Biomarker Expression and Risk of Subsequent Tumors After Initial Ductal Carcinoma In Situ Diagnosis

University of California, San Francisco, CA 94121, USA. [corrected]
Journal of the National Cancer Institute (Impact Factor: 12.58). 05/2010; 102(9):627-37. DOI: 10.1093/jnci/djq101
Source: PubMed
ABSTRACT
Studies have failed to identify characteristics of women who have been diagnosed with ductal carcinoma in situ (DCIS) and have a high or low risk of subsequent invasive cancer.
We conducted a nested case-control study in a population-based cohort of 1162 women who were diagnosed with DCIS and treated by lumpectomy alone from 1983 to 1994. We collected clinical characteristics and information on subsequent tumors, defined as invasive breast cancer or DCIS diagnosed in the ipsilateral breast containing the initial DCIS lesion or at a regional or distant site greater than 6 months after initial treatment of DCIS (N = 324). We also conducted standardized pathology reviews and immunohistochemical staining for the estrogen receptor (ER), progesterone receptor, Ki67 antigen, p53, p16, epidermal growth factor receptor-2 (ERBB2, HER2/neu oncoprotein), and cyclooxygenase-2 (COX-2) on the initial paraffin-embedded DCIS tissue. Competing risk models were used to determine factors associated with risk of subsequent invasive cancer vs DCIS, and cumulative incidence survival functions were used to estimate 8-year risk.
Factors associated with subsequent invasive cancer differed from those associated with subsequent DCIS. Eight-year risk of subsequent invasive cancer was statistically significantly (P = .018) higher for women with initial DCIS lesions that were detected by palpation or that were p16, COX-2, and Ki67 triple positive (p16(+)COX-2(+)Ki67(+)) (19.6%, 95% confidence interval [CI] = 18.0% to 21.3%) than for women with initial lesions that were detected by mammography and were p16, COX-2, and Ki67 triple negative (p16(-)COX-2(-)Ki67(-)) (4.1%, 95% CI = 3.4% to 5.0%). In a multivariable model, DCIS lesions that were p16(+)COX-2(+)Ki67(+) or those detected by palpation were statistically significantly associated with subsequent invasive cancer, but nuclear grade was not. Eight-year risk of subsequent DCIS was highest for women with DCIS lesions that had disease-free margins of 1 mm or greater combined with either ER(-)ERBB2(+)Ki67(+) or p16(+)COX-2(-)Ki67(+) status (23.6%, 95% CI = 18.1% to 34.0%).
Biomarkers can identify which women who were initially diagnosed with DCIS are at high or low risk of subsequent invasive cancer, whereas histopathology information cannot.

Full-text

Available from: Karla Kerlikowske
jnci.oxfordjournals.org JNCI
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Articles 627
DOI: 10.1093/jnci/djq101 Published by Oxford University Press 2010.
Advance Access publication on April 28, 2010.
Ductal carcinoma in situ (DCIS) has become a relatively common
diagnosis (1,2), yet the clinical and biological significance of DCIS
lesions is not fully understood. It appears that 5%–10% of women
diagnosed with DCIS who are treated by lumpectomy alone de-
velop a subsequent invasive cancer within 5 years, and a similar
proportion develops a subsequent DCIS lesion (3–7). Adjuvant
radiation and tamoxifen have been shown to decrease the rate of
subsequent tumors (3,8,9), but not to influence breast cancer mor-
tality (4–6,10–12).
Clinical trials and population-based studies have failed to con-
sistently identify which women will be at high vs low risk of sub-
sequent invasive cancer among those diagnosed with DCIS
(10,13), thereby creating a dilemma for physicians in choosing the
intensity of their treatment (14). Identification of biomarkers that
can accurately predict subsequent invasive cancer and/or DCIS
could aid in stratifying an individual’s risk for subsequent tumors.
A few studies have examined biomarkers including the estrogen
receptor (ER), progesterone receptor (PR), human epidermal
growth factor receptor-2 oncoprotein (HER2/neu, also known as
ERBB2), human epidermal growth factor receptor-4 oncoprotein
(HER4/neu), Ki67, and cyclooxygenase-2 (COX-2) as predictors
of subsequent tumors in women diagnosed with DCIS, but the
results have been inconsistent (15–20). These studies were based
primarily on follow-up of nonpopulation-based case series of
women in whom DCIS had been managed with a variety of treat-
ment modalities, making it difficult to know whether the results
ARTICLE
Biomarker Expression and Risk of Subsequent Tumors After
Initial Ductal Carcinoma In Situ Diagnosis
Karla Kerlikowske, Annette M. Molinaro, Mona L. Gauthier, Hal K. Berman, Fred Waldman, James Bennington, Henry Sanchez,
Cynthia Jimenez, Kim Stewart, Karen Chew, Britt-Marie Ljung, Thea D. Tlsty
Manuscript received June 22, 2009; revised July 14, 2009; accepted March 5, 2010.
Correspondence to: Karla Kerlikowske, MD, San Francisco Veterans Affairs Medical Center, General Internal Medicine Section, 111A1, 4150 Clement St,
University of California, San Francisco, San Francisco, CA 94121 (e-mail: karla.kerlikowkse@ucsf.edu) or Thea D. Tlsty, PhD, Box 0511, Core Campus, HSW
513, University of California, San Francisco, San Francisco, CA 94143 (e-mail: thea.tlsty@ucsf.edu).
Background Studies have failed to identify characteristics of women who have been diagnosed with ductal carcinoma in situ
(DCIS) and have a high or low risk of subsequent invasive cancer.
Methods We conducted a nested case–control study in a population-based cohort of 1162 women who were diagnosed
with DCIS and treated by lumpectomy alone from 1983 to 1994. We collected clinical characteristics and infor-
mation on subsequent tumors, defined as invasive breast cancer or DCIS diagnosed in the ipsilateral breast
containing the initial DCIS lesion or at a regional or distant site greater than 6 months after initial treatment of
DCIS (N = 324). We also conducted standardized pathology reviews and immunohistochemical staining for the
estrogen receptor (ER), progesterone receptor, Ki67 antigen, p53, p16, epidermal growth factor receptor-2
(ERBB2, HER2/neu oncoprotein), and cyclooxygenase-2 (COX-2) on the initial paraffin-embedded DCIS tissue.
Competing risk models were used to determine factors associated with risk of subsequent invasive cancer vs
DCIS, and cumulative incidence survival functions were used to estimate 8-year risk.
Results Factors associated with subsequent invasive cancer differed from those associated with subsequent DCIS.
Eight-year risk of subsequent invasive cancer was statistically significantly (P = .018) higher for women with
initial DCIS lesions that were detected by palpation or that were p16, COX-2, and Ki67 triple positive (p16
1
COX-
2
1
Ki67
1
) (19.6%, 95% confidence interval [CI] = 18.0% to 21.3%) than for women with initial lesions that were
detected by mammography and were p16, COX-2, and Ki67 triple negative (p16
2
COX-2
2
Ki67
2
) (4.1%, 95% CI =
3.4% to 5.0%). In a multivariable model, DCIS lesions that were p16
1
COX-2
1
Ki67
1
or those detected by palpation
were statistically significantly associated with subsequent invasive cancer, but nuclear grade was not. Eight-
year risk of subsequent DCIS was highest for women with DCIS lesions that had disease-free margins of 1 mm
or greater combined with either ER
2
ERBB2
1
Ki67
1
or p16
1
COX-2
2
Ki67
1
status (23.6%, 95% CI = 18.1% to 34.0%).
Conclusion Biomarkers can identify which women who were initially diagnosed with DCIS are at high or low risk of subse-
quent invasive cancer, whereas histopathology information cannot.
