.  
 
Concurrent use of warfarin and strong inhibitors of CYP2D6 (see Section 11.15 for a 
list) will not be allowed.  
 
A personal history or a strong family history of venous thromboembolism, (VTE), 
including deep venous thrombosis (DVT) or pulmonary embolus (PE). A one-time 
personal history of catheter-associated DVT may be allowed, as long as there were 
no subsequent VTE events and a strong family history is not present.  Examples of a 
strong family history include (but are not limited to): one first degree relative with 
more than one VTE event in the same individual, and two family members in the 
same lineage with unexplained VTE or two VTE events in the same individual.  If a 
family history is present, but it is not clear whether or not it is clinically significant, 
consideration of genetic testing to rule out a hereditary clotting syndrome (e.g. FVL, 
prothrombin G2010A mutations) may be considered. Because of a drug-drug 
interaction with tamoxifen, concurrent with warfarin use for any reason is not allowed.  

 
Protocol Version: 10/26/15 
23
For this reason, patients with atrial fibrillation may not participate. However, patients 
with coronary artery disease or congestive heart failure without atrial fibrillation may 
be allowed to participate. Patients with a personal history of a cerebrovascular 
accident (CVA), transient ischemic attack (TIA), or retinal vein thrombosis will not be 
allowed to participate. 
 
Current intrauterine pregnancy or plans to become pregnant within two years. In 
addition, currently nursing mothers will be excluded.  
 
Renal or hepatic insufficiency, defined as having a serum creatinine, total bilirubin, 
SGOT, or SGPT greater than 2x the institutional norm. 
 
Unable to provide consent. 
 
 
6.0 
DOSE SCHEDULE AND RULES FOR TREATMENT DISCONTINUATION 
 
6.1 
Dose Schedule 
Randomization will be stratified according to the rules outlined in Section 12.3. Subjects 
will be randomized 1:1 to receive low-dose tamoxifen 5 mg or placebo daily for 24 
months. Registration will be done via a central database, Interactive Web Response 
System (IWRS), which will also perform stratified treatment assignment. This system is 
maintained by Sharp Clinical Services, which is separate from the coordinating center 
and all participating sites such that all study staff (except for the statistician when 
preparing reports for the DSMB) will remain blinded.   
 
6.2 
Dose Escalation 
There will be no dose escalation in this Phase IIb study.  
 
6.3 
Criteria for Discontinuation of Study Protocol Therapy 
 
 
Patient requests to withdraw from study 
 
Treatment interruption longer than 90 consecutive days 
 
A breast or gynecologic event (see definitions below) 
 
Pregnancy 
 
The occurrence of a grade 2 toxicity with probable or greater attribution without 
resolution within 72 hours, or any grade 3 or 4 toxicity with possible or greater 
attribution meeting definitions below 
 
Death 
 
A breast event is defined as a new diagnosis of DCIS or invasive BC.  These events are 
expected to occur as a consequence of the high risk study population.  In the case that it 
should occur, the subject will be taken off study and referred to the appropriate breast 
surgeons and medical oncologists for proper diagnostic work-up and treatment.  For 
subjects who develop atypical hyperplasia (AH) or lobular carcinoma in-situ (LCIS), or 
any other form of benign breast disease, their specific histologic diagnosis and the date 
of diagnosis will be recorded, but they may continue on study.  
 
A gynecologic event is defined as a new diagnosis of endometrial carcinoma or uterine 
sarcoma.  Although these events may occur sporadically, they have been associated 
with standard dose tamoxifen and therefore are listed among criteria for discontinuation 
of study treatment.  In the case that a subject should experience dysfunctional uterine 

 
Protocol Version: 10/26/15 
24
bleeding of any etiology, the subject will be referred to a gynecologist for proper 
diagnostic work-up.  Should such a work-up lead to a diagnosis of endometrial 
carcinoma or uterine sarcoma, the subject will be taken off study and referred to a 
gynecologic oncologist for appropriate treatment.  Otherwise, their specific histologic 
diagnosis and the date of diagnosis will be recorded, but they may continue on study.  
 
Participants must not be planning to become pregnant for two years to be eligible for the 
study, and they will be warned about the importance of not becoming pregnant while on 
study at the time of enrollment. They will be advised to use either a barrier method or 
another nonhormonal form of contraception (eg. hormone-free IUD) for all sexual activity 
during the course of the study. Nevertheless, occurrence of an unplanned intrauterine 
pregnancy while on study will be an indication for immediately stopping treatment and 
the patient must withdraw from the study. If the patient should become pregnant while on 
study, or within two months after discontinuing therapy, her treatment assignment will be 
unblinded immediately. If the patient was randomized to tamoxifen, then they must be 
apprised of the potential risk of spontaneous abortion, birth defects, or a DES-like 
syndrome. 
 
The Principal Investigator (PI) at each site is responsible for monitoring protocol conduct 
and reporting to their Institutional Review Board (IRB) any adverse events (AEs) related 
to either the study medication per their institutional guidelines.  AEs must also be 
reported to the Coordinating Center (UAB) per instructions in Section 8.0.  The NCI 
common toxicity criteria for adverse events Version 4.0 (CTCAE v.4.0, Publish Date 
10/19/2009) will be used to assess toxicity.  
 
With the exception of vasomotor and gynecologic symptoms commonly associated with 
menopause, all grade 2 or greater toxicities of clinical significance with possible or 
greater attribution (please refer to Section 8.3 for definition) to the study drug will be 
reviewed by the Protocol Management Team (PMT) at the local site.  The PMT includes 
the consortium PI, the consortium co-investigator, and the CRA/CRN.   
 
Any grade 2 toxicity besides vasomotor and/or gynecologic symptoms commonly 
associated with menopause with probable or greater attribution to the study drug 
will be brought to the attention of the local PI (within 24 hours of knowledge of the 
event) for close monitoring.  If the symptom resolves or improves within 72 hours, 
the patient may continue on the study medication. Otherwise, the patient will be 
unblinded and taken off therapy.  
 
A patient reporting grade 3-4 toxicity with possible or greater attribution to the 
study drug will be unblinded and taken off protocol therapy. 
 
Serious adverse events (SAE) and all grade 3-4 toxicities with possible or greater 
attribution to the study drug must be submitted to the coordinating center PI and study 
staff immediately (within 24 hours of knowledge of the event), to allow reporting to the 
local and central IRBs and the central DSMB. If a Grade 3-4 AE is determined to have 
possible attribution to low-dose tamoxifen, an AE report will be submitted to the Food 
and Drug Administration (FDA) as described in Section 8.0. 
 
Because venous thromboembolic events have been associated with standard dose 
tamoxifen, we include CTCAE specific to DVT, PE, and stroke below.  
 
 
 

 
Protocol Version: 10/26/15 
25
CATEGORY: 
VASCULAR
Adverse Event: Thrombosis/embolism (vascular access-related) 
MedDRA Code: 10062169
 
  
Grade 
Description
1 
Venous thrombosis (e.g., superficial thrombosis) 
2 
Venous thrombosis (e.g., uncomplicated deep vein thrombosis), medical 
intervention indicated 
3 
Thrombosis (e.g., uncomplicated pulmonary embolism [venous], non-embolic 
cardiac mural [arterial] thrombus), medical intervention indicated 
4 
Life-threatening (e.g., pulmonary embolism, cerebrovascular event, arterial 
insufficiency); hemodynamic or neurologic instability; urgent intervention 
indicated 
5 
Death  
 
 
 
The following are considered clear indications for discontinuation of study protocol 
therapy (confirmed by clinical diagnosis):  
 
Deep venous thrombosis (DVT)  
 
Pulmonary embolism (PE) 
 
Transient ischemic attack (TIA) 
 
Cerebral vascular accident (CVA) 
 
Retinal vein thrombosis 
 
Cardiac ischemia/infarction 
 
6.4 
Study Stopping Rules 
 
6.4.1 Toxicity criteria for study stopping 
If two or more cases of the same related and unexpected grade 3-4 toxicity with 
probable or greater attribution are reported in separate patients, accrual will be 
held until further review by the central DSMB to determine whether or not the 
study should be stopped based on the unblinded data available at that time. 
Because low-dose tamoxifen is expected to have lower toxicity than standard-
dose tamoxifen, if two separate cases of grade 4 thromboembolism as listed in 
Section 6.3 occur and are attributed to the tamoxifen intervention in the 
unblinded DSMB review, with none observed in the placebo group, this will be a 
clear indication for stopping the study. If cases of grade 4 thromboembolic events 
are observed in the placebo group, then if occurrences in the tamoxifen group 
reach a risk ratio of 1.4, the trial will be stopped. Similarly, if 2.5-fold more 
endometrial cancers are observed in the tamoxifen group compared to the 
placebo group, the trial will be stopped. Both of these cut-offs are derived from 
the NSABP P-1 trial [33]. 
 
6.4.2 Efficacy criteria for study stopping 
We do not expect to see a greater efficacy of low-dose tamoxifen compared to 
the standard tamoxifen regimen for breast cancer risk reduction in any risk 
population. The NSABP P-1 trial found an overall magnitude of risk reduction of 
49% with the standard tamoxifen regimen of 20mg per day for 5 years (risk ratio 
0.51, 95% CI 0.39–0.66), with higher efficacy observed among women with 

 
Protocol Version: 10/26/15 
26
higher baseline risk. Specifically, the subgroup of women with a prior history of 
atypical hyperplasia had a reduction in breast cancer risk of 86% (risk ratio 0.14, 
95% CI 0.03–0.47) [33]. We believe that a similar risk reduction among radiation 
exposed CAYAC survivors would be of sufficient clinical significance to warrant 
early trial stopping and reporting of results. Thus, if a risk ratio of 0.14 or less is 
observed during DSMB review, the trial will be halted and subjects unblinded.  
 
 
7.0 
TREATMENT PLAN 
 
7.1 
Subject Treatment 
Subjects will be randomized 1:1 to receive low-dose tamoxifen 5 mg or placebo daily for 
24 months.  
Subjects will be instructed to swallow the capsule whole, with water or another non-
alcoholic liquid, preferably the same time each day. They may take the tamoxifen 
capsules with or without food. If they forget a dose, participants will be asked to take it 
when they remember, and then take the next dose as usual, but if they do not remember 
until their next dose, to not take extra capsules to make up the missed dose. 
For premenopausal subjects, they will be instructed to begin their study medication 
course while menstruating or after a negative beta-HCG or serum pregnancy test as 
below. For premenopausal subjects who do not have regular menstrual periods or are 
between periods, a beta-HCG level will be performed to confirm that the patient is not 
pregnant before starting study medication. Participants will be warned that they should 
avoid pregnancy while taking study medication and for two months after discontinuation. 
If they are sexually active, they will be advised to use barrier or nonhormonal 
contraceptive measures while on study.  
 
7.2 
Concomitant and Repeat Therapy 
 
No concomitant therapy is allowed with the exception of continued medications for 
chronic illnesses that are not excluded in Section 5.0, and necessary medications for 
unrelated acute illnesses that may occur during the study (cold, flu, infection, etc.). Any 
such medications must be recorded.  Ingestion of certain natural products should also be 
monitored while on trial, as outlined in Section 11.14.  
 
7.3 
Criteria for Removal from Treatment 
 
 
 
 
7.3.1   Treatment will be discontinued if a subject experiences an adverse event which 
the investigator deems related to the study medication or which will interfere with 
the ability of the subject to comply with the protocol.  See section 6.3 for specific 
rules.  
 
 
7.3.2    A subject may always voluntarily withdraw from treatment whenever she wishes. 
 
 
8.0 
TOXICITIES TO BE MONITORED AND DOSAGE MODIFICATIONS 
 
NCI CTCAE v.4.0 will be used to assess toxicity, study and medication stopping rules 
are outlined in Sections 6.3 and 6.4, respectively. The study will utilize Adverse Event 

 
Protocol Version: 10/26/15 
27
Collection Form for reporting all AEs and the FDA MedWatch for reporting serious 
adverse events as defined below.   
 
 
8.1 
Dosage Changes Based on Toxicity   
 
 
 
There will be no dose modifications on this trial.  See Section 6.3 for discontinuation 
rules.   
 
8.2 
Data Safety and Monitoring 
 
8.2.1 Monitoring and Personnel Responsible for Monitoring 
 
This is a multi-center phase II trial conducted at the University of Alabama at 
Birmingham and participating institutions, and monitored by the Clinical Trials 
Monitoring Committee (CTMC) housed at the UAB Comprehensive Cancer 
Center. 
8.2.2 Adverse Events: List and Reporting Requirements 
 
Adverse event (AE) monitoring and reporting is a routine part of every clinical 
trial. Serious Adverse Events (SAEs) are reported by the site PI within 24 hours 
to the coordinating center (survivorshiptrial@peds.uab.edu, phone: 205-638-
2127).  The Coordinating Center is then responsible for reporting SAEs to the 
UAB IRB and Clinical Trials Monitoring Committee in accordance with study-
specific requirements. SAEs occurring at participating sites are reported to the 
UAB IRB as “non-UAB” events. 
 
A Serious Adverse Event (SAE) is an AE that 1) results in patient hospitalization 
or prolongation of hospitalization; 2) results in persistent or significant disability or 
incapacity; 3) results in death; 4) is a cancer or congenital abnormality or 5) 
results in the development of drug dependence or abuse. An AE must be 
considered an SAE when the nature or severity of the event is not consistent with 
the current Investigator’s Brochure. Participating site SAEs must be reported by 
the site PI to the coordinating center as described above. It is also the 
responsibility of the site PI to report SAEs to the site IRB and to submit copies of 
that report to the coordinating center. It is the Coordinating Center’s  
responsibility to report the SAE to the appropriate regulatory agency and / or 
industry sponsor. This submission of IND Safety Reports will be cross referenced 
according to local regulations to Onyx Investigational Compound Number (IND) 
at the time of submission. 
 
 
8.2.3 Data and Safety Monitoring Plan 
 
 
 
Participants enrolled on this study will be monitored by the UAB Comprehensive 
Cancer Center’s Clinical Trials Monitoring Committee (CTMC). Adverse reactions 
observed during treatment will be closely monitored by the Clinical Trials 
Monitoring Committee (CTMC) on a monthly basis. The CTMC is responsible for 
data and safety monitoring of the trial and adherence to the DSMP, and is 
serving as the Central DSMB for this study. The independent Quality Assurance 
Committee (QAC) is responsible for oversight of the operation of CTMC, 
including adherence to the DSMP. Reports from the CTMC are reviewed monthly 
by the QAC. 

 
Protocol Version: 10/26/15 
28
 
9.0 
STUDY PARAMETERS AND CALENDAR   
These procedures are fully outlined in the study calendar and summarized below.  
9.1 
Screening visit   
After informed consent is obtained, a screening visit will be scheduled. This visit will 
include a history and physical, clinical breast exam, a complete blood count 
(includes platelet count, but no differential necessary), PT/PTT, metabolic panel 
(includes electrolytes, creatinine, and liver function tests), and a screening 
mammogram.  
 
For premenopausal patients, the screening mammogram should be performed between 
days 2 and 10 of their menstrual cycle (follicular phase). For premenopausal subjects 
who are unable to schedule a mammogram during the follicular phase of their menstrual 
cycle, the days since last menstrual period (LMP) will be recorded in the Breast Image 
Submission Form, and follow up mammograms will be scheduled on the same day of 
their cycle in which their screening mammogram was completed +/- 5 days.  All 
mammograms must be performed on a digital mammography machine using large 
paddles. Images should be taken using on-site mammography equipment (rather than 
off-site facilities), and serial measurements should be taken from the same 
mammography unit whenever possible. Each view from each breast must be saved as a 
single electronic image in raw DICOM (Digital Imaging and Communications in 
Medicine) format as instructed in Section 11.1.1. A site radiologist will perform a clinical 
reading of the screening mammogram; including a density score per American College 
of Radiology (ACR) BIRADS guidelines (Figure 9). If a screening mammogram was 
performed within 120 days of study registration as part of the patient’s routine clinical 
care, it can be estimated how many days into the menstrual cycle the patient was when 
it was performed (or the patient is postmenopausal), and it was obtained using a digital 
mammography unit, a digital copy of the original film will suffice, as long as that digital 
copy includes whole electronic images in raw DICOM format as outlined in Section 
11.1.1. In that case, a copy of the radiology report should be retrieved at the same time 
as the digital mammogram. If both clinical reading and clinical density assessment were 
provided on that report, that will suffice. Otherwise, the outside film must be submitted 
for a reading by a site radiologist who will provide a reading and density assessment 
according to Figure 9 in Section 5.2. Detailed instructions on electronic submission of 
study mammograms are in Section 11.1.1.  
 
