Annual Meeting Proceedings Part 1 - American Society of Clinical ...

573 General Poster Session (Board #5H), Sat, 8:00 AM-12:00 PM
In situ quantitative measurement of mRNA to predict response to trastuzumab
in a cohort of metastatic breast cancer patients. Presenting Author:
Maria Vassilakopoulou, Yale University, New Haven, CT
Background: Trastuzumab therapy is currently selected based on assessment
of HER2 by either immunohistochemistry (IHC) for protein overexpression
or fluorescence in situ hybridization (FISH) to detect gene
amplification. We sought to determine the predictive value of in situ
quantitative measurement of mRNA on a cohort of trastuzumab-treated
metastatic breast cancer patients. Methods: A tissue microarray composed
of ninety metastatic breast cancers treated with various chemotherapy
regimens combined with trastuzumab was constructed. HER2 intracellular
domain (ICD), HER2 extracellular domain (ECD) and HER2 mRNA were
assessed using the AQUA method for quantitative immunofluorescence.
For HER2 protein evaluation CB11was used to measure ICD and SP3 to
measure ECD of the HER2 receptor. In addition, HER2 mRNA status was
assessed using the RNAscope assay ERRB2 probe according to manufacturer’s
protocol modified for detection with Cy-5 Tyramide. Cytokeratin was
used in order to create the tumor mask and signal was quantified within the
mask. Primary endpoints included time to progression (TTP) from trastuzumab
initiation and overall survival (OS) times. Statistical analysis was
performed using Kaplan-Meier analysis and the Cox proportional hazard
model. Results: HER mRNA was tightly correlated with ICD HER2, as
measured by CB11 (r2�0.35), but not with ECD HER2 as measured by
SP3 (r2�0.14). Both ICD HER2 and HER2 mRNA were predictive of
response to trastuzumab, but ECD was not. Multivariate analysis including
age, histological grade and hormone receptor status shows the ICD to be
predictive of TTP (p�0.0377). HER2 mRNA levels were significant for
prediction of TTP (p�0.0344) in multivariate analysis including age and
histological grade. Conclusions: The expression of ICD, as detected by
CB11 and HER2 mRNA levels were significantly associated with TTP in this
trastuzumab-treated metastatic cohort. The ECD, as detected by SP3 is
neither predictive of response, nor tightly correlated with HER2 mRNA. In
situ assessment of HER2 mRNA has the potential to identify breast cancer
patients who derive benefit from trastuzumab treatment.
575 General Poster Session (Board #6B), Sat, 8:00 AM-12:00 PM
Fertility preservation options and the young breast cancer patient: A survey.
Presenting Author: Manuela Jacobsen Junqueira, Memorial Sloan-
Kettering Cancer Center, New York, NY
Background: Approximately 15% of breast cancers (BC) are diagnosed in
reproductive aged women. Management of the disease in this age group
frequently includes chemotherapy and hormonal therapy, which can both
affect fertility. Considering that age at first delivery has been steadily
increasing, young women may face BC before completion of childbearing.
Methods: In this prospective study, women referred to our institution for
surgical treatment of BC were asked, before their first visit, to fill out a
questionnaire regarding their reproductive history and fertility preservation
knowledge. Eligible patients included women between the ages of 18 and
45, with a newly diagnosed BC, who had not yet started treatment. Results:
Sixty women were eligible with a median age of 40 (range 20-45). 98% of
responders (59 out of 60) had been diagnosed within the previous 2
months. 78% (47/60) had a college or post-graduate degree. 80% (48/60)
had been pregnant before, while 86.5% (45/52) reported having had
children. 81% (47/58) were premenopausal, and only 3 patients reported
not having had periods for more than 1 year. 50% of responders (30/60)
declared no interest in future childbearing, 25% were definitely interested,
and 25% were undecided. However, only 9% (5/57) reported having
received information on fertility preservation options before the survey.
Women who have been pregnant were significantly less likely to consider
fertility preservation options prior to treatment (egg/embryo/ovarian tissue
cryopreservation [6% vs. 50%, p�0.001]), or after treatment (egg/embryo
donation, surrogacy or adoption [6% vs. 58%, p�0.0001]). Conclusions:
This pilot study was designed to gather information on reproductive health
of newly diagnosed young BC patients and to assess their willingness to
consider various fertility preservation options before or after treatment. Our
study population consisted of mostly women who had been pregnant and
had children. We found that 50% of the women were unsure or wanted
future children. Yet, only 9% had received information on fertility options
at diagnosis. This pilot study highlights the need for education and/or
intervention in fertility preservation options for young women with breast
cancer.
Breast Cancer—HER2/ER
25s
574 General Poster Session (Board #6A), Sat, 8:00 AM-12:00 PM
ER� mRNA expression in ER�-negative and triple-negative breast cancers.