J Natl Cancer Inst 2010;102:627–637
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were a function of biomarkers or treatment or both. In addition,
most of the studies were small, conducted at a single institution,
had short length of follow-up, tested only individual markers,
and did not stratify by type of subsequent tumor. These study
design restrictions limit the ability of published results to be
generalized.
The primary purpose of this study was to identify clinical, his-
topathologic, and molecular characteristics of initial DCIS lesions
that are associated with subsequent invasive cancer or DCIS. We
studied a large population-based cohort of women with DCIS who
were treated by lumpectomy alone to determine risk of subsequent
disease as a function of these factors.
Subjects and Methods
Subjects
The study sample and methods have been previously described (5).
In brief, we used data from the Surveillance, Epidemiology, and
End Results (SEER) program of Northern California to identify
women who were aged 40 years or older when diagnosed with
DCIS and who were treated by lumpectomy alone in one of the
nine San Francisco Bay Area counties from January 1, 1983, to
December 31, 1994. From an initial pool of 1568 women, we ex-
cluded 142 women who had DCIS treated by mastectomy or by
lumpectomy plus radiation within 6 months of the initial diagno-
sis, 19 women who had a prior diagnosis of breast cancer,
18 women who died within 6 months of the initial diagnosis,
37 women whose initial DCIS lesion was found to have invasive
cancer on standardized pathology review, and 20 women whose
DCIS diagnosis could not be confirmed. Of the 1332 eligible par-
ticipants, 29 women could not be located; 18 women did not speak
fluent English, Cantonese, Spanish, or Russian (the languages we
used to conduct the telephone interviews); 118 women refused to
participate; and five women had a doctor’s request not to be con-
tacted. The study cohort consisted of 1162 women with an overall
participation rate of 87%. This study was reviewed and approved
by the University of California, San Francisco Committee on
Human Research. Study participants provided verbal and/or
written informed consent.
Telephone Interviews and Vital Status
We obtained demographic information and a breast health history
from each woman during a telephone interview on average 7.5
years after initial diagnosis, as previously described (5). In brief,
the interview included questions about breast procedures a woman
had undergone, family history of breast cancer, detection method
at diagnosis, and menopausal status. To obtain information for
206 women who were either deceased or not able to participate in
an interview because of illness, we interviewed a proxy and/or
conducted medical record review. We obtained data regarding
vital status and underlying cause of death including breast cancer
as of December 31, 2005, from the California Department of Vital
Statistics and/or death certificates.
Standardized Pathology Review for Nested
Case–Control Study
Paraffin-embedded tissue samples and/or hematoxylin- and eosin-
stained slides of initial DCIS tissue from women who had subse-
quent tumors (case subjects) and women with DCIS who did not
have subsequent tumors (control subjects) were retrieved from
pathology laboratories. Control subjects were randomly selected
and frequency matched to case subjects by year of diagnosis before
retrieval of their DCIS tissue. We could not obtain paraffin-
embedded tissue blocks from some hospitals that had discarded
the tissues, had insufficient staff to collect the tissues, and/or
refused to provide tissue for research (80 case subjects and 93
control subjects). Subsequent tumors were defined as DCIS or
invasive breast cancer that was diagnosed in the ipsilateral breast
(that had contained the initial DCIS lesion) or at a regional or
distant site (bone, brain, liver, lung, and skin) more than 6 months
after the initial diagnosis and treatment of DCIS. Women who
had both DCIS and invasive cancer in subsequent tissue samples
were categorized as having a subsequent invasive cancer. To clas-
sify a woman as having had a subsequent tumor event as defined
above, we investigated the nature of all breast procedures reported
by women during the telephone interview by obtaining and
reviewing pathology reports for breast biopsies performed after
the initial diagnosis and linking to the Northern California SEER
program in 2002 and 2008. Pathology reports were available on
94% of breast biopsies performed after the initial diagnosis.
Women who developed only contralateral breast cancer during
the study period were included in the study as control subjects.
As previously described (5), study pathologists blinded to the
clinical outcome reviewed the slides stained with hematoxylin and
CONTEXT AND CAVEATS
Prior knowledge
Current biomarkers have been inadequate to distinguish which
women with a diagnosis of ductal carcinoma in situ (DCIS) have a
high risk of subsequent invasive breast cancer.
Study design
Clinical data and breast tissue specimens were collected from 1162
surgically treated DCIS patients from 1983 to 1994, and DCIS tissue
was subjected to immunohistochemical staining. Women whose
DCIS tissue expressed various combinations of biomarkers were
compared for 5- and 8-year risks of invasive cancer or subsequent
DCIS.
Contribution
Eight-year risk of invasive cancer was highest among women
whose DCIS lesions were detectable by palpation or were p16,
cyclooxygenase-2, and Ki67 positive. Eight-year risk of further DCIS
was highest among women with different specific biomarker
combinations.
Implications
These biomarker combinations may be useful to women and their
physicians as prognostic indicators.
Limitations
All data were collected retrospectively, subject to tissue avail-
ability. Patients had been treated by lumpectomy alone, so bio-
marker combinations could not be used to predict responsiveness
to adjuvant therapies.
From the Editors
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eosin (N = 502) of the original DCIS lesions from 114 women who
had a subsequent invasive cancer event, 109 women who had a
subsequent DCIS event, and 279 control subjects who did not
have a subsequent tumor event to verify initial diagnoses of DCIS;
to verify diagnoses of subsequent disease; and to determine nu-
clear grade, type, and quantity of necrosis, tumor size, and margin
width of the initial DCIS diagnosis. Our pathologists established
at least 80% agreement on identification of histopathologic char-
acteristics in a training set of women with DCIS before reviewing
study case and control subjects. Disagreements were resolved by
consensus.
Measurement of Biomarkers for Nested
Case–Control Study
We used immunohistochemical staining to identify DCIS phe-
notypes using slides from formalin-fixed paraffin-embedded
tumor blocks (N = 329) from 72 women who had a subsequent
invasive cancer event, 71 women who had a subsequent DCIS
event, and 186 control subjects who did not have a subsequent
tumor event. We scored the index lesion for the presence of the
following proteins using the indicated mouse monoclonal anti-
bodies: for ER using a 1:400 dilution of antibody 1D5 (DAKO,
Carpinteria, CA), for PR using a 1:25 dilution of antibody 1A6
(Novocastra, Bannockburn, IL), for Ki67 antigen (MKI67 [FHA
domain] interacting nucleolar phosphoprotein) using a 1:100
dilution of antibody MIB-1 (DAKO), for p53 (TP53) using a
1:200 dilution of antibody PAb 1801 (Neomarkers, Fremont,
CA), for human epidermal growth factor receptor-2 (ERBB2)
using a 1:200 dilution of antibody TAB250 (Invitrogen, Grand
Island, NY), for COX-2 using a 1:200 dilution of antibody
M3617 (DAKO), and for p16 (cyclin-dependent kinase inhibitor
2A) using a 1:200 dilution of antibody MS218 (Neomarkers)
(21,22). Staining with primary antibodies was followed by stain-
ing with biotinylated labeled secondary antibodies and detection
with an avidin–biotin–horseradish peroxidase system. Specimens
were counterstained with hematoxylin. Positive and negative
control tissues were used for assessment of each marker as fol-
lows: ER, breast tumor case and cell line MCF-7; PR, breast
tumor case and cell line T47D; Ki67, breast tumor case; p53,
colon tumor case and cell line T47D; ERBB2, breast tumor case
and cell line SKBR3; COX-2, a DCIS case; and p16, normal
breast tissue and colon case.