For subjects whose menopausal status is unclear based on clinical history alone (see 
definitions in inclusion criteria below), a FSH level will be drawn at the time of the 
screening visit. For subjects who are premenopausal and have not had a menstrual 
period within 28 days, a blood beta-HCG test will also be performed.  
9.2 
Other baseline study parameters 
Screen-eligible patients will be scheduled for baseline RPFNA (for consenting patients at 
FNA-participating sites) between days 2 and 10 of their menstrual cycle (follicular phase) 
and phlebotomy for circulating biomarker assays (all sites) to be performed within 60 
days of completion of the screening visit (for patients consented prior to protocol version 
3/9/2015 and all previously consented patients as of 10/26/15). A urine collection and a 
blood draw after at least an 8 hour fast must also be scheduled within the same time 
window for lipid profile, anti-thrombin III, bone turnover markers, and circulating 

 
Protocol Version: 10/26/15 
29
biomarkers (including insulin growth factors, sex steroid hormones, CYP2D6 SNPs, and 
tamoxifen metabolites). Please refer to Sections 11.2 [RPFNA], 11.3 [blood] and 11.4 
[urine] for details regarding specimen collection, processing, and transfer.  
 
In addition, if the patient is followed with annual screening breast MRI as part of their 
standard of care (applies to patients who have survived at least 8 years beyond their 
chest radiation exposure; please refer to the COG cancer survivorship guidelines 
[www.survivorshipguidelines.org] for those diagnosed with CAYAC prior to age 21 or the 
NCCN guidelines [www.nccn.org] for those diagnosed after age 21), then the most 
recent pre-registration MRI must be obtained for submission with the baseline screening 
mammogram described above. Patient reported symptoms will be assessed via 
standardized Symptom Log.  
 
For sites who prefer to combine screening and baseline enrollment visits, the screening 
blood collection must be performed after an 8 hour fast and include the additional tests 
outlined in this section. The tubes drawn for most of the baseline tests below may be 
held until screen eligibility is confirmed; however, the anti-thrombin III level must be 
processed right away. The baseline RPFNA (for FNA-participating sites, for patients 
consented prior to protocol version 3/9/2015 and all previously consented patients as of 
10/26/15) can be tentatively scheduled to follow completion of all screening activities, 
including registration in the Interactive Web Response System (IWRS), as described in 
Section 13.0, and then cancelled if eligibility is not confirmed by normal breast exam, 
benign mammogram, breast density reading of 2-3, and lab tests within stated 
parameters (see Section 5.0), or postponed if more time is needed. Study drug must be 
available for the participant to take home with them on the day the last baseline activity 
is performed.  
9.3 
Treatment initiation 
The study CRA or CRN will register subjects as outlined in Section 13.0.  The CRA or 
CRN will perform the randomization using the Sharp Clinical Services IWRS according 
to specifications in Section 12.3.  Upon completion of the randomization, the Sharp 
Clinical Services IWRS will assign kit numbers to be dispensed.   Kit labels include 
instructions on how to properly store and take the capsules as well as a contact number 
should any concerns regarding either the study medication or procedures arise.  
 
9.4 
Follow-up visits 
For patients who consented to the RPFNA procedure (prior to protocol version 3/9/2015 
and all previously consented patients as of 10/26/15), the study CRA or CRN will follow 
up with a call after the first week on study (Day 8) to confirm that the baseline RPFNA 
was tolerated well All patients will be called after the fourth week (Day 28) to confirm that 
the patient is tolerating the medication well.  Patient reported symptoms will be assessed 
via standardized Symptom Log administered via phone interview. 
 
The patient will be asked to return their pill kits on Day 90 for a pill count (reported on 
the Adherence Tracking Form) and refill dispensal, and patient reported symptoms will 
be assessed via standardized Symptom Log administered via phone interview.  
 
A return visit will be scheduled for Day 180, during which a clinical breast exam will be 
performed and reported on a Clinical Breast Exam Form. Patient reported symptoms 
will be assessed via standardized Symptom Log form administered via in-person 

 
Protocol Version: 10/26/15 
30
interview. Patient reported health status changes will be assessed using the Health 
Assessment Form.  In addition, health-related quality of life (QOL) will be assessed 
using the patient self-administered NSABP Quality of Life Form.  A medication refill will 
be dispensed at that time, and pill count will be reported on the Adherence Tracking 
Form.   
 
A phone follow-up will be conducted 3 months later (Day 270), including pill kit collection 
for assessment of adherence via pill count pill count (reported on the Adherence 
Tracking Form) and another refill dispensal. Patient reported symptoms will be 
assessed via standardized Symptom Log administered via phone interview.  
 
A return visit will be scheduled for Day 365, during which a clinical breast exam 
(reported on a Clinical Breast Exam Form), fasting blood draw and urine collection, 
and patient reported symptom assessment will be performed via in-person interview 
documented on the Symptom Log form.  Patient reported health status changes will be 
assessed using the Health Assessment Form.  In addition, the patient will be given the 
self-administered NSABP Quality of Life Form to fill out.  All lab results will be reported 
using a Lab Values Form. A repeat mammogram using a digital mammography 
machine with large paddles recommended to be timed to the follicular phase of the 
menstrual cycle (Days 2-10) will also be performed. The patient will be encouraged to 
have their mammogram at the same facility that did the initial study mammogram. If 
indicated (per COG or NCCN follow-up guidelines), a screening breast MRI should also 
be performed. If done while on study, it is suggested to time it to the follicular phase of 
the menstrual cycle (Days 2-10). A medication refill will be dispensed at that time, and 
pill count will be reported on the Adherence Tracking Form.   
 
A phone follow-up will be conducted 3 months later (Day 455), including pill kit collection 
for assessment of adherence, collection of patient reported symptoms via standardized 
Symptom Log administered via phone interview, and refill dispensal, with pill count 
reported on the Adherence Tracking Form.   
 
Another in-person visit will be scheduled for Day 540 for clinical breast exam (reported 
on a Clinical Breast Exam Form) and assessment of patient reported symptoms via 
Symptom Log. Patient reported health status changes will be assessed using the 
Health Assessment Form.  In addition, the patient will be given the self-administered 
NSABP Quality of Life Form to fill out.  The pill kit will be collected for assessment of 
adherence via pill count pill count, reported on the Adherence Tracking Form, and 
another refill will be dispensed. 
 
This will be followed by a phone follow-up 3 months later (Day 630) for pill adherence 
(reported on the Adherence Tracking Form), patient reported symptom evaluation via 
Symptom Log, and the last refill dispensal.  
 
The final study visit will be conducted on Day 730, during which a final history and 
physical (reported on the General History and Physical Form), fasting blood draw, and 
urine collection will be performed.  For patients who completed a baseline RPFNA, a 
post-treatment RPFNA will also be performed.  Adherence will be reported on the 
Adherence Tracking Form, and patient-reported symptoms will be assessed via 
Symptom Log. Patient reported health status changes will be assessed using the 
Health Assessment Form.  All lab results will be reported using a Lab Values Form. A 
repeat mammogram using a digital mammography machine with large paddles and 

 
Protocol Version: 10/26/15 
31
timed to the follicular phase of the menstrual cycle (Days 2-10) will also be performed. 
The patient will be encouraged to have their mammogram at the same facility that did 
the initial study mammogram. If annual screening breast MRI is indicated (per COG or 
NCCN follow-up guidelines), a repeat breast MRI should also be performed. If done 
while on study, it is suggested to time it to the follicular phase of the menstrual cycle 
(Days 2-10). A final telephone follow-up will occur one week later (Day 738) to ensure 
that the post-treatment RPFNA was well tolerated (only for patients consenting to the 
RPFNA procedure before protocol version 3/9/2015 and all previously consented 
patients as of 10/26/15).  
 
All forms are due within 28 days of each scheduled visit and should be entered on the 
study Electronic Data Capture (EDC) website. Please refer to the Treatment Monitoring 
and Follow-up SOP for detailed instructions.  
9.5 
Long-term Follow-up 
After trial completion, the patient should be referred back to their local long-term 
survivorship clinic for continued follow-up, with their first return visit scheduled one year 
after their Day 730 study visit. Their first return visit will represent a new baseline 
survivorship clinic visit, and all organ function baseline assessments included in the 
COG survivorship guidelines should be repeated. This includes a comprehensive 
metabolic panel, which is recommended to be performed at baseline and as needed. 
Because of the association between standard dose tamoxifen and hepatic disease, 
special attention should be paid to symptoms and signs of hepatic disease on all 
subsequent annual follow-up survivorship clinic visits.  
 
To monitor for significant medical events, including second malignancies that may occur 
after the two-year intervention, sites will submit an Annual Follow-Up Form for up to 10 
years post-study completion, including participants that complete the two-year 
intervention and those that withdrew prior to completion. Subjects who are lost to follow-
up will be reported to the coordinating center using this form. New cancer and any 
inpatient or outpatient procedures potentially related to the study will be reported using 
the Breast Event Form and Other Event Form.  

 
Protocol Version: 10/26/15 
32
STUDY CALENDAR* 
Treatment plan:    
Prior to Treatment  
Days on Study Treatment 
Off 
STUDIES  
Screeni
ng 
Baseline 
(Day 0)** 
8 
28
90 
180 
270 
365 
455 
540 
630 
730 
738 
Primary Cancer  Treatment History  
X 
 
 
 
 
 
 
 
 
 
 
 
 
General History and Physical  
X 
 
 
 
 
 
 
 
 
 
 
X 
 
Health Assessment 
 
 
 
 
 
X 
 
X 
 
X 
 
X 
 
Breast Evaluations 
Clinical Breast Exam 
X 
  
 
 
  
X 
  
X 
  
X 
  
X 
  
Breast Imaging a 
X 
 
 
 
 
 
 
X 
 
 
 
X 
 
Breast Tissue Sampling  b 
 
X 
 
 
 
 
 
 
 
 
 
X 
 
Blood/Urine Tests 
CBC and Metabolic Panel c  
X 
 
 
 
 
 
 
X 
 
 
 
X 
 
PT/PTT (pre-RPFNA) 
X 
 
 
 
 
 
 
 
 
 
 
X 
 
Beta-Human Chorionic 
Gonadotropin (-HCG), if needed d  
X 
 
 
 
 
 
 
 
 
 
 
 
 
FSH level, if needed  e 
X 
 
  
  
 
 
  
  
  
 
 
 
 
Blood for circulating biomarkers  f  
 
X 
 
 
 
 
 
X 
 
 
 
X 
 
Fasting lipid profile g  
 
X 
 
 
 
 
 
X 
 
 
 
X 
 
Anti-thrombin-III  h  
 
X 
 
 
 
 
 
X 
 
 
 
X 
 
Blood and urine to assess bone 
turnover  i  
 
X 
 
 
 
 
 
X 
 
 
 
X 
 
Self-Administered Questionnaires 
Demographics and BC risk Form  J   
X 
 
 
 
 
 
 
 
 
 
 
 
 
Family History Form J 
X 
 
 
 
 
 
 
 
 
 
 
 
 
NSABP Quality of Life Form J 
 
X 
 
 
 
X 
 
X 
 
X 
 
X 
 
Treatment Management 
Study tablet dispensal K 
  
X 
 
 
X 
X 
X 
X 
X 
X 
X 
  
  
Study tablet collectionm 
 
 
 
 
 
X 
 
X 
 
X 
 
X 
 
Symptom log  J 
 
X 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Adverse Event Collection Form J 
 
 
 
 
 
X 
 
X 
 
X 
 
X 
 
Follow up phone call L 
 
 
X 
X 
X 
 
X 
 
X 
 
X 
 
X 
a Annual mammograms will be required as standard of care follow-up for this patient population per COG cancer survivorship or NCCN guidelines.  For 
patients who are at least 8 years beyond their RT exposure, annual screening breast MRIs are recommended. Both forms of breast imaging will be timed to 
the menstrual cycle and submitted electronically as described in section 11.1.1. 
b  Will be done via RPFNA described in section 11.2, for patients consented prior to protocol version 3/9/2015 and discontinued for all participants 10/26/15. 
c Platelet count should be included in the CBC panel, differential is not needed. The metabolic panel should include liver function tests.  
d Only if premenopausal with irregular menstrual cycles, to rule out pregnancy (see eligibility criteria).  
e Only if status-post hysterectomy without bilateral salpingo-oophorectomy, or does not have regular periods and pregnancy has been ruled out, to 
determine menopausal status (see eligibility criteria). 
f Insulin growth factor-1 (IGF-1) and insulin growth factor binding protein-3 (IGFBP-3) will be determined (assay methods described in Section 11.7). Future 
analyses of sex steroid hormone levels, CYP2D6 SNPs and tamoxifen metabolite levels are also planned.  
g Lipid profile includes total cholesterol, LDL-cholesterol, HDL-cholesterol, and triglycerides, determined using standard clinical assays. 
h Anti-thrombin III (AT-III) is a measure of clotting propensity (a lower level suggests higher risk of clotting).  
i Submit for serum bone-specific alkaline phosphatase (BSAP) and urine N-telopeptide crosslinks (NTX), which measure bone formation and bone 
resorption, respectively. 
J Included in the appendix. 
K The initial Day 0 pill kit will be dispensed at the time of blood and/or tissue sampling. The Day 180, 365, and 540 refills will be dispensed at the time of their 
follow-up clinical breast exams. Study capsules may be dispensed without waiting for lab clearance.  
L Follow up calls are to confirm the patient is tolerating the RPFNA and to determine adherence and arrange medication refill dispensal. 
m Patients will return 2 study kits at the time of their visit  
 * The protocol allows for a +/- 60 business day window for all in person visits and +/- 5 business days for scheduled phone visits.  
** The Day 0 visit (must be done within 60 days of completion of the screen visit if scheduled separately) 

 
Protocol Version: 10/26/15 
33
10.0    CRITERIA FOR EVALUATION AND ENDPOINT DEFINITIONS 
 
 
A subject will be considered evaluable if they complete at least one year on the study, 
including both baseline and one-year study procedures.  The outcome status (in terms of 
toxicity and biological response) of all eligible subjects will be reported.  Subjects who 
begin treatment will be accounted for in the summaries of patient reported outcomes, 
regardless if they are considered evaluable or not.  
 
10.1 
Parameters to be Measured Prior to Initiation of Treatment 
There will be two visits prior to the initiation of treatment: a screening visit to identify 
eligibility, and a baseline visit. With careful planning and coordination, these visits may 
be combined as described in Sections 9.1 and 9.2. At the screening visit, a history and 
physical, CBC (with platelets), comprehensive chemistry panel will be performed.  A 
mammogram will also be performed at that time. (If the patient had a digital 
mammogram using a large-paddle digital mammography machine performed within 120 
days, the patient is either postmenopausal or is premenopausal and can estimate the 
day in the menstrual cycle when the mammogram was performed, and a raw DICOM 
format image of that mammogram can be obtained, then their prior mammogram may be 
used as the study mammogram). A FSH and/or HCG level will be measured at that time 
if it is needed to determine menopausal status (see definition in Section 5.0). For 
confirmed screen-eligible subjects enrolled at FNA-participating sites, an RPFNA 
procedure will be performed within 60 days of the screening visit to establish baseline 
breast tissue markers (for patients consenting prior to protocol version 3/9/2015 and all 
previously consented patients as of 10/26/15). Blood will be drawn after an 8-hour fast 
and urine collected within 60 days of the screening visit to measure circulating 
biomarkers as outlined in the study calendar.  
 
