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

670s Tumor Biology
10557 General Poster Session (Board #44F), Mon, 1:15 PM-5:15 PM
Exosomal-miRNA profiles as diagnostic biomarkers in cervical cancer.
Presenting Author: Hirotaka Nishi, Department of Obstetrics and Gynecology,
Tokyo Medical University, Tokyo, Japan
Background: MicroRNA (miRNA) expression is altered in cancer cells and
associated with the development and progression of various types of
cancer. miRNA could serve as diagnostic or prognostic biomarker for
cancer patients. Our study was designed to analyze circulating exosomalmiRNA
in patients with cervical cancer. Methods: Total RNA was extracted
from serum in healthy women and patients with cervical intraepithelial
neoplasia (CIN) and cervical cancer. We first explored miRNA expression
profiles using miRCURY LNA microRNA Array (Exiqon) in each six
malignant and healthy serum samples. miRNAs with significant differences
in expression were validated in larger sample sets by a quantitative
real-time RT-PCR using TaqMan gene expression assays (Applied Biosystems).
Results: Six of 1223 miRNAs compared in serum samples from
cervical cancer and normal control were found to have a �3.0-fold change
with a P value �0.01 using miRCURY LNA microRNA Array. In a validation
set (n�101), expression of 3 of the 6 miRNAs, miR-483-5p, miR-1246
and miR-1275, was significantly different between the cervical cancer
(n�40) and control (n�20) samples. We also found that the median levels
of these miRNAs were significantly higher in patients with cervical cancer
(n�40) than those in CIN (n�41). Circulating miRNAs were not correlated
with clinical-pathological parameters except for miR-1246. Compared to
patients with squamous cell carcinoma, the miR-1246 level was significantly
higher in those with adenocarcinoma. Whereas, receiver-operator
characteristic (ROC) curve analyses suggest that these serum miRNAs may
be useful markers for discriminating patients with cervical cancer from
healthy controls. Conclusions: Expression of circulating miR-483-5p,
miR-1246 and miR-1275 is significantly higher in the blood of cervical
cancer patients compared to controls and may potentially serve as a useful
biomarker for cervical cancer diagnosis. Especially, miR-1246 may be a
candidate for early detection of cervical adenocarcinoma. Larger-scaled
studies are warranted to fully explore the role of circulating exosomalmiRNAs
in cervical cancer.
10559 General Poster Session (Board #44H), Mon, 1:15 PM-5:15 PM
Use of next-generation sequencing (NGS) to detect high frequency of
targetable alterations in primary and metastatic breast cancer (MBC).
Presenting Author: Lajos Pusztai, University of Texas M. D. Anderson
Cancer Center, Houston, TX
Background: The aim of this study was to assess the frequency of genomic
alterations in breast cancer potentially treatable with approved targeted
agents or investigational drugs in clinical trials. NGS was performed in a
CLIA setting (Foundation Medicine). Methods: DNA was extracted from
needle biopsies of 33 pre-therapy primary and 17 MBCs (mean age 52 yrs;
58% ER�, 20% HER2�, 30% triple negative) obtained prospectively for
biomarker discovery and preserved in RNAlater. Patients with MBC
received an average of 7 drugs (range 5-17) including adjuvant therapy
before biopsy for this research; 13 biopsies were from soft tissues, 3 from
liver and 1 from bone. Sequencing was targeted to 3230 exons in 182
cancer-related genes and 37 introns in 14 genes often rearranged in
cancer. Average median depth was �1200x. Results: All biopsies yielded
sufficient DNA. NGS revealed a total of 117 known driver mutations across
36 genes (per-tumor average�2.5, range 1-6), including 37 base substitutions
(32%), 28 indels (24%), 42 amplifications (36%) and 10 homozygous
deletions (9%). NGS identified HER2 gene amplification in 6/7 cases
scored HER2� by FISH. The average number of functionally important
alterations was surprisingly similar, 2.3 in primaries vs 2.8 in heavily
treated MBCs (p�0.32). Remarkably, 25/33 (76%) of primary and 14/17
(82%) of MBCs had at least 1 genomic alteration targetable with an FDA
approved drug or novel agent in clinical trials. These included: ERBB2
alterations (n�9), PIK3CA mutations (n�8), NF1 mutations (n�4, candidate
for PI3K/MEK inhibitors), AKT1-3 mutations (n�5, PI3K inhibitors),
BRCA1/2, (n�6, PARP inhibitors), and CCND2 (n�3)/CDKN2A (n�3)
mutations (CDK inhibitors). Numerous other alterations with less apparent
therapeutic implications were also observed. Conclusions: Comprehensive
NGS profiling in breast cancer needle biopsies showed high frequency of
genomic alterations linked to a clinical treatment option or clinical trials of
targeted therapies. These results demonstrate it is feasible to use NGS to
guide targeted therapy. Prospective testing of the diagnostic/predictive
value of this patient selection approach is currently under way.