One investigator ( J . Bennington) scored ER, PR, ERBB2, and
p53 stains, and two investigators (M. L. Gauthier and H. K.
Berman) scored p16, COX-2, and Ki67 stains; all were blinded to
clinical outcomes. For p53, ERBB2, ER, and PR, the percentage
of tumor cells that showed staining of any intensity was estimated
and recorded. The marker p53 was considered to be overex-
pressed, and ER and PR were considered to be present when 10%
or more tumor cells showed staining. Similarly, ERBB2 was con-
sidered to be overexpressed when 10% or more tumor cells
showed moderate or strong membrane staining (+2 or higher);
these were criteria previously used for scoring DCIS lesions for
ERBB2 (23).
Using a condensed Allred score (24), COX-2 staining was eval-
uated on a scale of 0, 1, 2, or 3, with each value corresponding
to a combination of Allred classes (0 = Allred class 0; 1 = Allred
classes 2, 3, and 4; 2 = 5 and 6; 3 = 7 and 8; see Supplementary
Material, available online). Scoring of p16 was evaluated on a scale
of 0, 1, 2, or 3 based on the percentage of positively staining tumor
cells, irrespective of staining intensity (0 = no staining, 1 = fewer than
25% of cells stained, 2 = 25%–75%, 3 = more than 75% of cells
stained) (22). Tissues with a score of at least 2 were considered to
overexpress COX-2 or p16. For Ki67 scoring, a minimum of 1000
tumor cells were counted from at least three high-powered (×40)
fields in areas that showed the highest labeling. The labeling index
was expressed as a percentage and was calculated as the number of
positive cells divided by the number of positive plus negative cells.
Tissues were considered to have high Ki67 expression if more than
10% of tumor cells were stained, which was more than the median
value for all tumors evaluated. In a random sample of 45 speci-
mens, a comparison of two independent scorers of select immuno-
histochemical assays yielded a k statistic of 0.93 and concordance
of 98% for p16, a k statistic of 0.73 and concordance of 87% for
COX-2, and a k statistic of 0.82 and concordance of 91% for Ki67
(see Supplementary Material, available online, for representative
staining).
Statistical Analysis
We used Cox proportional hazards models to determine univar-
iate and multivariable hazard ratios (HR) for various clinical and
histopathologic characteristics and biomarkers among women in
the cohort who had a subsequent tumor compared with women
who did not. We examined combinations of biomarkers that were
found as individual markers in univariate analyses to be statistically
significantly associated with invasive cancer and/or DCIS or were
previously shown to have a biological basis for association with
subsequent tumors after a DCIS diagnosis (22) or were previously
reported to be associated with breast cancer survival (25). For in-
clusion in the multivariable models, we considered individual and
combinations of factors that were statistically significantly associ-
ated with invasive cancer and/or DCIS in univariate analyses. For
multivariable models, margin width was considered as an ordinal
variable (ordered as 10 mm, 2 to <10 mm, 1 to 1.9 mm, uncertain,
and positive). The validity of the proportional hazards assumption
was verified by log-cumulative hazard plots and, where appro-
priate, inclusion of a time-dependent variable. Subsequent inva-
sive cancer, DCIS, and death from causes other than breast cancer
were competing events. To calculate the appropriate hazard ratio,
we used the competing risk package cmprsk in R (http://cran
.rproject.org/doc/packages/cmprsk.pdf ) to estimate coefficients
in the “proportional subdistribution hazards” regression model
described by Fine and Gray (26). This model can be used to
directly assess the effect of covariates on the subdistribution of a
particular type of outcome, in this case invasive cancer or DCIS,
in a competing risk setting. In a sensitivity analysis, we excluded
women who developed contralateral breast cancer and results were
very similar to the results we present from the final models.
To estimate the risk of subsequent tumor events (invasive can-
cer or DCIS), we generated a standard Kaplan–Meier survival
curve. To estimate the 5- and 8-year probability of subsequent
tumor events for the population-based cohort by histopathologic
characteristics and biomarkers that were collected only for case
and control subjects, the results of the case–control study were
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converted to survival curves. To do so, we imputed histopatho-
logic characteristics and biomarker measurements for those
women in the cohort who were not included in the nested case–
control study. The imputed values were based on the observed
prevalence of the individual histopathologic and biomarkers in the
nested study stratified by case and control subject status as well as
by the type of subsequent tumor event as previously described (5).
To estimate the risk of subsequent invasive cancer with a DCIS
event and death from causes other than breast cancer (N = 125) as
competing risks, and to estimate risk of subsequent DCIS with an
invasive cancer event and death from causes other than breast
cancer as competing risks, we used the code from Pepe and Mori
(27) to estimate the marginal distribution, that is, the cumulative
incidence function. This process was repeated 2500 times, each
time generating a new imputed value for each woman for whom
we had missing data for a marker of interest. For each time point
t, the 2500 Kaplan–Meier or cumulative incidence function sur-
vival estimates were averaged and the 95% confidence interval
(CI) was reported as the 0.025 and 0.975 quantiles of those survival
estimates.
Four risk groups (ie, lowest, low, intermediate, and high risk)
were defined separately for subsequent invasive cancer and DCIS
based on statistically significant univariate and multivariable asso-
ciations as well as level of risk associated with clinical and histo-
pathologic characteristics and molecular markers and subsequent
invasive cancer or DCIS. Groups were defined by combining clin-
ical and histopathologic characteristics and molecular markers that
have similar strength associations and level of risk for subsequent
tumor events.
All statistical tests were two-sided. P values less than .05 were
considered statistically significant.
Results
From January 1, 1983, to May 1, 2008, 324 of the 1162 women in
the study cohort (27.9% overall or 3% per year) developed a
subsequent breast tumor (median follow-up = 98.0 months or 8.2
years [range = 6.3 to 299.1 months or 0.5 to 25 years]). Of the
1162 women, 154 (13.3%) had subsequent local DCIS lesions,
170 had subsequent invasive cancer (of these, 120 [10.3%] had
local disease, 33 [2.8%] regional disease, eight [0.7%] distant
disease, and nine [0.7%] disease of unknown location), and 125
(10.8%) died of a cause other than breast cancer. Among the
women who had subsequent invasive cancer, 34 (2.9%) died of
breast cancer. The 8-year risk of subsequent invasive cancer was
similar to the 8-year risk of subsequent DCIS (11.1% vs 11.6%,
respectively).
Univariate Results of Factors Associated With
Subsequent Invasive Cancer vs DCIS
The risk of subsequent invasive cancer was increased among
women whose initial DCIS was detected by palpation compared
with that for women whose DCIS was detected by mammography
(HR = 2.0, 95% CI = 1.3 to 2.9). The proportional incidence of
DCIS by mode of detection did not vary by year of diagnosis (data
not shown). We observed that risk of subsequent DCIS varied by
age: There was increased risk for women aged 40–49 years
compared with women aged 70 years and older (HR = 2.2, 95%
CI = 1.4 to 3.4). Race and/or ethnicity, family history of breast
cancer, and menopausal status were not associated with incidence
of subsequent tumors (Table 1), and neither was oral contraceptive
or postmenopausal hormone therapy use or body mass index (data
not shown).
Whereas histopathologic characteristics were not associated
with subsequent invasive cancer (Table 2), several such character-
istics were associated with an increased risk of subsequent DCIS:
initial DCIS lesions that were larger than 10 mm, had positive or
uncertain margins, were of high nuclear grade, or had extensive
necrosis (Table 2).