10.2 
Specifications by Methods of Measurements 
The same method and laboratory will be used to characterize each identified and 
reported biological endpoint at baseline and during follow-up (see Section 11.0).  
 
10.3 
Parameters to be Measured During Treatment 
All evaluations will be measured per the study calendar schedule using the assay 
methodology outlined in Section 11.0. Analysis of results is fully described in Section 
12.0.   
 
The following are biological response endpoints: 
1. Mammographic Breast Density 
2. Tissue biomarkers of BC risk – Cytomorphology,  proliferative (Ki67) 
index, and apoptotic (caspase) index 
3. Insulin Growth Factors – IGF-1 and IGFBP-3 
 
The following endpoints are measures of safety and tolerability: 
1. Bone metabolism markers - serum bone specific alkaline phosphatase 
(BSAP) and urine N-telopeptide crosslinks (NTX)  
2. Fasting lipid panel – total cholesterol, HDL-cholesterol, LDL-
cholesterol, and triglycerides 
3. Clotting propensity – anti-thrombin III (AT-III) 
4. Symptom logs 
 

 
Protocol Version: 10/26/15 
34
 
11.0    SPECIAL INSTRUCTIONS 
 
11.1 
Mammographic Breast Density (MBD) 
Mammograms will be collected within 120 days prior to treatment initiation and at one 
and two years after initiation of treatment as described in the study calendar. After a 
clinical reading by a local radiologist, who will evaluate density according to the ACR 
BIRAD scoring system used to determine eligibility, MBD will be determined from digital 
mammogram images by study radiologists experienced in breast imaging. This density 
assessment method, fully outlined below, yields a value expressed as a continuous 
percentage, which is more sensitive to modulation by an intervention than the 
categorical BIRADS density score. Thus, while BIRAD score will be used for determining 
eligibility, percent breast density will be subjectively evaluated by study radiologists, and 
objectively measured using area- and volume-based mammographic density software.  
The area-based software measurement will be the primary endpoint.  MBD will be 
determined from each breast.  If the patient had a history of DCIS or IEN prior to study 
entry, or if one breast had a baseline BIRAD density score of 1, only the contralateral 
breast measurement will be used for analysis.  Otherwise, the two measurements will be 
modeled as described in the statistical section.  
 
11.1.1 Digital mammogram handling and transfer:   
Mammography images will be performed on digital mammography machines 
using large paddles. Serial mammograms for each patient will be performed at 
the same facility using the same machine whenever possible. Sites will save 
post-processed and pre-processed or raw mammography images whenever both 
are available.  The pre-processed or raw image is also called the “for processing” 
image, and may require coordination with radiology staff to obtain before being 
cleared from the PACS system. The post-processed image (also called “for 
presentation” image) is the file typically displayed by a standard PACS system. 
Each digital image (up to 8 mammogram files per patient time point) should be 
de-identified (headers excluded) and submitted on CD-ROM to the address 
below within 28 days of the screening, Day 365, and Day 730 visits.  
If the subject has been followed with screening breast MRI as part of their 
standard of care screening regimen, the full electronic MRI file from the last 
breast MRI procedure performed prior to starting study drug and any breast MRI 
procedures performed during the 2-year protocol treatment period will be saved 
in de-identified DICOM format. All clinical MRI series obtained during the pre-
treatment breast MRI procedure will be submitted at the same time as the 
screening mammogram. All clinical MRI series obtained during all breast MRI 
procedures performed on protocol treatment (up to two annual exams per 
participant) may be submitted at the end of study with the Day 730 mammogram. 
The CD-ROM will be labeled with the subject’s name, central participant ID, site, 
and the date of preparation of the CD-ROM. A Breast Imaging Submission 
Form will be submitted along with the CD-ROM, on which the CRA/CRN will 
specify the date(s) of procedure(s) and the imaging equipment used. The 
electronic MRI will only be referred to as needed in the case of a breast event on 
trial. The MRI may also be stored for future research.   

 
Protocol Version: 10/26/15 
35
Address for site submission of breast imaging:  
Smita Bhatia, MD, MPH 
ATTN: LDtam study coordinator 
UAB Division of Peds Hem/Onc 
1600 7th Ave S, Lowder 500Birmingham, AL 35233 
United States 
Phone: 205-638-2127 
Fax: 205-638-2121 
Email: Survivorshiptrial@peds.uab.edu  
 
After the CD-ROM arrives, study staff at the coordinating center will remove any 
electronically embedded patient identifiers and procedure dates from the header 
of each image file. Then, film IDs will be assigned to each mammogram image, 
consisting of the participant ID plus a dummy code in random order with respect 
to timing. The procedure date for each mammogram will be tracked in a 
database that documents which dummy code was assigned to the serial images. 
Anonymized films will sent to the study radiologists, organized in batches at 
multiple reading time points: when the 10th patient completes the study (initial 
data check), and subsequently when the 25th, 50th, 100th (halfway point), 150th, 
and the last patient completes the study.  
 
MBD will be assessed in 12 batches (2 batches at each of the 6 reading time 
points) that contain all raw (batch A) and processed (batch B) films collected for 
10-80 subjects (up to 320 films per reading time point).  Views for 10% of 
subjects in each batch will be included a subsequent batch to obtain a second 
reading to assess reader reliability.   
11.1.2 MBD assessment:   
MBD will be assessed by study radiologists  using previously published area [65] 
and volumetric [66] software methods.   
11.1.3 MBD quality assurance:   
Since all subject identifiers and dates will be removed from the DICOM file prior 
to transfer, study radiologists will be blinded to treatment arm as well as timing of 
the mammograms. To minimize random measurement error, all mammograms 
from each subject will be read in the same session, but they will be in random 
order. A random 10% of subject’s films will be read twice to estimate quality 
control, as described above. Results will be delivered electronically, according to 
the batching procedures described above.  
 
11.2     Breast tissue sampling 
Breast epithelia will be sampled at baseline and after the two-year treatment period. 
Normal breast epithelial tissue will be acquired via RPFNA, which is a well-tolerated 
procedure that has been found to yield adequate tissue for multiple biomarkers [52]. The 
RPFNA procedure is fully described below, as well as the assays planned for tissue 
specimens obtained. The tissue biomarkers will be considered as secondary endpoints.  
NOTE: The study was amended to discontinue this procedure for new patients 
consenting to the study (protocol version 3/9/2015) and all previously consented 
patients as of 10/26/15. 

 
Protocol Version: 10/26/15 
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11.2.1 RPFNA procedure 
 
 
After local anesthesia, a 21-gauge needle attached to a 10-mL syringe will be 
used to sample normal breast glandular tissue of the upper outer and upper inner 
quadrants by entering just immediately adjacent to the areola at approximately 3 
o’clock and 9 o’clock, varying the position slightly as needed to avoid superficial 
blood vessels. Tissue is probed deeply to sample the terminal lobular–duct unit, 
where most cancers are thought to arise, obtaining 8-10 aspirations per breast; 
half from the upper outer quadrant site and half from the upper inner quadrant 
site. After the procedure, cold packs are applied to the aspiration sites for 
approximately 10 minutes, the breasts and chest wall are firmly bound in gauze, 
and a tight-fitting sports bra was worn over the gauze wrap to minimize breast 
movement and to decrease the chance of hematoma formation. Performing the 
RPFNA procedure in this manner typically yields 1000 epithelial cells per 
cytology slide [94, 95]. Minimum epithelial cell yields to allow evaluation are 10 
for cytomorphology and 25 for IHC per slide. 
 
 
The RPFNA procedure is well tolerated by most women, with any temporary 
discomfort experienced typically relieved with acetaminophen. In a large study 
including 480 women, 408 of 439 women invited for a repeat aspiration (93%) 
agreed to a second procedure, with 225 (51%) doing so as soon as 6 months 
after the first procedure. Infection requiring oral antibiotics or large hematoma 
formation requiring surgical evacuation occurred in fewer than 1% of the 
procedures performed [94].  Women will be asked to stop any medication that 
they may be taking that might increase their chances of bruising for at least ten 
days prior to the procedure (aspirin, non-steroidal, anti-inflammatory, or anti-
coagulant medications, as well as any herbal supplements).  In addition, as long 
as there is no history of venous thromboembolism, they may be given vitamin K 
10mg to take daily for 3 days prior to the procedure to avoid bleeding and 
hematoma formation. Patients should be offered lorazepam 2 mg for anxiety and 
be instructed to take it 30 minutes before the procedure if they feel they need it. 
In that case, they should arrange for transportation and avoid driving that day. 
Also, the patient should be provided with four extra-strength Tylenol after the 
procedure in case of discomfort.  
 
11.2.2 RPFNA specimen collection 
 
 
All cells collected by the RPFNA procedures will be pooled into one tube 
containing 6 mL of Modified CytoLyt per the Specimen Handling SOP. Briefly, the 
sample is labeled with participant ID, date of procedure, and sample type 
(RPFNA) and then capped and gently inverted 2-3 times to mix. The sample is 
placed on a test tube rocker at low speed (if test tube rocker is not available, then 
leave tube in wet ice). Within 24 hours, the sample must be shipped with the 
RPFNA Cytology Sample Submission Form via FedEx priority overnight 
service on a cold pack to the central processing lab at KUMC at the address 
below.  
Address for site submission of RPFNA samples:  
Carol J. Fabian, M.D. 
c/o Trina Metheny  
University of Kansas Medical Center 
Breast Cancer Prevention Center Laboratory 

 
Protocol Version: 10/26/15 
37
3901 Rainbow Blvd 
1005 WHE, Mail Stop 3003 
Kansas City, KS 66160 
Phone:  913-588-3917 
Email: tmetheny@kumc.edu 
 
11.2.3 Processing the cytology sample at the central pathology lab:  
 
Upon arrival to the KUMC central processing lab, one third of the CytoLyt 
suspended cellular sample will be processed for cytomorphology, and one third 
for evaluation of proliferative index and apoptotic index via Ki67 and caspase 
immunohistochemistry (IHC) staining, respectively. The remainder of the sample 
will be designated for tissue banking will be stored at -80C in RNAlater®. The 
cytomorphology sample will be processed first as outlined below, and that will be 
used to determine how the IHC slides will subsequently be prepared.  
 
Cytomorphologic preparations are filtered through a 25-mm Millipore filter (5-m 
pore size) and cytocentrifuged, rather than smeared, to reduce cellular distortion 
and maximize cell yield per slide. If there are fewer than 10 cells on the 
cytomorphology slide, this will be noted in the pathology report, and the sample 
will be considered unevaluable for cytomorphology. The method for 
cytomorphologic assessment is described in Section 11.5.1.  
 
Slides designated for IHC are sequentially fixed in 10% buffered formalin, 
methanol, and acetone. If there were fewer than 100 cells on the cytomorphology 
slide, the aliquots designated for Ki67 and caspase will be combined and used 
together for determination of proliferative index via IHC. In that case, the sample 
will be considered unavailable for apoptotic index. Proliferative index (and 
apoptotic index, when available) will be read as the number of cells with 
unequivocal nuclear staining per total number of ductal epithelial cells. If there 
were fewer than 500 cells on the slide, but more than 100, results will be 
considered evaluable but suboptimal. If there were fewer than 100 cells on the 
slide, that IHC measure will be considered unevaluable. Both of these IHC 
measures a described in detail in Sections 11.5.2 and 11.5.3, respectively.  
 
RPFNA specimens processed in this manner typically result in slides with 
approximately 1000 epithelial cells per slide. Thus, we expect that 
cytomorphology, Ki67, and caspase, will be evaluable from nearly all breast 
epithelial specimens obtained by RPFNA.  
 
11.3     Blood collection and processing  
Peripheral blood will be sampled at baseline and annually while on treatment. For 
premenopausal participants, blood draws will be timed to occur during the follicular 
phase of the menstrual cycle (Days 2-10). All subjects will be asked to fast for at least 8 
hours prior to blood draw.  
Whole blood will be drawn into the following tubes: 
 
one 10 mL EDTA-containing lavender-top  
 
one 5 mL citrate-containing blue-top 
 
one 10 mL red -top vacutainer tube 
 
two 10 mL red tiger-top vacutainer SST tubes 

 
Protocol Version: 10/26/15 
38
All tubes are gently inverted 2-3 times after being drawn. Two of the red tiger-top tubes 
will require processing at the local site and should be placed on wet ice immediately 
after blood draw for transport to the specimen processing center. Specimen processing 
and handling are fully detailed in the Specimen Handling SOP, and briefly discussed 
below.  
11.3.1 Processing and handling the K2EDTA-coated lavender-top tube  
 
 
This tube should be inverted 8-10 times and then labeled with type of sample 
(whole blood), participant ID, and date of blood draw. Pre-printed cryogenic 
labels with the sample type will be provided. PLEASE WRITE IN THE PTID AND 
THE DATE OF THE SAMPLE WITH A FINE TIP SHARPIE PERMANENT 
MARKER. It will then be shipped ON DRY ICE (≥10 pounds of dry ice) in the 
provided shipping container via FedEx priority overnight service (ship Monday 
thru Wednesday only), accompanied by the Blood and Urine Specimen 
Submission Form. If there is a delay between blood draw and shipment, the tube 
must be kept frozen at -80°C. Do NOT store the blood at -20°C for any length of 
time. This sample will be used for pharmacogenetic studies.  
Address for site submission of blood and urine samples. The study coordinator 
will document the receipt of the samples and transfer to the coordinating center 
laboratory:  
Smita Bhatia, MD, MPH 
ATTN: LDtam study coordinator 
UAB Division of Peds Hem/Onc 
1600 7th Ave S, Lowder 500 
Birmingham, AL 35233 
 
 
 
Phone: 205-638-2127 
Email: Survivorshiptrial@peds.uab.edu  
 
11.3.2 Handling the blue-top tube  
 
 
The blue-top tube should be inverted 2-3 times to gently mix the blood in citrate 
and immediately transported to the CLIA-approved hematology laboratory at the 
local site for determination of anti-thrombin III (AT3) per institutional protocol.  
Results will be entered into a secure study website.  
 
11.3.3 Processing and handling the red-top tubes:  
 
 
The red-top tube will be immediately transported to the CLIA-approved 
chemistry laboratory at the local site for determination of a fasting lipid profile per 
institutional protocols.  Lipid profiles consist of triglycerides (TG), total cholesterol 
(TC), high-density lipoprotein (HDL), and low-density lipoprotein (LDL).  Results 
will be entered into a secure study website. 
 
 
The two red tiger-top tubes should be inverted five times immediately after 
blood draw to ensure mixing of clot activator with blood (do not shake 
vigorously). Label the tubes with participant ID and date of blood draw using a 
permanent marker. Tubes should be allowed to clot at room temperature, upright 
in a test tube rack for 30 minutes. The sera is then harvested by spinning in a 
balanced centrifuge at 1200 x g for 10 minutes. Sera is aliquoted into up to 10 
study-labeled cryovials with 1 ml per aliquot and placed in a -80°C freezer within 
2 hours of collection time. All aliquots will be labeled with the provided cryogenic 

 
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labels (pre-printed with the aliquot number and sample type). PLEASE WRITE IN 
THE PTID AND THE DATE OF THE SAMPLE WITH A FINE TIP SHARPIE 
PERMANENT MARKER. Frozen serum samples should be shipped on DRY ICE 
(≥10 pounds of dry ice) via FedEx priority overnight service (ship Monday thru 
Wednesday only) to the coordinating center using the provided shipping 
container, FedEx shipping label, and accompanied by the Blood and Urine 
Specimen Submission Form. 
11.3.4 Storage of blood samples  
The whole blood and serum samples will be transferred to the UAB Institute for 
Cancer Outcomes and Survivorship laboratory, where they will be bar-coded, 
entered into a specimen tracking database, and stored at -80°C until assays are 
performed.  
 
11.4 
Urine collection and processing  
A spot urine sample will be collected at baseline and annually while on treatment. Two 
aliquots (3.5mL each) will be frozen at -20°C and then shipped on dry ice as described in 
the Specimen Handling SOP to the address in Section 11.3.1, accompanied by a Blood 
and Urine Specimen Submission Form.  This sample will be used for evaluation of N-
telopetides (NTX) as described in Section 11.8.2.  
 