Presenting Author: Young Choi, Yale University School of Medicine,
Bridgeport, CT
Background: ER� is the main prognostic and therapeutic marker in breast
cancer (BC). About 30% of BC cases are negative for ER (ER�-) and do not
benefit from antiestrogen therapy (TAM). We aim to study ER-beta (ER�)
expression in ER�- and triple negative (TN) cancers to explore alternate
pathway of treatment in this cohort. Methods: We studied 67 ER�- BC cases
including 44 TN together with 74 ER� �BC cases obtained from patients
aged 29 to 97 years old between 2003 and 2010. The histology included
110 intraductal, 12 medullary and 19 other types. 78 cases were grade 3,
52 were grade 2, and 11 were grade 1. RNA was extracted from FFPE and
mRNA levels of ER� isoform and ER� were determined by real-time
quantitative reverse transcription PCR. IHC stains were done on TMA the
sections for ER�, PR, Her-2, Ki-67, CK5/6 and Cyclin D1. Results: ER�
isoforms were highly expressed in ER�-, TN, basal-like and HER2 type BC
cases. ER�2 was the major ER� variant expressed. Ki-67 proliferating cells
(�20% nuclear staining) were mostly in ER�- rather than ER�� cases
(69.0% vs. 31.0%) as were cyclin D1- cells (82.2% vs. 17.8%). On the
other hand, in ER�� BC, ER� mRNA expression was consistently high and
upregulated, and ER�, low and down regulated, and the ratio of ER�� to
ER�� ranged from 3 to 100. ER�1, 2 and 5 were co-expressed with ER� in
56%, 63%, and 30% of cases, respectively. Overall, ER� mRNA levels did
not show any significant correlation with age, tumor size, lymph node status
and histological grades. Conclusions: ER�-dependent proliferating tumor
cells may render them more sensitive to TAM, and increase the effectiveness
of TAM and its metabolites in ER�- and TN cases. Increased overall
survival after adjuvant TAM ER�-BC may be directly related to ER�
over-expression. ER� isoform is potential selective therapeutic target in a
sub-cohort of ER�- BC. Additionally, when ER� and ER� are co-expressed,
ER� appears to play a distinct role from its action in ER�- BC.
Types (# of cases) ER�1* ER�2* ER�5*
ER�- (67) 39 42 15
TN (44) 41 46 25
HER2� (38) 66 82 37
HER2 type (19) 74 85 47
Basal-like type (11) 51 64 36
Ki-67� /ER�- (27) 55 70 37
Grade 3 (78) 41 46 31
Grade 2 (52) 39 54 29
* % mRNA expression.
576 General Poster Session (Board #6C), Sat, 8:00 AM-12:00 PM
Estrogen receptor (ER) signaling in normal, BRCA (B) 1 and B2 mutation
associated, and ER-positive breast cancer (BC) mammary cells. Presenting
Author: Elgene Lim, Dana-Farber Cancer Institute, Boston, MA
Background: ER is expressed in �70% of sporadic BC and activates genes
driving cell proliferation and tumorigenesis. B1 and B2 mutation associated
BC have predominant triple negative and ER� phenotypes respectively,
yet hormonal manipulation with oophorectomy reduces BC risk in
both B1 and B2 carriers. We have previously performed genome-wide
analysis of ER binding sites in MCF-7 BC cells, and identified distinct
mechanisms of ER signaling. Here, we seek to elucidate differences in ER
signaling between non-malignant (normal, B1 and B2) and ER� BC
mammary cells. Methods: Breast tissue were obtained from patients
undergoing reduction mammoplasties (normal), prophylactic mastectomies
(B1 and B2 carriers) and surgical excision of ER� BC. After
dissociation into a single-cell suspension, ER� EpCAM�CD49f- mature
luminal (ML) and EpCAM�CD49f� luminal progenitor (LP) subpopulations
were obtained by flow cytometry. Following estrogen stimulation, RNA
was extracted for gene microarray analysis. ER chromatin immunoprecipitation
and DNA sequencing (ChIP-seq) was performed and DNA libraries
prepared and sequenced. Results: Triplicates ofnormal, prophylactic B1
and B2, and ER� BC tissues were analyzed.Gene ontology analysis
indicates different gene expression signatures between non-malignant
tissue and MCF-7 BC cells following estrogen stimulation. Genes that
promotes cell cycling and proliferation were downregulated in nonmalignant
tissue, but were upregulated in BC cells (P � 10-5 ). Hierarchical
clustering of the gene microarray by mutation status revealed distinct LP
gene signatures in normal, B1 and B2 breast tissue, whilst the ML
signatures were largely indistinguishable between the three tissue types.
Conclusions: There are contrasting differences in ER signaling between
normal mammary and BC cells, with estrogen having anti-proliferative
effects in normal luminal cells compared to pro-proliferative effects in BC.
Our data suggest that alterations in ER signaling play a major role in breast
tumorigenesis. Differences in the LP population between B1 and B2 breast
tissue may account for their distinct BC phenotypes.
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