Whereas DCIS lesions with individual expression of the bio-
markers ER, PR, p53, ERBB2, and COX-2 were not statistically
significantly associated with subsequent invasive cancer, p16 and
selected combinations of markers did provide stratification of risk
(Table 3). Women whose initial DCIS lesions were p16 positive
(p16
1
) or p16 and Ki67 positive (p16
1
Ki67
1
) or p16, COX-2, and
Ki67 positive (p16
1
COX-2
1
Ki67
1
) had an increased risk of subse-
quent invasive cancer compared with women whose DCIS lesions
did not express these combinations of markers (Table 3). Of note,
Ki67 in combination with ER, PR, p53, or ERBB2 was not associ-
ated with subsequent invasive cancer nor was p16 in combination
with ER, PR, p53, or ERBB2 (data not shown).
Markers associated with subsequent DCIS differed from those
associated with subsequent invasive cancer. Women whose initial
DCIS lesions were ER negative (ER
2
), ERBB2 positive (ERBB2
1
),
or Ki67 positive (Ki67
1
) among individual markers, or were
ER
2
ERBB2
1
or ER
2
Ki67
1
among marker combinations, had an
increased risk of subsequent DCIS compared with women who
had lesions that did not express these individual markers or com-
binations of markers. Subsequent DCIS also was associated with
initial DCIS lesions that were p16
1
Ki67
1
or p16
1
COX-2
2
Ki67
1
.
Distributions by tumor size, margin status, and nuclear grade
according to case–control status were similar for women for whom
we could obtain tumor blocks and those we could not (data not
shown).
Multivariable Results of Factors Associated With
Subsequent Invasive Cancer vs DCIS and Risk of
Subsequent Tumors by These Factors
In a multivariable competing risk model, we found both DCIS
lesions that were detected by palpation and those that were
p16
1
COX-2
1
Ki67
1
were statistically significantly associated
with subsequent invasive cancer, whereas nuclear grade was not
(Table 4). When we examined the subgroup of women whose
initial DCIS was detected by mammography, the independent
association of p16
1
COX-2
1
Ki67
1
lesions with subsequent inva-
sive cancer remained (HR = 2.3, 95% CI = 1.0 to 5.3). Among
DCIS lesions associated with a subsequent invasive cancer, 25%
were detected by palpation and 23% were p16
1
COX-2
1
Ki67
1
;
only two case subjects had both these traits. The 5- and 8-year
risks of subsequent invasive cancer were high for women whose
initial DCIS lesions were detected by palpation (13.2% and
17.8%, respectively; Table 5) and highest for women whose ini-
tial DCIS lesions were p16
1
COX-2
1
Ki67
1
(19.6% and 27.3%,
respectively).
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Factors that were independently associated with subsequent
DCIS included positive or uncertain margins, DCIS lesions that
were p16
1
COX-2
2
Ki67
1
, and those that were ER
2
ERBB2
1
Ki67
1
,
whereas nuclear grade was no longer statistically significant.
The 5- and 8-year risks of subsequent DCIS were highest for
women with defined molecular subtypes of DCIS (Tables 4 and
5). The 5- and 8-year risks of subsequent DCIS were lowest for
women who had disease-free surgical margins of 10 mm or
larger (Table 5).
Risk of Subsequent Invasive Cancer or DCIS by
Risk Group
We next estimated the 5- and 8-year risks of subsequent invasive
cancer and DCIS for four risk groups based on the statistically
significant univariate and multivariable factors reported in Tables
3 and 4, and 5- and 8-year risks reported in Table 5. Among
women who were initially diagnosed with DCIS, 17.3% were in
the lowest-risk group, which had an 8-year risk of subsequent in-
vasive cancer of 4.1%, and 26.8% were in the next to lowest-risk
group, which had an 8-year risk of 6.9% (Table 6). Over a quarter
(27.6%) of the women were in the high-risk group, which had an
8-year risk of subsequent invasive cancer of 19.6%. The 8-year
risk of subsequent invasive cancer was statistically significantly
(P = .018) higher for women with initial DCIS lesions that were
detected by palpation or that were p16
1
COX-2
1
Ki67
1
(19.6%,
95% CI =18.0% to 21.3%) than for women with initial lesions that
were detected by mammography and were p16
2
COX-2
2
Ki67
2
(4.1%, 95% CI = 3.45 to 5.0%).
Women with DCIS could also be divided into groups accord-
ing to the risk for further DCIS. Here, 19.9% of the women ini-
tially diagnosed with DCIS were in the lowest-risk group and had
an 8-year risk of subsequent DCIS of 3.9%, and 21.2% were in the
low-risk group, with an 8-year risk of 10.2% (Table 6). In this
case, only 5.1% of these women were in the high-risk group and
had an 8-year risk of subsequent DCIS of 23.6%.
Discussion
We examined the clinical characteristics of women with DCIS
who were treated by lumpectomy alone and determined the histo-
pathologic and molecular characteristics of their breast lesions to
identify factors associated with the occurrence of subsequent
Table 1. Prevalence of risk factors among women initially treated for ductal carcinoma in situ (DCIS) by lumpectomy alone according to
the type of subsequent tumor event (invasive cancer or DCIS)*
Variable†
No subsequent tumor
event‡ (N = 838), % (No.)
Invasive event
(N = 170), % (No.)
DCIS event
(N = 154), % (No.)
Age at diagnosis, y
40–49 18 (154) 26 (44) 34 (53)
50–59 23 (194) 22 (38) 23 (35)
60–69 24 (198) 22 (38) 21 (33)
70 35 (292) 29 (50) 21 (33)
P§ Referent .6 <.001
Race and/or ethnicity
White 77 (643) 77 (131) 82 (125)
African American 7 (58) 9 (15) 5 (8)
Hispanic 8 (65) 8 (14) 6 (9)
Asian 8 (64) 6 (10) 7 (10)
P§ Referent .6 .4
Family history of breast cancer||
Negative 74 (459) 70 (97) 73 (95)
Positive 26 (164) 30 (42) 27 (36)
P§ Referent .4 .9
Menopausal status¶
Postmenopausal 96 (791) 93 (150) 93 (143)
Premenopausal 4 (32) 7 (12) 7 (11)
P§ Referent .13 .8
Detection method
Mammography 81 (519) 73 (97) 88 (112)
Palpation# 19 (120) 27 (37) 12 (16)
P§ Referent <.001 .06
* Excludes women with a history of breast cancer and women who had radiation therapy or mastectomy.
There was no race and/or ethnicity data missing. However, 22.7% of subjects had missing data for family history, 2.0% for menopausal status, and 22.5% for
detection method.
Control subjects were women with ductal carcinoma in situ who did not have a subsequent tumor event.
§ Wald test calculated from the proportional subdistribution hazards regression coefficients; age-adjusted two-sided test.
|| Defined as at least one first-degree relative (mother, sister, or daughter) with breast cancer.
Women were considered to be postmenopausal if both ovaries had been removed, if they reported their periods had stopped permanently for reasons other than
hysterectomy, if they were currently using postmenopausal hormone therapy, or if they were aged 55 or older.
# Palpable mass found by the woman or by her physician upon physical examination at the time of diagnosis.
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tumors and to determine the risk of subsequent tumors as a func-
tion of these factors. We found initial DCIS lesions that were
detected by palpation or had p16
1
COX-2
1
Ki67
1
expression were
the two factors most strongly associated with risk of subsequent
invasive cancer; however, these factors were not associated with
risk of subsequent DCIS. A little more than a quarter of these
women (27.6%) were categorized as having a high risk of subse-
quent invasive cancer (ie, 19.6% at 8 years). Importantly, many
women (44.1%) who did not demonstrate one of these two factors
were categorized as having a low risk of subsequent invasive cancer
at 8 years (4.1% and 6.9% for the lowest- and low-risk groups,
respectively). In addition, we developed the ability to distinguish
factors associated with risk of subsequent invasive cancer vs risk of
subsequent DCIS, an important clinical goal that could guide
initial therapeutic decisions. We found that initial lesions that
were ER
2
ERBB2
1
Ki67
1
, lesions that were p16
1
COX-2
2
Ki67
1
,
and lesions that had positive or uncertain surgical margins were
strongly associated with risk of subsequent DCIS; however, these
factors were not associated with risk of subsequent invasive
cancer.