11.5     Tissue-based biomarkers  
 
11.5.1 Cytomorphologic scoring 
Cytomorphology will be scored according to the Masood classification from 
epithelial cells retrieved via RPFNA. This scoring system assigns a score of 1 to 
4 points to each of 6 morphologic characteristics: cellular arrangement, cellular 
pleomorphism, prevalence of myoepithelial cells, anisonucleosis, nucleoli, and 
chromatin clumping. A score of 6 to 10 is associated with nonproliferative 
specimens, 11 to 14 with hyperplasia without atypia, 15 to 18 with hyperplasia 
with atypia, and 19 to 24 with malignancy (Table 2) [99]. For cytology, 
intraobserver variance has been reported to range from 8-24% [100]. Slides will 
be hand scored by two readers. In the case of a difference between the two 
readers, the scores will be averaged. The first reading and second reading will be 
performed by KUMC-associated cytopathologists. De-identified stained slides will 
be sent to each reader for blinded review:  
 
11.5.2 Determination of tissue proliferative index 
Proliferative Index will be determined using an immunohistochemistry (IHC) 
assay to evaluate Ki-67 expression from epithelial cells contained in RPFNA 
samples. A categorical estimate of the number of ductal epithelial cells present 
on the slides designated for proliferative index evaluation will first be assessed: 
less than 100, 100 to 500, 500 to 1,000; 1,000 to 5,000; or more than 5,000. 
Slides containing at least 100 epithelial cells will be processed for Ki-67 staining; 
if no slides meet this criterion, the subject will be deemed unevaluable for this 
endpoint at that time point. Antigen retrieval will be performed with 10 mmol/L 
citrate buffer (pH 6) and then will be stained with Ki67 clone MiB-1 monoclonal 
antibody (Dako North America, Carpinteria, CA) at a 1:20 dilution. Hyperplastic 
cell clusters will be preferentially assessed for nuclear staining. The number of 
cells with unequivocal nuclear staining out of 500 cells assessed will be recorded 

 
Protocol Version: 10/26/15 
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as the proliferative index. Slides will be hand scored by two readers as described 
in Section 11.5.1, and in the case of a difference between the two readers, the 
scores will be averaged. Agreement between two readers using this method is 
excellent (Cronbach’s  = 0.99; reported in [86]). Within-person reliability is also 
superb, with a previously reported intraobserver variance of 4% [100]. 
11.5.3 Determination of tissue apoptotic index 
RPFNA cytology samples will be stored for future analysis of tissue apoptotic 
index. Apoptotic index will be determined using an IHC assay to evaluate cleaved 
caspase-3 expression from epithelial cells contained in RPFNA samples. After 
categorical estimation of the number of ductal epithelial cells present on the 
slides designated for apoptotic index, antigen retrieval will be performed as 
described above and slides stained with cleaved Caspase 3 rabbit polyclonal 
antibody (Cell Signaling Technologies, Boston, MA). The number of cells 
positively stained out of 100 will be recorded as the apoptotic index, with the 
subject deemed unevaluable for this endpoint if less than 100 cells are present 
on the slide. As above, slides will be scored by two readers as described in 
Section 11.5.1, and scores will be averaged if discrepant.  
 
11.6     Estrogen Assays 
Serum and RPFNA samples collected into saline will be stored at the coordinating center 
for future analysis of sex steroid hormone levels.  At the time of analysis, the samples 
will be shipped to the Reproductive Endocrinology Research Laboratory of Dr. Frank Z. 
Stanczyk at the University of Southern California. The methodology used is a 
radioimmunoassay (RIA) after a unique in-house extraction using column 
chromatography as outlined below. This technique is highly sensitive for monitoring 
circulating estrogens in postmenopausal women.  
 
Total Estradiol (E2) – One-tenth of a ml of serum is incubated with 3H-E2 and extracted 
with hexane and ethyl acetate to remove unconjugated steroids. After evaporating under 
nitrogen, the residue is redissolved in isooctane and applied on a column of Celite 
impregnated with ethylene glycol. E2 is eluted in ethyl acetate in isooctane and 
redissolved in buffer. Then it is incubated with 125I-E2 and anti-E2 serum. Antibody-bound 
and unbound 125I-E2 are separated by adding a second goat anti-rabbit antibody, 
centrifuging, and aspirating the supernatant. Total estradiol is then quantitated using a 
gamma counter.   
 
Bioavailable E2 – is calculated by determining the sex hormone binding globulin (SHBG) 
level and subtracting that from the total estradiol level. SHBG is measured by a solid-
phase, two-site chemiluminescent immunoassay using the Immulite analyzer. Alkaline 
phosphatase conjugated anti-SHBG polyclonal antibodies are introduced into the 
reaction tube, and the tube is incubated for 30 minutes at 37C. SHBG in the sample is 
bound, forming an antibody sandwich complex. Unbound conjugate is then removed by 
a centrifuge wash, after which the chemiluminescent substrate is added, and the 
reaction tube is incubated for another 5 min. The chemiluminescent substrate undergoes 
hydrolysis in the presence of alkaline phosphatase, yielding an unstable intermediate. 
Continuous production of this intermediate results in the sustained emission of light. The 
light is measured by an illuminometer and is proportional to the concentration of SHBG 
in the sample 
 

 
Protocol Version: 10/26/15 
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Estrone (E1) – is determined using 1 ml of serum, which is incubated with 3H-E1 and 
treated with the same extraction and column chromatography methods as with total 
estradiol but using 125I-E1 and anti-E1 serum with a second goat anti-rabbit antibody. 
Estrone is quantitated similarly after centrifugation and separation.   
 
Estrone Sulfate (E1S) - One-third of a ml of plasma is incubated with 3H-E1S and is 
extracted with hexane and ethyl acetate, then deproteinized with methanol. After 
evaporating under nitrogen, the residue is reconstituted with sodium acetate buffer and 
hydrolyzed with arylsulfatase. The hexane:ethyl acetate extraction step is repeated, the 
extract is redissolved in buffer, and then the E1 RIA is carried out.  
 
The assay sensitivities are as follows: E2 = 5 pg/ml = 1.36 pmol/L, E1 = 5 pg/ml = 1.35 
pmol/L, and E1S = 0.16 ng/ml. Intra-assay and inter-assay coefficients of variation (CV) 
for all four assays are well below 15%, as tabulated below. 
 
Assay 
Intra-assay CV 
Inter-assay CV 
Total Estradiol 
7.0% 
9-13% 
Sex Hormone Binding Globulin 
4.1-7.7% 
6-13% 
Estrone 
7.4% 
8-10% 
Estrone Sulfate 
7.5% 
11% 
 
11.7     Insulin Growth Factors (IGFs) 
These tests will be performed by ARUP Laboratories at the University of Utah. ARUP is 
a national full-service reference laboratory that serves more than 50 academic medical 
centers, as well as many community hospitals, commercial laboratories, military and 
government facilities, and pharmaceutical firms. One 1mL aliquot will be sent by the 
coordinating center to ARUP Laboratories for measurement of the following IGFs. A 
second 1mL aliquot will be held for IGF evaluation in the case extra serum is needed.  
 
11.7.1 IGF-1 will be measured as circulating markers of BC risk. The assay will be 
conducted using Chemiluminescent Immunoassay. The sensitivity of the assay is 
5.1 ng/mL. The expected postmenopausal range is 58-318ng/mL. The expected 
premenopausal range is 89-397ng/mL.  
 
11.7.2 IGFBP3 will be measured as circulating markers of BC risk. The assay will be 
conducted using Chemiluminescent Immunoassay. The sensitivity of the assay is 
38.2 ng/mL. The expected postmenopausal range is 2514-6014 ng/mL. The 
expected premenopausal range is 2926-5858ng/mL. 
11.8     Bone metabolism marker assays 
 
Blood and urine will be collected at baseline and at the one-year and two-year visits at 
the local site and sent directly to ARUP Laboratories per instructions in Sections 11.3 
and 11.4 (see address in Section 11.7).   
 
11.8.1 Serum bone-specific alkaline phosphatase (BSAP) will be measured as a marker 
of bone formation. It is an isoenzyme of alkaline phosphatase localized on the 
cell membrane of osteoblasts. The assay employs a solid phase two-site 
immunoradiometric assay (IRMA). The sensitivity of the assay is 2.0 µg/L. The 
expected postmenopausal range is 7.0-22.4 µg/L. The expected premenopausal 
range is 4.5-16.9 µg/L. 

 
Protocol Version: 10/26/15 
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11.8.2 Urine N-telopeptide crosslinks (NTX) will be measured as a marker of bone 
resorption. Type I collagen comprises 90% of organic bone. It is a helical protein 
that is cross-linked at the N-terminal and C-terminal ends. These crosslinks are 
released during bone remodeling by osteoclasts. They are then found in the urine 
as a stable end-product of bone metabolism. The procedure is chemiluminescent 
immunoassay. Final values are reported as bone collagen equivalents (BCE) in 
nM/L corrected for creatinine clearance in mM/L. The expected range in 
postmenopausal women is 26-124 nM BCE/mM creatinine. The expected 
premenopausal range is 17-94 nM BCE/mM creatinine. 
 
11.9 
Lipid assays 
Blood will be drawn into a red-top tube at baseline before the first dose of treatment and 
then annually (Day 365 and 730), each time after an 8-hour fast.  This blood sample will 
be sent directly to the CLIA-approved chemistry lab for determination of triglycerides 
(TG), total cholesterol (TC), high-density lipoprotein (HDL), and low-density lipoprotein 
(LDL).  Results will be faxed back to the coordinating site.   
 
Although lipid profile abnormalities alone will not be considered an indication for stopping 
study protocol therapy, lipids will be monitored and treated according to NCEP ATP 
guidelines (http://www.nhlbi.nih.gov/guidelines/cholesterol/atglance.pdf).   
 
11.10   Clotting assays 
Blood will be collected into at baseline and at the one-year and two-year visits at the 
local site and sent directly to the CLIA-approved hematology laboratory at each local site 
for determination of anti-thrombin III per institutional protocol. Results will be entered into 
the study EDC website. 
 
11.11   Pharmacokinetic and pharmacogenetic measures 
Plasma will be processed from each scheduled blood draw and will be stored for the 
measurement of tamoxifen levels and levels of the tamoxifen metabolites, N-desmethyl-
tamoxifen (also known as endoxifen) and 4-OH-tamoxifen, at the time of trial completion.  
Blood will be also processed for DNA to allow for analyses of P450 SNPs including 
CYP2D6 polymorphisms after trial completion and unblinding. Stored DNA may also be 
used to explore DNA adduct formation during the course of tamoxifen treatment.  
The SNP studies will be performed using commercial assays developed by Roche 
Laboratories.  
 
11.12 Specimen storage 
Unused biological materials will not be discarded, and will be stored for possible future 
research purposes in the UAB Institute for Cancer Outcomes and Survivorship 
laboratory. IRB approval will be sought prior to their use for purposes other than those 
specifically outlined in this protocol. In addition, ancillary studies must be approved by 
the Consortium for Pediatric Interventional Research (CPIR) and all other participating 
institutions.  
 

 
Protocol Version: 10/26/15 
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11.13 Questionnaire data 
Self-reported and CRA/CRN administered questionnaires will be used to obtain baseline 
data on demographics, family history, reproductive history, and other BC risk factor 
exposure, as well as to assess patient reported symptoms during the course of the 
study. The self-reported questionnaires (Demographic and Breast Cancer Risk 
Factors Form, QOL Form, Family History Form and the Symptom Log) are 
considered source documents themselves and are not required to be accompanied by a 
source document, because of the design of the study.  Utilizing the study EDC website, 
these questionnaires will be entered electronically, in compliance with FDA Regulation 
21 CFR Part 11.  
Patient demographics and baseline BC risk factors:  
Standardized questionnaires will be used to obtain the following prior to study entry: 
 demographic reproductive history, and BC risk factors information 
(Demographics and Breast Cancer Risk Factors Form),  
 health and physical (H&P Form) 
 family history (COH DPS Family History Form),  
The salient domains captured by these questionnaires include date of birth, gender, 
race/ethnicity, socioeconomic status, and both familial and environmental BC risk 
factors. This questionnaire will be administered at the time of initial registration.  
 
Clinical data regarding prior Cancer diagnosis and treatment:  
The following information will be obtained from medical records using a standardized 
data abstraction form (the Primary Cancer Treatment History Form): date of 
diagnosis, stage of disease (with or without “b” symptoms), histological subtype, data 
elements regarding chemotherapy including dates, protocols/regimens, cumulative 
doses of therapeutic agents per square meter; and data regarding radiation therapy 
including dates, total lifetime dose, field, fractions, dose per fraction. A copy of 
institutional radiation oncology summary report will be obtained whenever possible. 
 
 
Patient reported symptoms and adherence to the chemopreventive agent:  
The National Surgical Breast and Bowel Project (NSABP) developed a standardized 
quality of life (QOL) questionnaire to assess patient reported symptoms in response to 
hormonal interventions, which they used in the NSABP P-1 and P-2 breast cancer 
prevention trials (NSABP QOL questionnaire).  The most recent version of this 
questionnaire, which was used in the NSABP P-2 trial [101], will be used. The salient 
domains captured by this questionnaire include both physical and mental well-being. In 
addition, a Symptom Log including the HEAL hormone-related symptoms instrument 
will be used [102]. This questionnaire captures general symptoms along with specific 
hormone-related symptoms along the following scales: vasomotor, vaginal, incontinence, 
cognitive/mood, and weight gain/appearance. Both questionnaires will be administered 
at baseline and during the two-year treatment period according to the study calendar. Pill 
counts will also be used to determine adherence to the prescribed intervention using the 
Adherence Tracking Form. 
Patients who report intolerable hot flashes may be managed with any of the following 
interventions (please refer to the Treatment and Follow-up SOP for recommended 
algorithm):  
 
Vitamin E 800 IU po daily 
 
Effexor (venlafaxine) XL 37.5-75 mg po daily  

 
Protocol Version: 10/26/15 
44
 
Neurontin (gabapentin) 300mg po TID  
 
Lyrica (pregabalin) 75mg po BID 
 
Patients who report vaginal atrophic symptoms may be managed with any of the 
following interventions (please refer to the Treatment and Follow-up SOP for 
recommended algorithm):  
 
Replens 
 
Astroglide  
 
Vagifem 
 
Estring 
 
11.14 Natural products known to modulate estrogens 
The following herbal supplements have been associated with hormonal activity. The 
primary endpoint, mammographic density, is positively correlated with estrogen levels. 
Therefore, subjects will be asked to avoid these items before during the treatment 
period. Any use will be recorded along with medication changes during the course of the 
trial.  
 
