Annual Meeting Proceedings Part 1 - American Society of Clinical ...
Annual Meeting Proceedings Part 1 - American Society of Clinical ...
Annual Meeting Proceedings Part 1 - American Society of Clinical ...
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664s Tumor Biology<br />
10532 General Poster Session (Board #41E), Mon, 1:15 PM-5:15 PM<br />
VEGF polymorphism may be associated with overall survival in lapatinibtreated<br />
breast cancer patients with brain metastases. Presenting Author:<br />
Colin F. Spraggs, GlaxoSmithKline, Stevenage, United Kingdom<br />
Background: Lapatinib is a dual HER2/EGFR tyrosine kinase inhibitor (TKI)<br />
approved in combination with capecitabine or letrozole for patients with<br />
HER2� metastatic breast cancer (MBC). Consistent with TKI therapies,<br />
patient response is variable and suggestive <strong>of</strong> additional determinants <strong>of</strong><br />
sensitivity and resistance. This exploratory pharmacogenetic study sought<br />
to identify germline genetic variants that associate with lapatinib treatment<br />
outcomes in HER2� MBC patients. Methods: Fifty five functional single<br />
nucleotide polymorphisms (SNPs) in 24 candidate genes were evaluated in<br />
a subset <strong>of</strong> MBC patients participating in two clinical trials: EGF105084:<br />
lapatinib monotherapy in HER2� MBC patients with recurrent brain<br />
metastases following trastuzumab and cranial radiotherapy (n�120) and<br />
EGF104900: lapatinib plus trastuzumab (n�92) and lapatinib monotherapy<br />
(n�103) in HER2� MBC patients with disease progression<br />
following trastuzumab. Testing for associations <strong>of</strong> SNPs with progression<br />
free survival (PFS) and overall survival (OS) during lapatinib treatment was<br />
performed using Cox proportional hazards methods, with covariate adjustment.<br />
Markers were considered to be significantly associated if they<br />
achieved a predefined multiple testing threshold <strong>of</strong> p�0.0003. Results: No<br />
SNPs were significantly associated with PFS in either study. A SNP in<br />
VEGFA (rs3025039, 936C�T) was significantly associated with improved<br />
OS (p�0.0002) for the T allele carriers, with an allelic hazard ratio <strong>of</strong> 0.21<br />
(0.08-0.52) in EGF105084. The association was not seen in EGF104900.<br />
This SNP is located in the 3’ UTR gene region, the T allele is associated<br />
with lower serum VEGFA levels and reduced breast cancer risk [Krippl et al,<br />
2003, Int J Cancer 106: 468; Kataoka et al, 2006, Cancer Epidemiol<br />
Biomarkers Prev, 15:1148]. Conclusions: A germline variant in VEGFA may<br />
be associated with survival outcome in MBC patients with brain metastases<br />
who are treated with lapatinib. This may represent activation <strong>of</strong> VEGF<br />
angiogenic pathways to overcome HER2 inhibition in patients carrying the<br />
higher expression genotype. These associations are considered exploratory<br />
and require confirmation in an independent dataset.<br />
10534 General Poster Session (Board #41G), Mon, 1:15 PM-5:15 PM<br />
Molecular characterization <strong>of</strong> circulating tumor cells in human metastatic<br />
colorectal cancer. Presenting Author: Jorge Barbazan, Translational Oncology<br />
Laboratory, Medical Oncology Department, Santiago de Compostela,<br />
Spain<br />
Background: Metastatic colorectal cancer (mCRC) relies on the detachment<br />
<strong>of</strong> aggressive malignant cells from the primary tumor into the bloodstream,<br />
being this Circulating Tumor Cells (CTC) the principal source <strong>of</strong> the further<br />
metastasis. <strong>Clinical</strong>ly, the presence <strong>of</strong> CTC is associated with poor<br />
prognosis and there exists a clear necessity for more specific and efficient<br />
chemotherapies in the treatment <strong>of</strong> mCRC. Our aim was to overcome this<br />
adverse scenario through the massive molecular pr<strong>of</strong>iling <strong>of</strong> the CTC<br />
population isolated from mCRC patients. Methods: CTC’s were magnetically<br />
isolated by using anti-EpCAM coupled magnetic beads from 6 mCRC<br />
patients and 3 healthy controls. The presence <strong>of</strong> CTCs was evaluated by<br />
citokeratins 8, 18 and 19 staining. RNA from CTCs was amplified by a<br />
whole transcriptome amplification system (WTA) and resulting material was<br />
hybridized onto gene expression arrays. Bioinformatics were used to array<br />
analysis. Selected genes were validated by RT-qPCR. Results: 410 transcripts<br />
were found to specifically characterise the CTC population after<br />