Recent molecular studies on DCIS lesions may provide insights
about the biological contributions of p16, COX-2, and Ki67 ex-
pression in p16
1
COX-2
1
Ki67
1
lesions. Molecular studies have
identified markers that distinguish different subtypes of DCIS
(22,28–31) that may relate, in an unknown fashion, to molecularly
defined subtypes of invasive breast cancer. Previously, we reported
in a pilot study that DCIS lesions that express p16 and COX-2 and
have a high proliferative capacity share characteristics with basal-
like invasive tumors (22). Overexpression of p16 has been vali-
dated as a basal-like marker in two recent studies (32,33). In this
report, we demonstrated that expression of these markers results
in a high risk of subsequent invasive cancer but not DCIS. This is
similar to studies of invasive breast cancer where a basal-like sub-
type is associated with worse clinical outcomes (25). Furthermore,
the established role of COX-2 in promoting invasive potential
(34–36) provides a biological rationale for why the p16
1
COX-
2
1
Ki67
1
lesions tend to recur as invasive carcinomas, whereas
p16
1
COX-2
2
Ki67
1
lesions tend to recur as DCIS. Moreover, the
p16
1
COX-2
1
Ki67
1
phenotype is independent of risk conferred by
DCIS lesions detected by palpation. Palpable DCIS lesions
accounted for 15%–20% of DCIS lesions in this study, consistent
with recent studies of women undergoing screening mammog-
raphy (1). That palpable DCIS lesions appear to be more aggres-
sive than mammography-detected lesions is consistent with the
observation that palpable invasive cancer lesions tend to be more
aggressive than mammography-discovered invasive lesions (37).
Attempts to predict risk of subsequent invasive cancer vs DCIS
using a woman’s age at diagnosis and nuclear grade of the DCIS
lesion have met with limited success (5,38), in part, because there
is only moderate agreement in assessing histopathologic character-
istics, such as nuclear grade (39,40). We combined biomarker data
with data pertaining to diagnosis age and nuclear grade to predict
Table 2. Univariate results of histopathologic factors associated with type of subsequent tumor event (invasive cancer or ductal
carcinoma in situ [DCIS])*
Factor†
No subsequent
tumor event‡
(N = 279), % (No.)
Invasive event
(N = 114), % (No.)
Risk of invasive
event, HR§ (95% CI)
DCIS event
(N = 109), % (No.)
Risk of DCIS event,
HR§ (95% CI)
Tumor size, mm
>10 30 (194) 39 (70) 1.2 (0.8 to 1.8) 41 (64) 1.4 (1.0 to 2.1)
10 70 (85) 61 (44) 1.0 (referent) 59 (45) 1.0 (referent)
Margins
Positive 23 (62) 36 (39) 1.6 (0.9 to 2.7) 37 (38) 3.6 (1.8 to 7.2)
Uncertain|| 22 (58) 22 (24) 1.1 (0.6 to 2.1) 24 (25) 2.7 (1.3 to 5.8)
1–1.9 mm disease free 22 (57) 15 (16) 0.8 (0.4 to 1.6) 20 (20) 2.5 (1.1 to 5.4)
2 to <10 mm disease free 10 (26) 9 (10) 1.1 (0.5 to 2.3) 10 (10) 2.3 (0.9 to 5.5)
10 mm disease free 23 (62) 18 (19) 1.0 (referent) 9 (9) 1.0 (referent)
Nuclear grade¶
High 35 (92) 44 (47) 1.2 (0.8 to 2.1) 60 (61) 2.7 (1.5 to 4.8)
Intermediate 33 (85) 35 (38) 1.3 (0.8 to 2.2) 26 (26) 1.4 (0.8 to 2.7)
Low 32 (83) 21 (22) 1.0 (referent) 14 (14) 1.0 (referent)
Necrosis type
Comedo 39 (100) 45 (48) 1.1 (0.8 to 1.6) 45 (46) 1.1 (0.8 to 1.6)
Focal punctate 61 (159) 55 (59) 1.0 (referent) 55 (56) 1.0 (referent)
Quantity of necrosis
Extensive 18 (48) 25 (26) 1.2 (0.8 to 1.8) 28 (29) 1.5 (1.0 to 2.3)
Moderate/scant 82 (212) 75 (80) 1.0 (referent) 72 (73) 1.0 (referent)
* CI = confidence interval; HR = hazard ratio.
Here, 4.0% of specimens had missing data regarding margins, 6.8% for nuclear grade, type of necrosis, and extent of necrosis.
Control subjects were a random sample of women with DCIS who did not have a subsequent tumor event and were frequency matched by year of diagnosis to
the case subjects who were women who had a subsequent tumor event.
§ Adjusted for diagnosis age.
|| Unknown or could not be assessed.
For lesions with more than one type of nuclear grade, an overall grade was assigned according to the highest grade present.
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Table 3. Univariate results of molecular markers associated with type of subsequent tumor event (invasive cancer or ductal carcinoma
in situ [DCIS])*
Factor†
No subsequent
tumor event‡
(N = 186), % (No.)
Invasive event
(N = 72), % (No.)
Risk of invasive
event, HR§ (95% CI)
DCIS event
(N = 71), % (No.)