 
Alfalfa 
 
Bee Pollen 
 
Black Cohosh  
 
Blue Cohosh  
 
Cats Claw 
 
Chasteberry 
 
Chysin  
 
Coumestans 
 
Cramp Bark 
 
Dong Quai, or Chinese       
      Angelica 
 
DHEA 
 
Evening Primrose 
 
Feverfew 
 
Flaxseed 
 
Ginseng 
 
Hops 
 
Isoflavones 
 
Kava  
 
Kudzu  
 
Licorice   
 
Lignans 
 
Passionflower  
 
Propolis  
 
Red clover or other clover extracts  
 
Sarsaparilla  
 
Saw palmetto  
 
Shepherds purse  
 
Soy extracts 
 
St. Johns Wort  
 
Valeriana  
 
Wild yam  
 
11.15 Drugs described to modulate CYP2D6 
The following drugs have been reported as inhibitors or inducers of CYP2D6, which is 
the major P450 enzyme that metabolizes tamoxifen. Concomitant use of strong and 
clinically significant moderate inhibitors will not be allowed while on trial. Other moderate 
inhibitors should be avoided while on study.  
Strong CYP2D6 inhibitors  
(not allowed on study): 
bupropion (Wellbutrin) 
fluoxetine (Prozac) 
paroxetine (Paxil) 
quinidine (Quinidex) 
 
Clinically significant  
moderate CYP2D6 inhibitor  
(not allowed on study): 
duloxetine (Cymbalta) 
 
Other moderate CYP2D6 
inhibitors  

 
Protocol Version: 10/26/15 
45
(should be avoided while on 
study): 
sertraline (Zoloft) 
terbinafine (Lamisil) 
 
Weak CYP2D6 inhibitors  
(will be tracked on study): 
amiodarone (Cordarone) 
cimetidine (Tagamet) 
thioridazine (Mellaril) 
 
 
Possible CYP2D6 inhibitors  
(will be tracked on study): 
celecoxib (Celebrex) 
chlorpheniramine* 
chlorpromazine (Thorazine) 
citalopram (Celexa)  
clemastine (Tavist)* 
clomipramine (Anafranil) 
diphenhydramine (Benadryl)* 
doxepin (Sinequan) 
doxorubicin (Adriamycin) 
escitalopram (Lexapro)  
halofantrine (Halfan) 
haloperidol (Haldol) 
hydroxyzine (Vistaril, Atarax)* 
levomepromazine (Nozinan) 
methadone 
metoclopramide (Reglan) 
midodrine (ProAmantine) 
moclobemide (Aurorix, Manerix) 
perphenazine (Trilafon) 
ranitidine (Zantac)* 
ritonavir (Norvir) 
ticlopidine (Ticlid) 
tripelennamine (Pyribenzamine) 
other histamine H1 receptor antagonists** 
Possible CYP2D6 inducers  
(will be tracked on study): 
dexamethasone (Decadron, 
Hexadrol) 
rifampin (Rifadin, Rimactane) 
 
Virtually no effect on CYP2D6: 
venlafaxine (Effexor) – preferred 
antidepressant while on study 
 
*available over the counter 
**examples: cetirizine (Zyrtec), dimenhydrinate (Dramamine), doxylamine (included in NyQuil), 
fexofenadine (Allegra), loratidine (Claritin), meclizine (Antivert, Dramamine [less drowsy 
formulation]), pheniramine (Avil) 
 
11.16 Other drugs to avoid while on trial 
 
Warfarin or other coumarin-type anticoagulants 
 
Systemic exogenous hormonal agents, including 
o Other selective estrogen receptor modifiers, such as raloxifene (Evista). 
o Systemic hormone replacement therapy (includes oral or transdermal 
formulations). Vagifem and Estring, two formulations of locally applied vaginal 
estrogen associated with minimal systemic absorption, may be allowed. 
Other estrogen-containing vaginal creams, while not an exclusion, should be avoided 
whenever possible.  Patients with a history of hormone modifying herbal supplements 
(see section 11.3) are eligible, but patients will be asked to avoid their use after on 
study.  
o Hormonal forms of contraception (includes oral, transdermal, implanted, and 
injectable formulations),  
o Aromatase inhibitors,  
o GnRH analogs,  
o Prolactin inhibitors, and 
o Androgens or antiandrogens.  
 
 

 
Protocol Version: 10/26/15 
46
12.0 
STATISTICAL CONSIDERATIONS 
 
12.1 
Study design  
This is a Phase IIb randomized placebo controlled clinical trial.  
 
 
 
12.2 
Accrual plan 
This study is expected to accrue 230 evaluable subjects.   
12.3 
Stratified randomization 
Central participant IDs will be generated by the coordinating center and will be issued to 
sites as they are activated. Participant IDs for all sites will be sent to Sharp Clinical 
Services, along with the site names and strata as defined below. Sharp Clinical Services 
designed a system (IWRS) that will manage blocked stratified randomization, with 
menopausal status (2 strata: pre- and post-menopausal), radiation dose to the chest (2 
strata: 1200-2599 cGy, ≥2600 cGy), and age at radiation exposure (2 strata: <18y, 18-
40y) as stratification factors (8 strata total) and a block size of 4 to balance the number 
of participants in each arm. Blinded study drug will be shipped to sites as they are 
activated by Sharp Clinical Services. Once a subject has completed all screening 
assessments and is eligible, the site CRA or CRN will randomize the subject to low-dose 
tamoxifen or placebo using the Sharp Clinical Services IWRS, which will assign a 
blinded study kit. This will generate an e-mail notification to the site and the coordinating 
center that randomization has been completed (as described in Section 9.3).  
 
12.4 
Data analytic plan 
Aim 1: To determine the impact of low-dose tamoxifen on surrogate endpoint 
biomarkers of BC risk.  
Mammographic breast density (MBD). MBD is the ratio of absolute density to the total 
breast area. Since participants are randomized, we expect MBD to not differ by 
treatment status at baseline (t0). However, we will evaluate the success of 
randomization by examining between group differences in demographic and clinical 
characteristics (i.e. age, radiation dose, age at radiation exposure), study site, as well as 
the average MBD of the two breasts (or the available one) at baseline. If imbalance is 
evident, analysis will take into account the covariates with significant imbalance. 
Using an intention-to-treat analysis, the efficacy of low dose tamoxifen in reducing MBD 
will be compared between patients in the low dose tamoxifen intervention and placebo 
group by applying the linear mixed effects model for bivariate normally distributed data 
[103]. All patients with a minimum of baseline (t0) mammographic data will be included 
in the analysis. MBD data from each breast will be square root transformed (RTMBD) to 
normality before model fitting. RTMBD for each breast will be modeled by the 
unstructured mean model using two indicator variables of time. We will assume the 
effects of time on RTMBD to be the same in both breasts by setting their coefficients 
equal. Correlation between the breasts will be introduced by assuming a bivariate 
normal random intercept model with a common fixed effects intercept and breast-specific 
random effects. Residual errors will be assumed to be bivariate normally distributed with 
mean zero and non-zero correlation. Unstructured and compound symmetry covariance 
structures will be considered for residual errors. The fixed effects intercept will be 
allowed to differ between treatment groups. Interactions of the time indicators with 
treatment indicator (0 for placebo, 1 for tamoxifen) will be included to allow for treatment 

 
Protocol Version: 10/26/15 
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differences at t1 and t2, and their significance tested using a 2-df chi-square test. Low 
dose tamoxifen will be considered effective if 1) the 2-df test is significant and the 
expected RTMBD is lower both at t1 and t2 or at t2 compared to that of the placebo 
group, or 2) the interaction term at t2 is significant and the expected RTMBD there is 
lower than that of placebo. It should be noted that because common fixed effects for 
intercept and time coefficients are assumed for both breasts, the expected RTMBD is 
the same for both breasts within a treatment group. This model enables use of the data 
from both breasts to estimate the common treatment effect while accounting for the 
between-breast correlation and the correlation among repeated measurements over time 
within a woman. The validity of these restrictions, e.g. common intercept and treatment 
effect between left and right breast, while biologically reasonable, will be examined and 
tested. This method also enables women with incomplete RTMBD data, i.e. data from 
only one breast or missing data at specific time points, to be included in the analysis. In 
addition to the unstructured mean model, we will also fit a linear model in time, with an 
interaction of time by group indicator to allow for treatment difference. A significant time 
by group interaction, with a larger negative slope for the low dose tamoxifen group 
compared to placebo, will indicate the effectiveness of low dose tamoxifen in reducing 
MBD. 
In addition to unadjusted analyses which are appropriate under the randomization 
principal, adjusted analysis also will be performed. Significantly imbalanced covariates 
and covariates known to be prognostic for BC, such as radiation dose, age at irradiation, 
premature menopause, and family history for BC, will be incorporated in the longitudinal 
models before including and testing the time by treatment group interaction. 
To address potential noncompliance, we will also conduct an as-treated analysis, 
considering patients’ actual treatment uptake. Instead of the treatment indicator (PBO or 
LDTAM), we will include adherence rate, i.e. 0 for PBO and a number between 0 and 1 
for LDTAM determined by pill count, in the LME model. Time-specific adherence rate 
and the overall adherence rate will be considered. If LDTAM is effective, we may expect 
to see a decrease in MBD with adherence rate. 
The estimates from linear mixed effects model are valid in the presence of incomplete 
response data if missing data occurs at random. However, non-participation and 
dropouts may occur differentially between treatment groups for various reasons, possibly 
resulting in non-ignorable missing data. In addition to collecting information on reasons 
for nonparticipation, we will apply two analytic approaches to examine the effects of 
missing data on study results [104]: 1) pattern mixture models and 2) selection models. 
In pattern mixture models, the four possible missing data patterns (arising from 3 time 
points with no missing data at t0 and considering a response as missing if data from 
both breasts are missing) will be used as covariates in the above-described bivariate 
linear mixed effects model. The effects of missing data on the estimate of treatment 
effect can be assessed. An overall estimate of the treatment effect may be obtained by 
averaging the estimates over the missing pattern groups. In selection model, we will 
model the probability of dropout using baseline covariates to estimate the propensity for 
dropout. The propensity scores will be used as a covariate in the bivariate linear mixed 
effects model to adjust for the effects of dropout on treatment effects. 
Secondary efficacy objectives will be evaluated by applying the methods for longitudinal 
analysis as described below. Significance of the low dose tamoxifen effects is evaluated 
in the manner described above based on testing the significance of the interaction of 
time by group indicator variables. The distribution of continuous variables will be 

 
Protocol Version: 10/26/15 
48
examined graphically and appropriate transformations made before applying analytical 
methods based on normal assumption. 
Cytomorphology and proliferative index. Samples for cytomorphology and 
proliferative index will be collected at two time points, at pre-treatment (t0) and at post-
treatment (t2). The Masood score for cytomorphology ranges from 6 to 24, which can be 
classified into 4 groups of worsening morphology: 1) nonproliferative (6-10), 2) 
hyperplasia without atypia (11-14), 3) hyperplasia with atypia (15-18), 4) malignancy (19-
24). A patient is considered to have cellular atypia if their Masood score is 15 or above. 
We will compare the effects of low dose tamoxifen versus placebo on changes in 
Masood score between pre- and post-treatment. Masood score will be treated as a 
continuous variable, transformed to normality as appropriate, and the linear mixed 
effects model applied using an indicator variable for pre- and post-treatment time points. 
We will also consider Masood score as an ordinal outcome with 4 categories and as a 
dichotomous outcome (cellular atypia or not), and apply the generalized linear mixed 
model (GLMM) for non-normal data [105]. 
Proliferative index, defined as the number of cells with unequivocal nuclear staining (Ki-
67 expression) out of a total number of ductal epithelial cells contained in RPFNA 
samples, will be analyzed using longitudinal logistic regression with two time points. If 
cell counts are better approximated by a Poisson distribution, we will apply the 
longitudinal Poisson regression method.  
Insulin growth factors. Insulin growth factor-1 (IGF1) and insulin growth factor binding 
protein-3 (IGFBP3) will be treated as continuous measures. We will apply the linear 
mixed effects model for between group comparisons of measures from the three time 
points. The unstructured mean model and linear in time model will be employed. 
 
Aim 2: To establish safety and tolerability of low-dose tamoxifen. 
Objective safety measures:  
Adverse events. The number of grade 2-4 toxicities observed will be tabulated by 
treatment arm. Differences by treatment arm will be evaluated using Fisher exact tests.  
Biomarkers. Total cholesterol, low and high density lipoprotein, triglycerides, anti-
thrombin III enzymatic assay (for clotting propensity), and serum bone-specific alkaline 
phosphatase (BSAP, for bone formation) and urine N-telopeptides (NTX, for bone 
resorption) measurements will be treated as continuous variables. Transformed to 
normality as appropriate, the linear mixed effects model with be applied, using the 
unstructured mean model using and linear in time model, to assess the effects of low 
dose tamoxifen on these measurements over time. 
Subjective safety and tolerability measures:  
Treatment adherence. Compliance will be measured by pill counts performed every 3 
months. The number of pills taken out of the total prescribed in a 3-month period will be 
modeled as a random effects binomial regression model [105]. The binomial rates from 8 
time points (month 3 to 24) will be modeled as unstructured mean model with 7 indicator 
variables as well as polynomial models over time. The random-intercept and the random 
intercept and slope models will be considered. The significance of the time indicators or 
parameters by treatment interaction will be evaluated for treatment difference in 
compliance.  

 
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Voluntary withdrawals will be examined at the end of the study by comparing the percent 
of withdrawals between the treatment groups using a chi-square test or Fisher’s exact 
test.  
Patient reported symptoms. The outcomes will be scored as a 5-point Likert-type scale 
(0 to 4) in response to questions on how much the patients are bothered by certain 
symptoms. The questionnaire will be administered every six months, for a total of five 
time points. The responses will be treated as normally distributed, as ordinal or 
dichotomized variable, and we will apply the linear mixed effects model or GLMM 
methods to compare changes between treatment groups. Because of additional time 
points available, in addition to the unstructured mean model, we will also fit piecewise 
models with join point at 6 months, considering linear and curvilinear trajectories 
between 6m and 24m time points.  
Exploratory Aim 1: To examine the modifying effects of demographic, clinical, and 
molecular characteristics on both efficacy and safety markers.  
As the primary aim of this trial is to examine the main effects of low dose tamoxifen 
treatment on MBD, we will examine the modifying effects of demographic, clinical, and 
molecular characteristics as exploratory analysis. Even if time by treatment interaction 
(main treatment effect) is not significant, examining the modifying effects of covariates 
may reveal subgroups that respond differently to low dose tamoxifen. A three-way 
interaction of time by treatment by modifying variable will be included in the longitudinal 
model containing a two-way time by treatment group interaction. Modifying effects will be 
considered statistically significant if the three-way interaction is significant. The 
covariates of interest are described below.  
A. Patient demographics. 
The following variables are known to affect breast cancer risk and MBD.  
Attained age – continuous, time-varying variable.  
Menopausal status – status at study enrollment (premenopausal or postmenopausal, as 
defined in Section 5.0) will be considered as constant over the study period. We will also 
consider trichotomized status (pre-menopausal, typical post-menopausal, and early 
post-menopausal), constant over the study period. Menopause will be defined as “early” 
if onset was before age 50.  
Prior hormone use – categorical variable (estrogen plus progestin menopausal hormone 
therapy, estrogen-only menopausal hormone therapy, hormonal contraception and/or 
fertility drugs, no prior hormonal therapy). We will also consider duration of use, as a 
continuous and dichotomous variable (5+ years, <5 years); and most recent use (within 
1 year, >1 year).  
Body mass index (BMI) – continuous, time-varying variable. It will also be grouped into 
ordinal categories: underweight (<19), normal weight (19-24), overweight (25-29), obese 
(30-39), morbidly obese (40+).  
Personal history of benign breast disease – dichotomous (yes/no), constant over the 
study period. We will also examine whether the number of prior breast biopsies, and the 
presence of atypia or LCIS on a prior biopsy further modifies the intervention effect.  
Family history of cancer – dichotomous (yes/no), constant over the study period. We will 
also examine the frequency of the following classifications: 0 = no family members with 
breast cancer; 1 = one or more first or secondary degree relative(s) with BC all 
diagnosed at 50+ years of age; 2= one first or second degree relative with BC diagnosed 