10558 General Poster Session (Board #44G), Mon, 1:15 PM-5:15 PM
Evaluation of gene expression by RNA-seq after single dose of trastuzumab
(T) reveals predictors of pathologic complete response (pCR) in HER2positive
early breast cancer. Presenting Author: Natalie Galanina, Yale
University School of Medicine, Yale Comprehensive Cancer Center, New
Haven, CT
Background: The use of a ‘brief exposure’ to single agent T allows the
measurement of dynamic changes in the transcriptome that may predict
response to T-based combinations. We have shown that most gene
expression changes in HER2� tumors treated with T occur in tumors that
ultimately achieve a pCR. Our further analysis suggests several patterns of
transcriptional change in pCR tumors suggesting different mechanisms of
action of T. RNA-seq analysis provides more in-depth annotation of these
mechanisms. Methods: Fresh tumor core biopsies were taken at a 2 week
time point after a single dose of T (8mg/m2) from 80 HER2� early breast
cancer patients enrolled on a clinical trial of T�T�C. Nucleic acids were
extracted using Qiagen AllPrep and were analyzed with Illumina HT12v3
Beadchip and Illumina 610 QUAD V1 SNP arrays. RNA was also processed
for sequencing using the Ovation RNA-Seq System and paired-end sequenced
using an Illumina Genome Analyzer IIxRNA-seq data was analyzed
with Tophat/Cufflinks. Network analysis was performed with Metacore.
Results: Among pCR tumors, distinct patterns of differential expression
pre/post T were observed, in both microarray and RNA-seq data. ERBB2
down-regulation was characteristic of pCR in one subgroup by microarray.
In this group, differentially expressed genes belonged to interaction
networks involved in apoptosis and cell cycle regulation. In contrast,
tumors with no change in ErbB2 showed differentially expressed genes that
belonged to networks related to chromatin assembly and regulation of
immune pathways. NOLC1, RPL41, ZCHHC17, and B2M had altered
alternative splicing product distributions in both groups. In the ERBB2
down-regulated group, genes with changed expression were enriched for
targets of STAT3 and YY1. Conclusions: RNA-seq and microarray reveal
distinct responses in tumors that achieve pCR to T-containing regimens.
These methods provide predictive markers for validation in subsequent
clinical trials.
10560 General Poster Session (Board #45A), Mon, 1:15 PM-5:15 PM
EGFR exon 20 insertion mutations: Incidence and clinicopathologic
characteristics in U.S. patients with lung adenocarcinoma. Presenting
Author: Maria E. Arcila, Memorial Sloan-Kettering Cancer Center, New
York, NY
Background: Activating insertion mutations in exon 20 of EGFR are reported
in a small subset of lung adenocarcinomas (ADC). In contrast to the classic
EGFR mutations, they appear to confer primary resistance to currently
approved EGFR tyrosine kinase inhibitors. Their incidence and clinicopathologic
features are not well established. Methods: Lung ADCs (n�1500)
were screened for major activating mutations in EGFR (exons 19 and 21)
and KRAS (exon 2). Negative cases were tested for EGFR exon 20
insertions by a PCR-based sizing assay. Extended testing for additional
recurrent point mutations in EGFR, KRAS, BRAF, NRAS, PIK3CA, MEK1
and AKT was performed in all cases by Sequenom mass spectrometry. A
subset of cases was also tested for ALK rearrangements by FISH. Results:
We identified 32cases withEGFRexon 20 insertions, accounting for 11% of
all EGFR mutations. EGFRexon 20 insertions were mutually exclusive with
the other genetic alterations tested except for PIK3CA mutations. The
incidence was higher among never-smokers (p�0.0001) but there was no
association with sex, ethnic origin or stage at diagnosis. Insertions were 3,
6, 9 or 12bp; 9bp insertions were most common (50%, 16/32). Morphologically,
90% of tumors were moderate to poorly differentiated with a
predominant mixed ADC phenotype. Conclusions: EGFR exon 20 testing
may identify a unique subset of EGFR mutant lung ADCs which is
significantly larger than previously reported, making this the third most
common type of EGFR mutation after exon 19 deletions and L858R. This
population could potentially benefit from alternate targeted therapies,
many of which are currently in clinical development.
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