Risk of DCIS event,
HR§ (95% CI)
ER
Negative 20 (35) 20 (13) 0.8 (0.4 to 1.5) 31 (21) 1.7 (1.0 to 2.9)
Positive 80 (143) 80 (53) 1.0 (referent) 69 (47) 1.0 (referent)
PR
Negative 21 (36) 31 (20) 1.3 (0.7 to 2.1) 33 (21) 1.5 (0.9 to 2.5)
Positive 79 (138) 69 (45) 1.0 (referent) 67 (42) 1.0 (referent)
p53
Positive 10 (17) 10 (6) 0.8 (0.4 to 1.9) 17 (10) 1.8 (0.9 to 3.5)
Negative 90 (153) 90 (57) 1.0 (referent) 83 (49) 1.0 (referent)
ERBB2 oncoprotein
Positive 13 (25) 19 (14) 1.1 (0.6 to 1.9) 30 (21) 2.0 (1.2 to 3.2)
Negative 87 (161) 81 (58) 1.0 (referent) 70 (50) 1.0 (referent)
Ki67
Positive|| 36 (62) 59 (38) 1.7 (1.0 to 2.7) 67 (40) 2.3 (1.3 to 4.1)
Negative 64 (109) 41 (26) 1.0 (referent) 33 (20) 1.0 (referent)
p16
Positive 30 (43) 57 (37) 2.3 (1.4 to 3.8) 41 (26) 1.1 (0.7 to 1.8)
Negative 70 (98) 43 (28) 1.0 (referent) 59 (38) 1.0 (referent)
COX-2
Positive 46 (68) 50 (34) 1.3 (0.8 to 2.0) 34 (22) 0.6 (0.4 to 1.1)
Negative 54 (79) 50 (34) 1.0 (referent) 66 (42) 1.0 (referent)
p16/Ki67
Positive/positive 11 (14) 34 (18) 2.1 (1.2 to 3.8) 33 (18) 2.0 (1.1 to 3.6)
All other groupings 89 (111) 66 (35) 1.0 (referent) 67 (36) 1.0 (referent)
COX-2/Ki67
Positive/positive 18 (24) 33 (18) 1.8 (1.0 to 3.2) 25 (14) 1.1 (0.6 to 2.1)
All other groupings 82 (106) 67 (37) 1.0 (referent) 75 (41) 1.0 (referent)
p16/COX-2/Ki67
Positive/positive/positive 8.5 (10) 23 (12) 2.2 (1.2 to 4.2) 15 (8) 1.2 (0.5 to 2.5)
All other groupings 91.5 (107) 77 (40) 1.0 (referent) 85 (44) 1.0 (referent)
p16/COX-2/Ki67
Positive/negative/positive 2.6 (3) 12 (6) 1.5 (0.6 to 3.6) 19 (10) 3.2 (1.5 to 6.9)
All other groupings 97.4 (114) 88 (46) 1.0 (referent) 81 (42) 1.0 (referent)
ER/PR/ERBB2
Negative/negative/negative 5 (9) 6 (4) 1.1 (0.4 to 3.3) 6 (4) 1.1 (0.4 to 3.0)
All other groupings 95 (172) 94 (64) 1.0 (referent) 94 (65) 1.0 (referent)
ER/ERBB2
Negative/positive 6.4 (11) 5 (3) 0.5 (0.2 to 1.5) 19 (12) 3.0 (1.6 to 5.7)
All other groupings 93.6 (161) 95 (61) 1.0 (referent) 81 (53) 1.0 (referent)
ER/Ki67
Negative/positive 9 (14) 13 (7) 0.8 (0.4 to 1.8) 28 (15) 2.8 (1.5 to 5.2)
All other groupings 91 (136) 87 (48) 1.0 (referent) 72 (39) 1.0 (referent)
ERBB2/Ki67
Positive/positive 7 (11) 18 (10) 1.6 (0.8 to 3.2) 21 (12) 1.9 (1.0 to 3.5)
All other groupings 93 (146) 82 (46) 1.0 (referent) 79 (46) 1.0 (referent)
ER/ERBB2/Ki67
Negative/positive/positive 2.7 (4) 6 (3) 0.9 (0.3 to 2.7) 15 (8) 3.6 (1.7 to 7.8)
All other groupings 97.3 (143) 94 (50) 1.0 (referent) 85 (45) 1.0 (referent)
* CI = confidence interval; COX-2 = cyclooxygenase-2; ER = estrogen receptor; ERBB2 = human epidermal growth factor receptor-2 (HER2/neu-oncoprotein);
HR = hazard ratio; PR = progesterone receptor.
Missing data: 5.2% for ER status, 7.9% for PR status, 10.6% for p53 status, 0% for ERBB2, 10.3% for Ki67, 17.6% for p16, and 15.2% for COX-2.
Control subjects were a random sample of women with DCIS who did not have a subsequent tumor event and were frequency matched by year of diagnosis to
the case subjects who were women who had a subsequent tumor event.
§ Adjusted for diagnosis age.
|| More than 10% positive cells.
risk of invasive cancer. Only initial DCIS lesions that had been
detected by palpation and those mammography-detected lesions
that were p16
1
COX-2
1
Ki67
1
retained a statistically significant
association with invasive cancer in a multivariable analysis. We
have previously reported an association between high nuclear and
subsequent invasive cancer at a median follow-up of about 6 years
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Table 4. Hazard ratios (HRs) and 95% confidence intervals (CIs)
from final multivariable models of clinical and histopathologic
characteristics and molecular markers independently associated
with subsequent tumor events*
Variable
Invasive cancer,
HR (95% CI)
Age at diagnosis, y
1.0 (0.8 to 1.3)
Detection by palpation
(vs mammography)
2.7 (1.4 to 5.5)
Nuclear grade
High vs low 1.0 (0.4 to 2.3)
Intermediate vs low 1.9 (0.8 to 4.3)
p16/COX-2/Ki67
Positive/positive/positive 2.2 (1.1 to 4.5)
All other groupings 1.0 (referent)
Variable† DCIS*, HR (95% CI)
Age at diagnosis, y
0.9 (0.7 to 1.1)
Margins ordinal
(per category increase)
1.3 (1.1 to 1.7)
Nuclear grade
High vs low 1.7 (0.6 to 4.8)
Intermediate vs low 1.3 (0.4 to 4.1)
p16/COX-2/Ki67
Positive/negative/positive 3.7 (1.7 to 7.9)
All other groupings 1.0 (referent)
ER/ERBB2/Ki67
Negative/positive/positive 5.8 (2.4 to 14)
All other groupings 1.0 (referent)
* COX-2 = cyclooxygenase-2; DCIS = ductal carcinoma in situ; ER = estrogen
receptor; ERBB2 = human epidermal growth factor receptor-2 (HER2/
neu-oncoprotein).
Palpable mass found by the woman or by her physician upon physical
examination.
Margins ordinal defined as margin 10 mm disease free = 0; margin 2 to <10
mm disease free = 1; margin 1–1.9 mm disease free = 2; margin uncertain = 3;
margin positive = 4.
margin status, we found that certain combinations of molecular
markers are present in a very small number (5.1%) of DCIS lesions
that are statistically significantly associated with a high risk of sub-
sequent DCIS. The high-risk lesions include lesions that are
ER
2
ERBB2
1
Ki67
1
or p16
1
COX-2
2
Ki67
1
. The striking differences
in lesion characteristics associated with subsequent invasive cancer
compared with subsequent DCIS suggest biological heterogeneity
among DCIS lesions.
Our study has several strengths. First, it is a large population-
based study of women with DCIS treated by lumpectomy alone
that has measures of clinical, histopathologic, and molecular
characteristics by type of subsequent tumor with a median fol-
low-up of about 8 years. Our results are directly applicable to
women with different standard histological types of DCIS
because this study included sufficient numbers of women within
each category. Second, we collected DCIS case subjects from 63
hospitals, thereby minimizing the chance of selection bias
because of specific clinical practices at some hospitals. Third,
our large sample size allowed us to assess the combinations of
biomarkers that independently associated with subsequent inva-
sive cancer vs DCIS by using a multivariable model.
The study also has possible limitations. Clinical factors were
assessed retrospectively, raising the possibility of recall bias.
However, factors that a woman might attribute as causes of
subsequent tumors and thus remember more readily when ques-
tioned, such as presence of family history of breast cancer, were
not associated with subsequent tumors, suggesting that recall
bias did not greatly affect our results. Because we studied
women treated by lumpectomy only, we could not determine
whether various biomarker profiles are more likely to respond
to adjuvant therapies. Additionally, we were only able to
measure biomarkers on a subset of women based on the avail-
ability of tumor blocks at participating hospitals. Our imputa-
tion of missing biomarker data may have resulted in a small
overestimation or underestimation of risk of subsequent tumors.
Similar to challenges presented by assessment of ERBB2 expres-
sion, the immunohistochemical interpretation of COX-2 and
p16 expression can be challenging because of heterogeneity
within DCIS. Refinement of immunohistochemical methods
and validation in additional cohorts and independent labora-
tories are required to validate our results. Likewise, identifica-
tion of additional markers may further refine risk groups and
increase the robustness of risk assessment.