 
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before age 50 (or two, but on different sides of the family); 3 = two or more first and/or 
second degree relatives with BC diagnosed before age 50 and/or ovarian cancer on the 
same side of the family; 4 = carrier of a germline mutation in a highly penetrant BC 
susceptibility gene (BRCA1, BRCA2, PTEN, p53), or a known mutation in one of these 
genes in the patient’s family and patient has not had genetic testing.  If this results in 
sparse cells, we will consider collapsing them: A = no family history, B = some family 
history (group 2 above), and C = strong family history (groups 3 and 4 above). We will 
also examine if the number of first & second degree relatives (constant over the study 
period) is a significant effect modifier.  
B. Clinical primary cancer treatment characteristics.  
The following variables are known to influence breast cancer risk among radiation 
exposed CAYAC survivors.  
Chest radiation dose – continuous, time-invariant. We will also dichotomize dose levels 
as described in Section 12.3.  
Age at exposure to chest radiation – continuous, time-invariant. We will also categorize 
as described in Section 12.3. 
Latency from radiation therapy – continuous, time-varying. We will examine if 
intervention effects vary by time since exposure to therapeutic radiation.  
Pelvic radiation – continuous, time-invariant. Depending on the range of doses, we will 
also consider dichotomizing or categorizing the dose levels.  
Alkylator containing chemotherapy regimen – dichotomous (yes/no), time-invariant.  
C. Molecular characteristics. 
The following variables are known to influence efficacy of tamoxifen in the treatment of 
BC patients and will be analyzed at a later date.  
Endoxifen levels – continuous, time-varying. These data will be available at t1 and t2. 
CYP variants - dichotomous (yes/no), constant over the study period.  
In addition to testing their significance as effect modifiers, we will also examine their 
relationship with the outcome variables (MBD, cytomorphology, proliferative index, etc) 
to validate known relationships. Proc Mixed for normal data and Proc GLMMIX for non-
normal data in SAS 9.1 (Cary, North Carolina) will be used for analysis.  
Exploratory Aim 2: To examine the relationship between a low-dose tamoxifen regimen 
and clinical measures of efficacy and toxicity. 
For measures of clinical efficacy, we will document the occurrence of new breast cancer 
and DCIS diagnoses. 
The assessment of clinical toxicity will include the following outcomes/measures: 
1) incidence of uterine and other cancers during the study; 
2) gynecological symptoms: any and the frequency of hot flashes, night sweats, vaginal 
discharge, vaginal bleeding or spotting, missed menstrual periods for pre-menopausal 
women, genital itching/irritation, and pain with intercourse, to be obtained from symptom 
log administered 10 times (baseline, days 28, 90, 180, 270, 365, 455, 540, 630, 730) ; 
3) incident thromboembolic events (DVT, PE, TIA or stroke, and retinal vein 
thromboses); 

 
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4) incidence of cataract; 
5) liver function abnormalities: total bilirubin, SGOT/AST, SGPT/ALT, measured at 
baseline, year 1, and year 2. 
The incidence of new breast cancer, DCIS, non-breast malignancies, thromboembolic 
events, and cataract are expected to be low (≤5 events per arm) during the study period 
given the sample size, based on statistics quoted in Investigational Brochure. Thus, we 
will initially compare the frequency of events by treatment group in a 2x2 table using the 
Fisher exact test, stratified on CYP2D6 status. We will also use poisson regression in 
cohort analysis to examine treatment differences in these outcomes adjusted for 
covariates, including CYP2D6. 
The presence/absence of gynecological symptoms (for each symptom mentioned 
above) will be compared between treatment groups using 2x2 tables using the Fisher 
exact test, stratified on CYP2D6 status. Logistic regression will be used to compare the 
frequency between treatments adjusted for covariates, including CYP2D6. Longitudinal 
analysis also will be conducted to compare the trends of the presence/absence of each 
of the reported gynecological symptoms over the 2 year period using the generalized 
linear mixed-effects model for binary outcomes.  
For each liver function measure, a woman will be considered to have elevated liver 
function if the measurements at years 1 and/or 2 are above the upper limit of the 
institutional reference range and higher than her baseline pre-treatment level. These 
data will be coded such that those who are greater than 2-times the upper limit of the 
reference range will be considered to be clinically significant (grade 1 if asymptomatic 
and requiring no intervention; grade 2 if minimal/local/non-invasive intervention 
required). The frequency of women with elevated liver function measures will be 
compared between treatment groups using an exact test on 2x2 tables, stratified on 
CYP2D6. Logistic regression analysis will also be used to compare the frequency of 
elevated liver function between treatments, adjusted for covariates. Linear mixed-effects 
model for normally distributed data will also be used to compare the trends in liver 
function levels between the treatment groups. Procs MIXED and GLIMMIX will be used 
for longitudinal analysis of normally and non-normally distributed data, respectively. 
Poisson regression analysis will be conducted using Proc GENMOD, all in SAS 9.1 
(Cary, North Carolina). 
 
 
12.5 
Power and sample size:  
Aim 1: To determine the impact of low-dose tamoxifen on surrogate endpoint 
biomarkers of BC risk.  
Mammographic breast density (MBD): The primary efficacy endpoint of the trial is MBD. 
The efficacy of low dose tamoxifen will be examined by comparing the slopes in the 
longitudinal linear regression of the MBD on time since treatment initiation between the 
low dose tamoxifen group and placebo control group. For sample size calculation, we let 
Y, the response, as the mean of the square root of MBD (RTMBD) from both breasts. 
Because of randomization, we expect no difference between the expected values of Y at 
t0 between treatments but expect the linear slopes of the two groups to diverge over 
time, if the intervention is efficacious. Power/sample size calculation was conducted by 
assuming a linear change in RTMBD over time and various magnitudes of RTMBD 
difference expected at t2 between the treatment and control groups.  

 
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Longitudinal power calculation requires specification of the expected linear trend (thus 
the expected values of Y at t0, t1, t2) for the controls, the variance/covariance of the 
repeated measurements, the expected difference in Y at t2 between the treatment 
groups, and the correlations among repeated measurements. These estimates were 
obtained from 41 control women in the Puget Sound NSABP P-1 participating sites, 
[106] 16 pre-menopausal women (median age=50, range=38-50) and 25 post-
menopausal women, (median age=57, range=36-74), with data on MBD expressed as a 
percent breast density at pre-, 1-yr, and 2-yr post-treatment. Although this entire control 
sample (median age=50, range=36-74) is older than the available CPIR HL cohort 
(median age=35, range 25-50), based on changes in MBD observed with hormonal 
fluctuations during the menstrual cycle [66] and after discontinuation of exogenous 
hormone in postmenopausal women [70], it can be reasonably assumed that estrogen 
withdrawal associated with menopause has a greater impact on variation in MBD than 
age alone. Thus, we assumed that the estimates obtained from these women can be 
applied to the anticipated CAYAC patient control sample. 
The expected RTMBD at the three time points in the pre-menopausal and post-
menopausal women in the Puget Sound sample were estimated longitudinally using 
SAS Proc Mixed with a compound symmetry covariance structure. Square root 
transformation was used to normalize the distribution. A significant decline in MBD with 
time was detected. Since the covariance matrices of the pre- and post-menopausal 
women looked very similar, for simplicity, they were assumed to be equal. To determine 
the variance/covariances of the MBDs and the expected values at t0, t1, t2 in the 
anticipated CAYAC sample, 1000 random trials of size n were generated from 
multivariate normal distributions with parameters estimated from the Puget Sound 
sample to simulate the CAYAC sample expected to contain 67% pre-menopausal and 
33% post-menopausal women. Sixty-seven percent of n were drawn randomly from a 
multivariate normal distribution with expected MBD of (24.7, 23.57, 22.27) at (t0, t1, t2) 
as predicted for Puget sound pre-menopausal women, and 33% of n were drawn 
randomly from a multivariate normal distribution with expected Y of (15.49, 11.99, 13.03) 
as predicted for post-menopausal women. For each of the 1000 simulated samples, a 
longitudinal linear model was fitted assuming a compound symmetry structure for the 
covariance. The average of the 1000 estimates of the linear parameter was used to 
estimate the expected RTMBD at t0, t1, t2 for the CAYAC control sample. 
The expected RTMBDs for CAYAC controls obtained from simulations were used to 
specify the effect size, defined as the difference between expected values at t2 for the 
two groups divided by the common sd, in the longitudinal power calculations were 
conducted using the program RMASS2. [107]. We assumed a Type I error of .05, power 
of .80, 2-sided test, no attrition over time, linear trend, compound symmetry covariance 
structure, and a correlation of 0.8 between measurements (estimated from the Puget 
Sound sample).  
The size of the difference in MBD that can be detected between the low dose tamoxifen 
and placebo groups at year 2 (t2) are shown in the table below. Column 4 indicates the 
expected MBD before randomization (at t0). The expected MBDs at t2 for the control 
and treatment groups are shown in columns 5 and 6, respectively. Column 7 and 
Column 8 are the % reduction in MBD at t2 relative to t0 in the placebo control group 
and low dose tamoxifen treatment group, respectively.  The last column shows the % 
reduction in PD in the treatment group at t2 relative to controls. 
 

 
Protocol Version: 10/26/15 
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Placebo 
(PBO) 
Low Dose 
Tamoxifen 
(LDTAM) 
Power 
Exp(MBD) 
at t0 
Exp(MBD)  
at t2 
% reduction in 
MBD at t2 relative 
to t0 
% reduction 
in MBD at t2 
for LDTAM 
group relative 
to PBO 
Both 
treatment 
groups 
PBO 
LDTAM 
PBO 
 
LDTAM 
50 
50 
.8 
21.2 
18.75 
13.42 
11.6% 
36.5% 
28.4% 
 
 
.9 
“ 
“ 
12.68 
“ 
40.0% 
32.4% 
75 
75 
.8 
21.1 
18.67 
14.27 
“ 
32.5% 
23.5% 
 
 
.9 
“ 
“ 
13.70 
“ 
35.2% 
26.6% 
100 
100 
.8 
21.2 
18.72 
14.89 
“ 
29.6% 
20.5% 
 
 
.9 
“ 
“ 
14.32 
“ 
32.3% 
23.5% 
150 
150 
.8 
21.1 
18.64 
15.47 
“ 
26.8% 
17.0% 
 
 
.9 
“ 
“ 
15.00 
“ 
29.1% 
19.5% 
200 
200 
.8 
21.1 
18.67 
15.91 
“ 
24.8% 
14.8% 
 
 
.9 
“ 
“ 
15.50 
“ 
26.7% 
17.0% 
 
If we anticipate the MBD in the low dose tamoxifen group to change from 21.1 at t0 to 
14.89 at t2 compared to 18.72 at t2 in controls (in other words, a reduction of ~30% with 
low dose tamoxifen versus 11.6% with placebo over a 2 year time, or about 20% 
reduction in MBD with low dose tamoxifen relative to placebo, or an effect size of 0.25 at 
t2), a sample size of 100 per arm will be able to detect this magnitude of change with 
80% power. With 10% attrition at each annual follow-up visit, 115 women per arm (or a 
total of 230) would be required. Thus, the goal accrual for this trial will be 115 women 
per arm, for a total of 230 women at t0.   
 
Tissue and blood biomarkers: Masood score (cytomorphology) and Ki67 expression 
(proliferative index) evaluated from breast tissue samples will be obtained at t0 and t2. 
Insulin growth factors (IGF1 and IGFBP-3) will be measured at 3 time points. With a 
baseline sample size of 115 women per arm (assuming 10% yearly attrition), assuming 
Type I error=0.05, 2-sided test, and a within-person correlation of 0.5 between pairs of 
normally distributed measurements, there is 80% power to detect an effect size of 0.34 
(absolute difference between treatment and control means divided by the common 
standard deviation) at year 2. A similar effect size is detectable for Ki67 for which the 
problem was formulated as that of detecting the difference at t2 in percent of women 
with positive expression (at least one cell expressing Ki67) between the treatment 
groups. Kahn et al [86] reported a median Ki67 expression level of 1.4% in RPFNA 
samples collected from 147 women at high risk for breast cancer. Thus, we 
hypothesized that 50% (conservative estimate) to 85%.of our cohort will have Ki67 
expression in their healthy breast tissue samples collected via RPFNA at their baseline 
visit.  
 
Detectable values for the various outcomes are shown in the table below. Column 2 
shows the assumed common mean (± SD) at t0 for both treatment groups (obtained 
from the literature, adjusted for the age structure of our cohort, if possible). Column 3 
shows the minimum detectable level at t2 for the tamoxifen group in the hypothesized 

 
Protocol Version: 10/26/15 
54
direction. The mean levels at t2 in the placebo group are assumed to remain the same 
as those at t0. Column 4 shows the detectable % change at t2 relative to t0 in the 
tamoxifen group. 
 
 
 
 
 
Outcome 
Assumed levels at t0 for
Placebo and Low dose 
Tamoxifen groups 
Detectable levels at t2
(80% power)  
for Low dose Tamoxifen group 
Mean ± SD
Mean
% change (from t0)
Masood score [86] 
14 ± 1.8 
13.4 
4% ↓ 
Women positive for Ki67 
expression [86] (%) 
50% 
32% 
36% ↓ 
65% 
47% 
28% ↓ 
85% 
70% 
18% ↓ 
IGF1 [108]  (ng/ml) 
260 ± 82.5  
230 
12% ↓ 
IGFBP-3 [108] (ng/ml) 
3346 ± 1226  
3787 
13% ↑ 
 
Thus, we expect to be able to detect a 5-36% difference in the BC risk biomarkers 
proposed as secondary endpoints.  
 
Aim 2: To establish safety and tolerability of low-dose tamoxifen. 
Adverse events: We assume the rate of grade 2-4 toxicities in the placebo group to be 
low, at 0.5%. At the end of the study, with 94 women in each arm (accounting for 10% 
attrition per year), the study will have at least 80% power to detect an adverse event rate 
of 9.1% in the low dose tamoxifen group based on Fisher’s exact test and Type I 
error=0.05. 
Adherence to therapy: We will examine the difference in adherence rates of the 
treatment groups by comparing the percent of pills taken across 8 time points. With 115 
women at baseline per arm, assuming a constant adherence rate of 90% in the placebo 
group and a between-measurement correlation of 0.6 and 2-sided alternative, there will 
be 80% power to detect an adherence rate of about 77% in the low dose tamoxifen 
group.  
Voluntary withdrawals: If the proportion of patients in the placebo arm who withdraws 
voluntarily by the end of the study is 10%, then with 115 women in each arm at baseline, 
assuming type I error=0.05, the study will have 80% power to detect a significant 
difference in the voluntary withdrawal rate in the low dose tamoxifen group if the 
withdrawal rate in the intervention group is about 28%. 
Safety biomarkers: We focus here on total cholesterol (TC), anti-thrombin-III (AT-III), 
bone-specific alkaline phosphatase (BSAP), and urine N-telopeptide crosslinks (NTX). 
With the proposed sample sizes, the detectable effect size (described earlier) is 0.34. A 
relatively small % change in mean levels is detectable for TC and AT-III but larger % 
changes are needed to detect a significant treatment difference at t2 for BSAP and NTX. 
 
 
 
Outcome 
Assumed levels at t0 for
Placebo and Low dose 
Tamoxifen groups 
 
Detectable levels at t2 (80% power) for 
Low dose Tamoxifen group 
Mean ± SD 
Mean 
Relative % change (from 
t0) 

 
Protocol Version: 10/26/15 
55
TC*  (mg/dL) 
199 ± 45  
214.3 
8% ↑ 
AT-III**  (%) 
100 ± 13.3  
95.5 
5% ↓ 
BSAP [109] (µg/L) 
7.7 ± 4.5  
6.2 
20% ↓ 
NTX[110]  (nM BCE/mM) 
24.4 ± 16.1  
18.9 
23% ↓ 
 * Mean obtained from [111], sd obtained from [112] 
** per ARUP reference laboratory 
 
Thus, we expect to be able to detect a 5-23% difference in the objective safety 
biomarkers proposed as secondary endpoints.  
Exploratory Aim 1: To examine the modifying effects of demographic, clinical, and 
molecular characteristics on MBD.  
These analyses will be considered exploratory. We anticipate relatively low power as 
interaction effects are typically smaller and have relatively larger standard errors than 
main effects. Examining interactions is similar to comparing treatment effects among 
smaller sub-samples, thus power is reduced. An example is comparing the treatment 
effects of low dose tamoxifen between women who are pre- and post-menopausal at the 
time of study enrollment. Power estimates for this example were obtained by simulation. 
First, we estimated the linear mixed effects models of RTMBD for the 16 pre- and 25 
post-menopausal control women in the Puget Sound NSABP P-1 data [106]. Based on 
our power calculation which showed that a 20% reduction in MBD is detectable at year 2 
at 80% power with 100 women per arm without attrition, linear mixed effects models for 
the pre- and post-menopausal groups in the treatment arm were set up that included the 
treatment by time coefficient β=0.2338 corresponding to the detectable effect size. We 
also included another treatment by time parameter ∆ to allow differential treatment effect 
by menopausal status (+∆/2 in pre-menopausal women and –∆/2 in post-menopausal 
women, i.e. an assumption of larger treatment effect in pre- than in post-menopausal 
women). Random sets of RTMBD at (t0, t1, t2) were generated from the corresponding 
trivariate normal distributions for the four groups (67 pre- and 33 post-menopausal 
women in each treatment arm). Menopause-specific covariance matrices (assuming 
compound symmetry) estimated from the Puget Sound NSABP P-1 data were used. 
Power for detecting a significant ∆ under the alternative hypothesis of 100%, 75%, and 
50% of the main treatment effects (β=0.2338) was estimated from 1000 random samples 
of the trial data by fitting the linear mixed effects model (linear in time) that included the 
treatment by time by menopausal status term. The model also included menopause 
status main effects and menopause by time interactions. Since interactions are expected 
to be smaller than main effects, ∆=0.2338 is likely an overestimate of the interaction 
effects. Even with this overestimation, the anticipated power is only 40% (corresponds to 
treatment-related reduction in MBD of 28% relative to placebo at year 2 in the pre-
menopausal group versus 13% reduction in the post-menopausal group). For the 75% 
and 50% alternative for ∆, power is even lower (24% and 12%, respectively). Thus, 
power for examining other modifying effects are anticipated to be similarly low or lower. 
Exploratory Aim 2: To examine the relationship between a low-dose tamoxifen regimen 
and clinical measures of efficacy and toxicity. 
This aim will also be considered exploratory as the study was not powered to detect 
treatment difference in these adverse outcomes. As the events of interest have very low 
incidence rates in both treatment groups (see Table 3 of Investigational Brochure), the 
study will not have any power (<5%) to detect the relative risks seen in NSABP P-1 Trial. 
The power for detecting treatment differences in the proportion of women anticipated to 
experience hot flashes and gynecological symptoms is somewhat better, 90% for vaginal 
discharge (prevalence rates of 35% in placebo and 55% in treatment groups), but only 

 
Protocol Version: 10/26/15 
56
41% for hot flashes (prevalence rates of 68% in placebo and 80% in treated groups), 
and 5% for vaginal bleeding (22% versus 23%). 
 
13.0     REGISTRATION GUIDELINES 
 
 
 
Patients will be registered using an online secure website developed by Sharp Clinical 
Services (IWRS). To register a subject, the CRN or CRA must complete the intake form 
and eligibility worksheet online. This process is fully described in the Screening and 
Enrollment SOP.  After verification of eligibility by the coordinating center, randomization 
will be performed and treatment assigned using the Sharp Clinical Services IWRS.  This 
procedure is fully described above in Section 12.3.  
 
13.1    Procedures for On-Study and Treatment Deviations   
Protocol waivers and/or treatment deviations are generally not allowed.  However, 
should the need arise to amend the protocol in order for the protocol document to more 
accurately reflect the initial intent of the investigator(s), then the treating physician must 
contact the PI obtain approval.  If approved, the protocol PI must immediately submit an 
amendment to the protocol.  A backup for the originating PI has been designated and is 
specified in the registration worksheet. 
 
 
14.0     RECORDS TO BE KEPT AND DATA SUBMISSION SCHEDULE 
 
14.1     Confidentiality of Records 
All research records will be stored in locked file cabinets at each site. Computer files with 
coded subject information will have restricted access passwords available only to the 
local PMT. 
 
14.2     Subject Consent Form  
The original signed Consent Form will be placed in the medical record.  A copy will be 
given to the subject and a copy will be filed in the research record. 
 
14.3    Data Collection Forms and Submission Schedule 
Specific patient-administered forms to be used and the timing that they will be submitted 
are outlined in Section 9.0.  In addition, the CRA/CRN will be responsible for completing 
the forms below and entering the information into the study EDC website. 
Screening and Baseline Forms:  
The CRA/CRN will fill out the Intake Form at the time of the prescreen interview, and 
the Eligibility Worksheet at the time of the patient’s screening visit for study enrollment. 
These forms and associated documentation must be fully submitted within 14 days of 
the prescreening interview and screening visit, respectively. Should the patient be 
registered as a participant, the CRA/CRN will complete the baseline Primary Cancer 
Treatment History Form, the General History and Physical Form, and the Clinical 
Breast Exam Form. These should be submitted within 28 days of the baseline visit. 
Please refer to the Screening and Enrollment SOP for submission instructions.  
Breast Imaging and Specimen Submission Forms:  

 
Protocol Version: 10/26/15 
57
The CRA/CRN will complete the Breast Imaging Submission Form to accompany 
submission of the baseline and annual screening mammograms and, if applicable, 
annual breast MRIs. Scheduling of these imaging tests are outlined in Section 9.1. 
RPFNA-protein, RPFNA-Cytology, and Blood and Urine Specimen Submission 
Forms will be completed to accompany submission of RPFNA tissue and blood 
specimens as outlined in Sections 11.2.3 (tissue) and 11.3.1–11.3.3 (blood).  
Breast Event Form:  
The NSABP developed a data capture form to record breast events, which they used in 
the NSABP P-1 and P-2 breast cancer prevention trials. This form has been adapted for 
use in this study. The CRA/CRN will complete the Breast Event Form should a subject 
receive a diagnostic breast biopsy while on study, or during follow-up. Should such a 
biopsy reveal DCIS or invasive breast cancer, the subject will be withdrawn from the 
study treatment as outlined in Section 6.3. For other breast events, the subject may 
continue on the trial. 
Other Event Form:  
The NSABP developed a data capture form to record other cancer and other potentially 
protocol-related medical events, which they used in the NSABP P-1 and P-2 breast 
cancer prevention trials. This form has been adapted for use in this study. The 
CRA/CRN will fill out the Other Event Form should a subject receive a cancer diagnosis 
other than breast cancer or require inpatient or outpatient procedure that could be 
potentially related to protocol while on trial or during follow-up. Only for specific events 
outlined in Section 6.3 will the subject necessitate withdrawal from the study. Otherwise, 
the subject may continue on the trial. 
Treatment Interruptions:  
The CRA/CRN will fill out a Treatment Withholding Form should a subject report an 
interruption for any reason while on trial. The patient-reported reason and both hold and 
restart dates will be recorded. If the subject voluntarily chooses to withdraw from study, 
then a Study End Form will be used instead. However, if the subject chooses to 
continue on trial, then the Treatment Withholding Form will be submitted. If a patient is 
off study medication for more than 90 consecutive days, then they must be withdrawn 
from the study. In that case, the CRA/CRN will submit both Treatment Withholding and 
Study End Forms.  
Study End Form:  
The CRA/CRN will fill out the Study End Form at the time the subject goes off trial, 
even if they complete the full treatment course. This form records whether or not the 
subject completed the trial per protocol, voluntarily withdrew, or was withdrawn for 
another reason. For voluntary withdrawals, subjects will be asked to clarify if it was due 
to intolerable side effects of treatment or if it was due to another reason.  
 
Annual Follow-up Form: 
To monitor for significant medical events, including second malignancies that may occur 
after the two-year intervention, the CRA/CRN will submit an Annual Follow-Up Form 
for up to 10 years post-study completion, including for participants who withdrew prior to 
completing the study. Subjects who are lost to follow-up will be reported to the 
coordinating center using this form. New cancers and any inpatient or outpatient 
procedures potentially related to the study will be reported using the Breast Event Form 
and Other Event Form.  

 
Protocol Version: 10/26/15 
58
 
 
15.0     MINORITIES AND WOMEN STATEMENT 
Female subjects of all racial/ethnic groups are eligible for this study if they meet the 
eligibility criteria outlined in Section 5.0.   
Efforts to accrue an ethnically diverse sample are outlined in Section 4.0.  Ethnic 
distribution will likely make up a representative ratio of participants, based on the 
diversity of the communities in Los Angeles, Memphis, and Toronto. Specific minority 
groups for which each center has particular outreach include African Americans for 
SJCRH and EU, Asian Americans for PMH, and Latino Americans for COH.   
To date, there is no information that suggests that differences in drug metabolism or 
response would be expected in one group compared to another.  If differences in 
outcome appear to be associated with gender or ethnic identity, then a follow-up study 
will be designed to investigate those differences more fully. 
 
 
16.0     ETHICAL AND REGULATORY CONSIDERATIONS 
All subjects will have signed an informed consent for participation in research activities in 
accord with all institutional, National Cancer Institute (NCI) and Federal regulations, and 
will have been given a copy of the Experimental Subject's Bill of Rights. 

 
Protocol Version: 10/26/15 
59
17.0  REFERENCES 
 
1. 
Cartwright, R.A. and G. Watkins, Epidemiology of Hodgkin's disease: a review. Hematol Oncol, 
2004. 22(1): p. 11-26. 
2. 
Kaplan, H., Hodgkin's Disease. 2nd ed. 1980, Cambridge, Massachusetts: Harvard University 
Press. 
3. 
Ries LAG, H.D., Krapcho M, Mariotto A, Miller BA, Feuer EJ,  Clegg L, Eisner MP, Horner MJ, 
Howlader N, Hayat M,  Hankey BF,  Edwards BK (eds). SEER Cancer Statistics Review, 1975-
2003.   [cited; Available from: http://seer.cancer.gov/csr/1975_2003/ and 
http://dccps.nci.nih.gov/ocs/prevalence. 
4. 
Donaldson, S.S. and M.P. Link, Combined modality treatment with low-dose radiation and MOPP 
chemotherapy for children with Hodgkin's disease. J Clin Oncol, 1987. 5(5): p. 742-9. 
5. 
Engert, A., et al., Two cycles of doxorubicin, bleomycin, vinblastine, and dacarbazine plus 
extended-field radiotherapy is superior to radiotherapy alone in early favorable Hodgkin's 
lymphoma: final results of the GHSG HD7 trial. J Clin Oncol, 2007. 25(23): p. 3495-502. 
6. 
Dickerman, J.D., The late effects of childhood cancer therapy. Pediatrics, 2007. 119(3): p. 554-
68. 
7. 
Aisenberg, A.C., et al., High risk of breast carcinoma after irradiation of young women with 
Hodgkin's disease. Cancer, 1997. 79(6): p. 1203-10. 
8. 
Hancock, S.L., M.A. Tucker, and R.T. Hoppe, Breast cancer after treatment of Hodgkin's disease. 
J Natl Cancer Inst, 1993. 85(1): p. 25-31. 
9. 
Ng, A.K., et al., Second malignancy after Hodgkin disease treated with radiation therapy with or 
without chemotherapy: long-term risks and risk factors. Blood, 2002. 100(6): p. 1989-96. 
10. 
Wolden, S.L., et al., Second cancers following pediatric Hodgkin's disease. J Clin Oncol, 1998. 
16(2): p. 536-44. 
11. 
Metayer, C., et al., Second cancers among long-term survivors of Hodgkin's Disease diagnosed 
in childhood and adolescence. Journal of Clinical Oncology, 2000. 18: p. 2435-2443. 
12. 
Taylor, A.J., et al., Risk of breast cancer in female survivors of childhood Hodgkin's disease in 
Britain: a population-based study. Int J Cancer, 2007. 120(2): p. 384-91. 
13. 
Maule, M., et al., Risk of second malignant neoplasms after childhood leukemia and lymphoma: 
an international study. J Natl Cancer Inst, 2007. 99(10): p. 790-800. 
14. 
Bhatia, S., et al., High risk of subsequent neoplasms continues with extended follow-up of 
childhood Hodgkin's disease: report from the Late Effects Study Group. J Clin Oncol, 2003. 
21(23): p. 4386-94. 
15. 
Kenney, L.B., et al., Breast cancer after childhood cancer: a report from the Childhood Cancer 
Survivor Study. Ann Intern Med, 2004. 141(8): p. 590-7. 
16. 
Travis, L.B., et al., Cumulative absolute breast cancer risk for young women treated for Hodgkin 
lymphoma. J Natl Cancer Inst, 2005. 97(19): p. 1428-37. 
17. 
Easton, D.F., D. Ford, and D.T. Bishop, Breast and ovarian cancer incidence in BRCA1-mutation 
carriers. Breast Cancer Linkage Consortium. Am J Hum Genet, 1995. 56: p. 265-271. 
18. 
Easton, D.F., et al., Cancer risks in two large breast cancer families linked to BRCA2 on 
chromosome 13q12-13. Am J Hum Genet, 1997. 61(1): p. 120-8. 
19. 
Ford, D., et al., Genetic heterogeneity and penetrance analysis of the BRCA1 and BRCA2 genes 
in breast cancer families. The Breast Cancer Linkage Consortium. Am J Hum Genet, 1998. 62: p. 
676-689. 
20. 
Struewing, J.P., et al., The risk of cancer associated with specific mutations of BRCA1 and 
BRCA2 among Ashkenazi Jews. N Engl J Med, 1997. 336(20): p. 1401-1408. 
21. 
Travis, L.B., et al., Breast cancer following radiotherapy and chemotherapy among young women 
with Hodgkin disease. Jama, 2003. 290(4): p. 465-75. 
22. 
Hill, D.A., et al., Breast cancer risk following radiotherapy for Hodgkin lymphoma: modification by 
other risk factors. Blood, 2005. 106(10): p. 3358-65. 
23. 
Segaloff, A. and W.S. Maxfield, The synergism between radiation and estrogen in the production 
of mammary cancer in the rat. Cancer Res, 1971. 31(2): p. 166-8. 

 
Protocol Version: 10/26/15 
60
24. 
Stone, J.P., S. Holtzman, and C.J. Shellabarger, Synergistic interactions of various doses of 
diethylstilbestrol and x-irradiation on mammary neoplasia in female ACI rats. Cancer Res, 1980. 
40(11): p. 3966-72. 
25. 
Segaloff, A. and H.M. Pettigrew, Effect of radiation dosage on the synergism between radiation 
and estrogen in the production of mammary cancer in the rat. Cancer Res, 1978. 38(10): p. 3445-
52. 
26. 
Bartstra, R.W., et al., The influence of estrogen treatment on induction of mammary carcinoma in 
rats by single-dose gamma irradiation at different ages. Radiat Res, 1998. 150(4): p. 451-8. 
27. 
Narod, S.A. and K. Offit, Prevention and management of hereditary breast cancer. J Clin Oncol, 
2005. 23(8): p. 1656-63. 
28. 
Hartmann, L.C., et al., Efficacy of bilateral prophylactic mastectomy in BRCA1 and BRCA2 gene 
mutation carriers. J Natl Cancer Inst, 2001. 93(21): p. 1633-7. 
29. 
Schwartz, M.D., et al., Impact of BRCA1/BRCA2 Counseling and Testing on Newly Diagnosed 
Breast Cancer Patients. J Clin Oncol, 2004. 22(10): p. 1823-9. 
30. 
Rebbeck, T.R., et al., Prophylactic oophorectomy in carriers of BRCA1 or BRCA2 mutations. N 
Engl J Med, 2002. 346(21): p. 1616-22. 
31. 
Kauff, N.D., et al., Risk-reducing salpingo-oophorectomy in women with a BRCA1 or BRCA2 
mutation. N Engl J Med, 2002. 346(21): p. 1609-15. 
32. 
Bevers, T.B., et al., Breast cancer risk reduction. J Natl Compr Canc Netw, 2007. 5(8): p. 676-
701. 
33. 
Fisher, B., et al., Tamoxifen for prevention of breast cancer: Report of the national surgical 
adjuvant breast and bowel project P-1 study. Journal of the National Cancer Institute, 1998. 
90(18): p. 1371-1388. 
34. 
Gail, M.H., et al., Projecting individualized probabilities of developing breast cancer for white 
females who are being examined annually. J Natl Cancer Inst, 1989. 81(24): p. 1879-86. 
35. 
Cummings, S.R., et al., The effect of raloxifene on risk of breast cancer in postmenopausal 
women: results from the MORE randomized trial. Multiple Outcomes of Raloxifene Evaluation. 
Jama, 1999. 281(23): p. 2189-2197. 
36. 
Martino, S., et al., Continuing outcomes relevant to Evista: breast cancer incidence in 
postmenopausal osteoporotic women in a randomized trial of raloxifene. J Natl Cancer Inst, 2004. 
96(23): p. 1751-61. 
37. 
Vogel, V.G., et al., Effects of tamoxifen vs raloxifene on the risk of developing invasive breast 
cancer and other disease outcomes: the NSABP Study of Tamoxifen and Raloxifene (STAR) P-2 
trial. Jama, 2006. 295(23): p. 2727-41. 
38. 
Baum, M., 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. 
Cancer, 2003. 98(9): p. 1802-10. 
39. 
Goss, P.E., et al., A Randomized Trial of Letrozole in Postmenopausal Women after Five Years 
of Tamoxifen Therapy for Early-Stage Breast Cancer. New England Journal of Medicine 2003. 
349(19): p. 1793-1802. 
40. 
Coombes, R.C., et al., A randomized trial of exemestane after two to three years of tamoxifen 
therapy in postmenopausal women with primary breast cancer. N Engl J Med, 2004. 350(11): p. 
1081-92. 
41. 
Decensi, A., et al., Randomized dose-ranging trial of tamoxifen at low doses in hormone 
replacement therapy users. J Clin Oncol, 2007. 25(27): p. 4201-9. 
42. 
Decensi, A., et al., A randomized double-blind 2x2 trial of low-dose tamoxifen and fenretinide for 
breast cancer prevention of high-risk premenopausal women. J Clin Oncol 2008. 26((May 20 
suppl; abstr 1503)). 
43. 
Johansson, H., et al., Effect of fenretinide and low-dose tamoxifen on insulin sensitivity in 
premenopausal women at high risk for breast cancer. Cancer Res, 2008. 68(22): p. 9512-8. 
44. 
Peterson, N.C., et al., Tamoxifen resistance and Her2/neu expression in an aged, irradiated rat 
breast carcinoma model. Carcinogenesis, 2005. 26(9): p. 1542-52. 