In conclusion, our study adds to the literature in that we iden-
tified combinations of biomarkers in DCIS lesions whose expres-
sion patterns improve estimation of a woman’s risk for subsequent
invasive cancer. Our results suggest that among initial DCIS le-
sions p16
1
COX-2
1
Ki67
1
expression or the ability to be detected
by palpation are the two most important factors that predict higher
risk of subsequent invasive cancer. Conversely, mammographically
detected Ki67-negative DCIS lesions, in particular those that are
also p16 and COX-2 negative, are associated with a lower risk of
subsequent invasive cancer that is similar to the risk of contralat-
eral invasive cancer in women after their first primary invasive
breast cancer (43). Of note, women in the lowest-risk group have
an 8-year risk of invasive breast cancer comparable to an average-
risk 60-year-old woman’s 10-year risk of invasive breast cancer.
(5). We did not observe an association between nuclear grade and
invasive cancer at a median follow-up of about 8 years, consistent
with a study that reported 10-year subsequent tumor rate was not
statistically significantly different between women with high nu-
clear grade and all other grades (41). One explanation for this ob-
servation is that the nuclear grade of the initial DCIS lesion may
be associated with short-term epithelial proliferation but not long-
term proliferation. The contribution of Ki67 to risk of subsequent
invasive cancer may capture, in part, the previously observed asso-
ciation of nuclear grade and subsequent invasive cancer (5) and has
the benefit of signifying short- and long-term risk of subsequent
tumor in this cohort.
Factors associated with subsequent DCIS differed from those
associated with subsequent invasive cancer. Disease-free surgical
margins have been strongly associated with a lower risk of subse-
quent tumors, in particular DCIS (5,42). In this study, when bio-
markers were combined with data pertaining to margin status and
nuclear grade, positive margins remained a strong predictor of
subsequent DCIS, suggesting that persistence of neoplastic cells
from the original DCIS lesion may contribute to subsequent DCIS.
Margin status did not predict subsequent invasive cancer, implying
that most subsequent invasive cancer is an independent process
from any residual nonsurgically removed DCIS. In addition to
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Table 6. Stratification of women into low, intermediate, and high 5- and 8-year risk by type of subsequent tumor event
Risk category* Prevalence in cohort, %†
5-y risk of invasive cancer,
% (95% CI)
8-y risk of invasive cancer,
% (95% CI)
Lowest‡ 17.3 2.1 (1.9 to 2.6) 4.1 (3.4 to 5.0)
Low§ 26.8 4.4 (4.0 to 5.0) 6.9 (6.1 to 8.0)
Intermediate|| 28.3 7.7 (7.0 to 8.5) 11.5 (10.3 to 12.8)
High¶ 27.6 14.1 (13.1 to 15.3) 19.6 (18.0 to 21.3)
Risk category* Prevalence in cohort, %†
5-y risk of DCIS,
% (95% CI)
8-y risk of DCIS,
% (95% CI)
Lowest# 19.9 2.7 (2.4 to 3.2) 3.9 (3.3 to 4.8)
Low** 21.2 7.8 (6.8 to 8.7) 10.2 (8.1 to 12.7)
Intermediate†† 53.8 12.0 (11.4 to 12.6) 14.4 (13.6 to 15.2)
High‡‡ 5.1 19.2 (15.3 to 23.9) 23.6 (18.1 to 34.0)
* Risk groups were defined separately for subsequent invasive cancer and ductal carcinoma in situ (DCIS) based on multivariable associations in Table 4 as well
as level of risk associated with factors in Table 5. CI = confidence interval.
Average prevalence estimated among 2500 cohorts of 1162 women with missing measures imputed as described in the statistical section.
DCIS mammographically detected plus Ki67, cyclooxygenase-2 (COX-2) and p16 triple negative (Ki67–COX-2
2
p16
2
).
§ DCIS mammographically detected plus Ki67 negative and either COX-2 positive (Ki67
2
COX-2
1
) or p16 positive (Ki67
2
p16
1
) or both positive (Ki67
2
COX-2
1
p16
1
).
|| DCIS mammographically detected plus Ki67 positive and either COX-2 positive (Ki67
1
COX-2
1
) or p16 positive (Ki67
1
p16
1
) or COX-2-negative/p16-negative
(Ki67
1
COX-2
2
p16
2
).
Detected by palpation or p16, Ki67, and COX-2 triple positive (p16
1
Ki67
1
COX-2
1
).
# DCIS with margins of 1 mm or greater disease free plus estrogen receptor (ER) positive and HER2/neu-oncoprotein (ERBB2) negative and Ki67 negative
(ER
1
ERBB2
2
Ki67
2
).
** DCIS with margins of 1 mm or greater disease free plus either ER negative, ERBB2 negative (ER
2
ERBB2
2
) or p16 and Ki67 positive (p16
1
Ki67
1
) or COX-2
negative, Ki67 positive (COX-2
2
Ki67
1
) or COX-2 positive, Ki67 positive (COX-2
1
Ki67
1
) or ERBB2 positive, Ki67 positive (ERBB2
1
Ki67
1
).
†† Positive or uncertain margins or ER negative, Ki67 positive (ER
2
Ki67
1
) or ER negative, ERBB2 positive (ER
2
ERBB2
1
).
‡‡ DCIS with margins of 1 mm or greater disease free plus ER negative/ERBB2 positive/Ki67 positive (ER
2
ERBB2
1
Ki67
1
) or p16/Ki67 positive and COX-2 negative
(p16
1
COX-2
2
Ki67
1
).
We also confirmed margin status as a strong predictor of subse-
quent DCIS and identified expression of novel combinations of
biomarkers predicting subsequent DCIS, which differ from those
of that predict subsequent invasive cancer. These markers, com-
pared with nuclear grade, improve the estimation of risk for subse-
quent DCIS.
Many women who have been diagnosed with DCIS have an
inaccurate perception of their risk of subsequent invasive cancer
Table 5. Estimate of 5- and 8-year risks of invasive cancer vs ductal carcinoma in situ (DCIS) for characteristics of women initially diag-
nosed with DCIS that were independently associated with subsequent invasive cancer or DCIS events*
Variable 5-y risk of invasive cancer, % (95% CI) 8-y risk of invasive cancer, % (95% CI)
Overall
7.8 (6.2 to 9.4) 11.1 (9.2 to 13.0)
Detection method
Palpation† 13.2 (12.3 to 14.3) 17.8 (16.2 to 19.4)
Mammography 6.5 (6.3 to 6.6) 9.3 (9.2 to 9.6)
p16/COX-2/Ki67
Positive/positive/positive 19.6 (16.6 to 23.4) 27.3 (22.9 to 33.9)
All other groupings 6.8 (6.6 to 7.0) 9.5 (9.2 to 9.8)
Variable
5-y risk of DCIS, % (95% CI) 8-y risk of DCIS, % (95% CI)
Overall
9.7 (7.9 to 11.4) 11.6 (9.7 to 13.5)
Margins
Positive or uncertain 12.4 (11.7 to 13.0) 14.6 (13.8 to 15.4)
1 to <10 mm disease free 10.2 (9.4 to 10.9) 11.7 (10.4 to 12.3)
10 mm disease free 2.8 (2.5 to 3.3) 4.4 (3.9 to 5.2)
p16/COX-2/Ki67
Positive/negative/positive 20.8 (17.3 to 25.3) 24.9 (20.3 to 33.4)
All other groupings 8.6 (8.4 to 8.8) 10.4 (10.1 to 10.7)
ER/ERBB2/Ki67
Negative/positive/positive 37.2 (29.3 to 49.0) 40.5 (31.7 to 54.3)
All other groupings 8.8 (8.7 to 9.0) 10.6 (10.4 to 10.8)
* CI = confidence interval; COX-2 = cyclooxygenase-2; ER = estrogen receptor; ERBB2 = human epidermal growth factor receptor-2 (HER2/neu-oncoprotein).