 
Protocol Version: 10/26/15 
61
45. 
Sanna, G., et al., Breast cancer in Hodgkin's disease and non-Hodgkin's lymphoma survivors. 
Ann Oncol, 2007. 18(2): p. 288-92. 
46. 
Blankenstein, M.A., et al., Oestrogen receptors in rat mammary tissue and plasma concentrations 
of prolactin during mammary carcinogenesis induced by oestrogen and ionizing radiation. J 
Endocrinol, 1981. 88(2): p. 233-41. 
47. 
Inano, H., et al., Estradiol-17 beta as an initiation modifier for radiation-induced mammary 
tumorigenesis of rats ovariectomized before puberty. Carcinogenesis, 1995. 16(8): p. 1871-7. 
48. 
Narod, S.A., et al., Tamoxifen and risk of contralateral breast cancer in BRCA1 and BRCA2 
mutation carriers: a case-control study. Hereditary Breast Cancer Clinical Study Group. Lancet, 
2000. 356(9245): p. 1876-81. 
49. 
Yahalom, J., et al., Breast cancer in patients irradiated for Hodgkin's disease: a clinical and 
pathologic analysis of 45 events in 37 patients. J Clin Oncol, 1992. 10(11): p. 1674-81. 
50. 
Wolden, S.L., et al., Management of breast cancer after Hodgkin's disease. J Clin Oncol, 2000. 
18(4): p. 765-72. 
51. 
Fabian, C.J. and B.F. Kimler, Use of biomarkers for breast cancer risk assessment and 
prevention. J Steroid Biochem Mol Biol, 2007. 106(1-5): p. 31-9. 
52. 
Fabian, C.J., et al., Breast-tissue sampling for risk assessment and prevention. Endocr Relat 
Cancer, 2005. 12(2): p. 185-213. 
53. 
Boyd, N.F., et al., Quantitative classification of mammographic densities and breast cancer risk: 
results from the Canadian National Breast Screening Study. J Natl Cancer Inst, 1995. 87(9): p. 
670-5. 
54. 
Boyd, N.F., et al., Mammographic density and the risk and detection of breast cancer. N Engl J 
Med, 2007. 356(3): p. 227-36. 
55. 
Byrne, C., et al., Mammographic features and breast cancer risk: effects with time, age, and 
menopause status. J Natl Cancer Inst, 1995. 87(21): p. 1622-9. 
56. 
Kato, I., et al., A nested case-control study of mammographic patterns, breast volume, and breast 
cancer (New York City, NY, United States). Cancer Causes Control, 1995. 6(5): p. 431-8. 
57. 
Saftlas, A.F., et al., Mammographic densities and risk of breast cancer. Cancer, 1991. 67(11): p. 
2833-8. 
58. 
Thomas, D.B., et al., Risk of subsequent breast cancer in relation to characteristics of screening 
mammograms from women less than 50 years of age. Cancer Epidemiol Biomarkers Prev, 2002. 
11(6): p. 565-71. 
59. 
Vachon, C.M., et al., Mammographic breast density as a general marker of breast cancer risk. 
Cancer Epidemiol Biomarkers Prev, 2007. 16(1): p. 43-9. 
60. 
Warner, E., et al., The risk of breast cancer associated with mammographic parenchymal 
patterns: a meta-analysis of the published literature to examine the effect of method of 
classification. Cancer Detect Prev, 1992. 16(1): p. 67-72. 
61. 
McCormack, V.A. and I. dos Santos Silva, Breast density and parenchymal patterns as markers 
of breast cancer risk: a meta-analysis. Cancer Epidemiol Biomarkers Prev, 2006. 15(6): p. 1159-
69. 
62. 
Boyd, N.F., et al., Mammographic signs as risk factors for breast cancer. Br J Cancer, 1982. 45: 
p. 185-193. 
63. 
Byng, J.W., et al., The quantitative analysis of mammographic densities. Phys Med Biol, 1994. 
39(10): p. 1629-38. 
64. 
Saha, P.K., et al., Breast tissue density quantification via digitized mammograms. IEEE Trans 
Med Imaging, 2001. 20(8): p. 792-803. 
65. 
 Byng JW, Boyd HF, Fishell E, Jong RA, Yaffe MJ.,The quantitative analysis of mammographic 
desities. Phys Med Biol, 1994. 10: p. 1629-38. 
66. 
Highnam R, Sauber N, Destounis S, Harvey J, McDonald D, Breast density into clinical practice. 
In: ADA Maidment, PR Bakic, S, Gavenois 11th International Workshop on Breast Imaging. Berlin: 
Springer-Verlag. p. 466-473. Available: http://www.volparadensity.com/wp-
content/uploads/IWDM2012-BreastDensityInto ClinicalPractice.pdf.  Accessed 2012 Sept 28. 
67. 
Boyd, N.F., et al., Mammographic densities as a marker of human breast cancer risk and their 
use in chemoprevention. Curr Oncol Rep, 2001. 3(4): p. 314-21. 

 
Protocol Version: 10/26/15 
62
68. 
Fabian, C.J. and B.F. Kimler, Mammographic density: use in risk assessment and as a biomarker 
in prevention trials. J Nutr, 2006. 136(10): p. 2705S-8S. 
69. 
Greendale, G.A., et al., Effects of estrogen and estrogen-progestin on mammographic 
parenchymal density. Annals of Internal Medicine, 1999. 130: p. 262-269. 
70. 
Rutter, C.M., et al., Changes in Breast Density Associated With Initiation, Discontinuation, and 
Continuing Use of Hormone Replacement Therapy. JAMA, 2001. 285(2): p. 171-176. 
71. 
Greendale, G.A., et al., Postmenopausal hormone therapy and change in mammographic 
density. J Natl Cancer Inst, 2003. 95(1): p. 30-7. 
72. 
Anderson, G.L., et al., Effects of conjugated equine estrogen in postmenopausal women with 
hysterectomy: the Women's Health Initiative randomized controlled trial. Jama, 2004. 291(14): p. 
1701-1712. 
73. 
Rossouw, J.E., et al., Risks and benefits of estrogen plus progestin in healthy postmenopausal 
women: principal results From the Women's Health Initiative randomized controlled trial. JAMA, 
2002. 288(3): p. 321-33. 
74. 
Chlebowski, R.T., et al., Influence of estrogen plus progestin on breast cancer and 
mammography in healthy postmenopausal women: the Women's Health Initiative Randomized 
Trial. Jama, 2003. 289(24): p. 3243-53. 
75. 
Brisson, J., et al., Tamoxifen and Mammographic Breast Densities. Cancer Epidemiology, 
Biomarkers & Prevention, 2000. 9: p. 911-915. 
76. 
Chow, C.K., et al., Effect of Tamoxifen on Mammographic Density. Cancer Epidemiol Biomarkers 
Prev, 2000. 9(9): p. 917-921. 
77. 
Cuzick, J., et al., Tamoxifen and breast density in women at increased risk of breast cancer. J 
Natl Cancer Inst, 2004. 96(8): p. 621-8. 
78. 
Freedman, M., et al., Digitized mammography: a clinical trial of postmenopausal women randomly 
assigned to receive raloxifene, estrogen, or placebo. J Natl Cancer Inst Monogr, 2001. 93(1): p. 
51-56. 
79. 
Palomares, M.R., et al., Tamoxifen modifies mammographic density, a noninvasive marker of 
breast cancer risk. Breast Cancer Research and Treatment, 2001. 70(Sup 1): p. 133. 
80. 
Cuzick, J., et al., Change in breast density as a biomarker of breast cancer risk reduction: results 
from IBIS-1. San Antonio Breast Cancer Symposium, 2008. 
81. 
Fearon, E.R. and B. Vogelstein, A genetic model for colorectal tumorigenesis. Cell, 1990. 61: p. 
759-767. 
82. 
Wang, J., et al., Lower-category benign breast disease and the risk of invasive breast cancer. J 
Natl Cancer Inst, 2004. 96(8): p. 616-20. 
83. 
Shoker, B.S., et al., Estrogen receptor-positive proliferating cells in the normal and precancerous 
breast. Am J Pathol, 1999. 155(6): p. 1811-5. 
84. 
Shaaban, A.M., et al., Breast cancer risk in usual ductal hyperplasia is defined by estrogen 
receptor-alpha and Ki-67 expression. Am J Pathol, 2002. 160(2): p. 597-604. 
85. 
Masood, S., et al., Cytologic differentiation between proliferative and nonproliferative breast 
disease in mammographically guided fine-needle aspirates. Diagn Cytopathol, 1991. 7(6): p. 581-
90. 
86. 
Khan, Q.J., et al., Ki-67 expression in benign breast ductal cells obtained by random periareolar 
fine needle aspiration. Cancer Epidemiol Biomarkers Prev, 2005. 14(4): p. 786-9. 
87. 
Mori, H., et al., Control of cell proliferation in cancer prevention. Mutat Res, 1999. 428(1-2): p. 
291-8. 
88. 
Chang, J., et al., Prediction of clinical outcome from primary tamoxifen by expression of biologic 
markers in breast cancer patients. Clin Cancer Res, 2000. 6(2): p. 616-21. 
89. 
Anderson, T.J., et al., Early changes in tumour Ki67 expression differentiatie for pathological 
response in breast cancers treated neo-adjuvantly with letrozole. Breast Cancer Treatment and 
Research, 2004: p. 1027. 
90. 
Jackson, J., et al., Short and long term effects of letrozole on tumour histo and immuno-pathology 
in breast cancer patients given neoadjuvant treatment. Breast Cancer Treatment and Research, 
2004: p. 430. 

 
Protocol Version: 10/26/15 
63
91. 
Assersohn, L., et al., Studies of the potential utility of Ki67 as a predictive molecular marker of 
clinical response in primary breast cancer. Breast Cancer Res Treat, 2003. 82(2): p. 113-23. 
92. 
Fabian, C.J., et al. Reduction in breast epithelial cell proliferation after six months of letrozole in 
high risk women on hormone replacement therapy withrandom periareolar fine needle aspiration 
evidence of atypia. in 27th Annual San Antonio Breast Cancer Symposium. 2004: Breast Cancer 
Research and Treatment. 
93. 
Tan-Chiu, E., et al., Effects of tamoxifen on benign breast disease in women at high risk for 
breast cancer. J Natl Cancer Inst, 2003. 95(4): p. 302-7. 
94. 
Fabian, C., et al., Short-Term Breast Cancer Prediction by Random Periareolar Fine-Needle 
Aspiration Cytology and the Gail Risk Model. Journal of the National Cancer Institute, 2000. 
93(15): p. 1217-1227. 
95. 
Fabian, C.J., et al., Breast cytology and biomarkers obtained by random fine needle aspiration: 
use in risk assessment and early chemoprevention trials. J Cell Biochem Suppl, 1997. 28-29: p. 
101-10. 
96. 
Fabian, C.J. and B.F. Kimler, Selective estrogen-receptor modulators for primary prevention of 
breast cancer. J Clin Oncol, 2005. 23(8): p. 1644-55. 
97. 
Decensi, A., et al., A randomized trial of low-dose tamoxifen on breast cancer proliferation and 
blood estrogenic biomarkers. J Natl Cancer Inst, 2003. 95(11): p. 779-90. 
98. 
Kisanga, E.R., et al., Tamoxifen and metabolite concentrations in serum and breast cancer tissue 
during three dose regimens in a randomized preoperative trial. Clin Cancer Res, 2004. 10(7): p. 
2336-43. 
99. 
Masood, S., et al., Prospective evaluation of radiologically directed fine-needle aspiration biopsy 
of nonpalpable breast lesions. Cancer, 1990. 66(7): p. 1480-7. 
100. 
Fabian, C.J., et al., Identification of a chemoprevention cohort from a population of women at high 
risk for breast cancer. J Cell Biochem Suppl, 1996. 25: p. 112-22. 
101. 
Land, S.R., et al., Patient-reported symptoms and quality of life during treatment with tamoxifen or 
raloxifene for breast cancer prevention: the NSABP Study of Tamoxifen and Raloxifene (STAR) 
P-2 trial. Jama, 2006. 295(23): p. 2742-51. 
102. 
Alfano, C.M., et al., Psychometric properties of a tool for measuring hormone-related symptoms 
in breast cancer survivors. Psychooncology, 2006. 15(11): p. 985-1000. 
103. 
Laird, N.M. and J.H. Ware, Random-effects models for longitudinal data. Biometrics, 1982. 38(4): 
p. 963-74. 
104. 
Little, R.J.A., Modeling the drop-out mechanism in repeated-measures studies. Journal of the 
American Statistical Association, 1995. 90: p. 1112-1121. 
105. 
Breslow, N.E. and D.G. Clayton, Approximate inference in generalized linear mixed models. 
Journal of the American Statistical Association, 1993. 88: p. 9-25. 
106. 
Palomares, M.R., et al., Mammographic density correlation with Gail model breast cancer risk 
estimates and component risk factors. Cancer Epidemiol Biomarkers Prev, 2006. 15(7): p. 1324-
30. 
107. 
Hedeker, D., R.D. Gibbons, and C. Waternaux, Sample size estimation for longitudinal designs 
with attrition: comparing time-related contrasts between two groups. Journal of Educational and 
Behavioral Statistics, 1999. 24: p. 70-93. 
108. 
Rinaldi, S., et al., IGF-I, IGFBP-3 and breast cancer risk in women: The European Prospective 
Investigation into Cancer and Nutrition (EPIC). Endocr Relat Cancer, 2006. 13(2): p. 593-605. 
109. 
Gonnelli, S., et al., Bone alkaline phosphatase measured with a new immunoradiometric assay in 
patients with metabolic bone diseases. Eur J Clin Invest, 1996. 26(5): p. 391-6. 
110. 
de Papp, A.E., et al., A cross-sectional study of bone turnover markers in healthy premenopausal 
women. Bone, 2007. 40(5): p. 1222-30. 
111. 
Schober, S.E., et al., High Serum Total Cholesterol - An Indicator for Monitoring Cholesterol 
Lowering Efforts:  U.S. Adults, 2005-2006. 2007, DHHS, NCHS. p. 1-8. 
112. 
Fridinger, F.W., A.W. Jackson, and J. Andresen, A comparison of results of a national cholesterol 
and blood pressure screening with the NHANES II Study: implications for further emphasis on 
reducing cardiovascular risk among Americans. J Community Health, 1992. 17(4): p. 247-57.