Palpable mass found by the woman or by her physician upon physical examination.
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(44). Here, we show that the mode of detection and the bio-
markers p16, COX-2, and Ki67 may be used to help stratify a
woman’s risk of subsequent invasive cancer and to help
her decide whether she should undergo adjuvant therapies. In
addition, these factors may provide insight for targeted
interventions.
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Funding
National Cancer Institute-funded University of California, San Francisco Breast
Cancer SPORE (P50 CA58207); California Breast Cancer Research Program
(2RB-0197). Technical support was received from the UCSF Cancer Center (P30
CA82103), UCSF Cancer Center Tissue and Immunohistochemistry Cores, and
Northern California Surveillance, Epidemiology, and End Results Program.
Notes
K. Kerlikowske, A. M. Molinaro, and M. L. Gauthier contributed equally to
this work.
The study sponsor had no role in the design of the study; the collection,
analysis, and interpretation of the data; the writing of the manuscript or the
decision to publish.
Affiliations of authors: Department of Medicine (KK, KS), General Internal
Medicine Section, Department of Veterans Affairs (KK), Helen Diller Family
Comprehensive Cancer Center (KK, FW, HS, KC, B-ML, TDT), Department
of Laboratory Medicine, Department of Urology, and Department of
Radiation Oncology (FW), Department of Pathology (HS, B-ML, TDT), and
School of Medicine (CJ), University of California, San Francisco, San
Francisco, CA; Division of Biostatistics, Department of Epidemiology and
Public Health in the Yale School of Medicine, New Haven, CT (AMM);
Department of Laboratory Medicine and Pathobiology (HKB) and
Department of Medical Biophysics (MLG), University of Toronto, Campbell
Family Institute for Breast Cancer Research, Princess Margaret Hospital,
Toronto, ON, Canada; California Pacific Medical Center Research Institute,
San Francisco, CA (JB).
Page 11
  • Source
    • "The transition from low-grade DCIS to high-grade DCIS or to high-grade invasive carcinoma is deemed unlikely [8][9][10] 12]. The grade distribution of DCIS has been studied in mostly small series [6,[14][15][16][17][18] , or only included screendetected cases (Table 1) [19] . More insight in this distribution based on larger numbers in both screened and non-screened populations is of paramount importance and may improve our estimates of overdiagnosis. "
    [Show abstract] [Hide abstract] ABSTRACT: Background The incidence of ductal carcinoma in situ (DCIS) has rapidly increased over time. The malignant potential of DCIS is dependent on its differentiation grade. Methods Our aim is to determine the distribution of different grades of DCIS among women screened in the mass screening programme, and women not screened in the mass screening programme, and to estimate the amount of overdiagnosis by grade of DCIS. We retrospectively included a population-based sample of 4232 women with a diagnosis of DCIS in the years 2007–2009 from the Nationwide network and registry of histopathology and cytopathology in the Netherlands. Excluded were women with concurrent invasive breast cancer, lobular carcinoma in situ and no DCIS, women recently treated for invasive breast cancer, no grade mentioned in the record, inconclusive record on invasion, and prevalent DCIS. The screening status was obtained via the screening organisations. The distribution of grades was incorporated in the well-established and validated microsimulation model MISCAN. Results Overall, 17.7 % of DCIS were low grade, 31.4 % intermediate grade, and 50.9 % high grade. This distribution did not differ by screening status, but did vary by age. Older women were more likely to have low-grade DCIS than younger women. Overdiagnosis as a proportion of all cancers in women of the screening age was 61 % for low-grade, 57 % for intermediate-grade, 45 % for high-grade DCIS. For women age 50–60 years with a high-grade DCIS this overdiagnosis rate was 21–29 %, compared to 50–66 % in women age 60–75 years with high-grade DCIS. Conclusions Amongst the rapidly increasing numbers of DCIS diagnosed each year is a significant number of overdiagnosed cases. Tailoring treatment to the probability of progression is the next step to preventing overtreatment. The basis of this tailoring could be DCIS grade and age.
    Full-text · Article · Dec 2016 · Breast cancer research: BCR
  • Source
    • "HER-2, a transmembrane tyrosine kinase, is over-expressed in 30-40% of DCIS [3][4][5]. High HER-2 and/or Ki-67 expression are risk factors of local recurrence of DCIS [6]. Since targeted antibodies have successfully demonstrated activity in treating HER-2 positive breast cancer, conjugated antibodies have been explored for enhancing their potency [7, 8]. "
    [Show abstract] [Hide abstract] ABSTRACT: The standard treatment for ductal carcinoma in situ (DCIS) of the breast is surgical resection, followed by radiation. Here, we tested localized therapy of DCIS in mice using the immunoconjugate 225Ac linked-trastuzumab delivered through the intraductal (i.duc) route. Trastuzumab targets HER-2/neu, while the alpha-emitter 225Ac (half-life, 10 days) delivers highly cytotoxic, focused doses of radiation to tumors. Systemic 225Ac, however, elicits hematologic toxicity and at high doses free 213Bi, generated by its decay, causes renal toxicity. I.duc delivery of the radioimmunoconjugate could bypass its systemic toxicity. Bioluminescent imaging showed that the therapeutic efficacy of intraductal 225Ac-trastuzumab (10-40 nCi per mammary gland; 30-120 nCi per mouse) in a DCIS model of human SUM225 cancer cells in NSG mice was significantly higher (p<0.0003) than intravenous (120 nCi per mouse) administration, with no kidney toxicity or loss of body weight. Our findings suggest that i.duc radioimmunotherapy using 225Ac-trastuzumab deserves greater attention for future clinical development as a treatment modality for early breast cancer.
    Full-text · Article · Apr 2016 · Oncotarget
  • Source
    • "Other studies have examined the utility of biomarkers in predicting subsequent invasive carcinoma. Kerlikowske et al. studied the characteristics of patients with DCIS who later developed invasive carcinoma [17]. In the 170 patients with progression, a combination of Ki67 positivity with positivity for p16 and COX2 in the initial in situ lesion was significantly associated with subsequent invasion. "
    [Show abstract] [Hide abstract] ABSTRACT: Breast cancer screening has led to a dramatic increase in the detection of pre-invasive breast lesions. While mastectomy is almost guaranteed to treat the disease, more conservative approaches could be as effective if patients can be stratified based on risk of co-existing or recurrent invasive disease.Here we use a range of biomarkers to interrogate and classify purely non-invasive lesions (PNL) and those with co-existing invasive breast cancer (CEIN). Apart from Ductal Carcinoma In Situ (DCIS), relative homogeneity is observed. DCIS contained a greater spread of molecular subtypes. Interestingly, high expression of p-mTOR was observed in all PNL with lower expression in DCIS and invasive carcinoma while the opposite expression pattern was observed for TOP2A.Comparing PNL with CEIN, we have identified p53 and Ki67 as predictors of CEIN with a combined PPV and NPV of 90.48% and 43.3% respectively. Furthermore, HER2 expression showed the best concordance between DCIS and its invasive counterpart.We propose that these biomarkers can be used to improve the management of patients with pre-invasive breast lesions following further validation and clinical trials. p53 and Ki67 could be used to stratify patients into low and high-risk groups for co-existing disease. Knowledge of expression of more actionable targets such as HER2 or TOP2A can be used to design chemoprevention or neo-adjuvant strategies. Increased knowledge of the molecular profile of pre-invasive lesions can only serve to enhance our understanding of the disease and, in the era of personalised medicine, bring us closer to improving breast cancer care.
    Full-text · Article · Dec 2015 · Oncotarget
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