array signals comparison between mCRC patients and controls. Gene-gene<br />
interaction analysis generated networks related with cell migration, adhesion<br />
or apoptosis resistance. Gene ontology revealed similar functions.<br />
Signalling pathways such as RhoA, PKA, ILK, integrins or actin cytoskeleton<br />
signalling were found as relevant in CTCs. Eleven genes (TGFB1, APP,<br />
TIMP1, CD9, CLU, ITGB5, LIMS1, RSU1, VCL, BMP6 and TLN1) were<br />
validated by real-time PCR in 20 mCRC patient and 10 control samples. All<br />
genes showed a significantly higher expression in patients (p�0.0001).<br />
Except from TLN1 and CD9, all genes had a prognostic value in terms <strong>of</strong><br />
progression free survival (p�0.05). Conclusions: We described the molecular<br />
pr<strong>of</strong>iling <strong>of</strong> CTCs in stage IV CRC patients, characterized by a migratory<br />
and invasive phenotype. Specific and sensitive diagnostic/prognostic markers<br />
were obtained. Our strategy represents an innovative and promising<br />
approach in the clinical management <strong>of</strong> mCRC patients.<br />
10533 General Poster Session (Board #41F), Mon, 1:15 PM-5:15 PM<br />
Isolation <strong>of</strong> circulating tumor cells using a novel EMT-based capture<br />
method. Presenting Author: Rhonda Lynn Bitting, Duke Cancer Institute,<br />
Durham, NC<br />
Background: Circulating tumor cells (CTCs) have potential prognostic,<br />
predictive and surrogate implications in oncology. In patients with metastatic<br />
castration-resistant prostate cancer (CRPC) and breast cancer (BC),<br />
we have shown that CTCs isolated using epithelial cell adhesion molecule<br />
(EpCAM) ferromagnetic capture express mesenchymal markers, including<br />
N- and O-cadherin, suggesting phenotypic plasticity and the presence <strong>of</strong><br />
epithelial-mesenchymal transitions (EMT). Therefore, we postulate that<br />
during metastasis, tumor cells exist as a spectrum <strong>of</strong> epithelial to<br />
mesenchymal phenotypes and may not be captured with existing EpCAMbased<br />
CTC technology. The goal <strong>of</strong> this study is to identify CTCs in CRPC<br />
and BC patients using a novel mesenchymal-based capture method.<br />
Methods: In patients with advanced CRPC and BC, two EDTA and one<br />
CellSave tube <strong>of</strong> blood are collected. Using the CellSearch system (Veridex,<br />
USA), CTCs are captured with either anti-N-cadherin (N-cad) or anti-Ocadherin<br />
(O-cad) ferr<strong>of</strong>luid and detected cells (events) are defined as<br />
beta-catenin and DAPI positive, CD45 negative intact cells. We evaluated<br />
the performance <strong>of</strong> these EMT-based ferr<strong>of</strong>luids in healthy volunteers,<br />
control cells, and in a pilot study <strong>of</strong> patients with CRPC, and compared<br />
enumeration <strong>of</strong> cells using novel vs. standard EpCAM-based capture<br />
methods. Results: In healthy volunteers, rare events were detected using<br />
the novel capture methods. In CRPC patients, O-cad capture detected more<br />
events in 3 <strong>of</strong> 5 subjects than EpCAM-based capture, and the majority <strong>of</strong><br />
captured cells were cytokeratin negative. See table below. Conclusions:<br />
These preliminary results suggest that a novel CTC phenotype exists in<br />
CRPC based on EMT properties, particularly overexpression <strong>of</strong> O-cadherin.<br />
Further characterization <strong>of</strong> these cells from patients with advanced PC, BC,<br />
and other solid tumors will provide insight into EMT and metastasis biology.<br />
O-cad capture, beta-catenin� N-cad capture, beta-catenin� EpCAM capture, cytokeratin�<br />
Healthy volunteers 0-51 events (mean 5.95) n�21 0-4 events (mean 0.28) n�25 NA<br />
CRPC patients 0-465 events (mean 138.4) n�5 0-2 events (mean 1.2) n�5 1-123 CTCs (mean 50.8) n�5<br />
10535 General Poster Session (Board #41H), Mon, 1:15 PM-5:15 PM<br />
Pr<strong>of</strong>iling <strong>of</strong> circulating tumor cells isolated from 105 metastatic gastric<br />
cancer patients revealed HER2 overexpression/activation for potential use<br />
in clinical setting. Presenting Author: Saswati Hazra, Prometheus Laboratories<br />
Inc., San Diego, CA<br />
Background: Gastric cancer (GCA) is the second leading cause <strong>of</strong> cancer<br />
mortality in the world. Survival <strong>of</strong> patients with advanced GCA treated with<br />
chemotherapy remains low. New targeted therapies are urgently needed.<br />
There is mounting evidence <strong>of</strong> the role <strong>of</strong> HER2 overexpression in patients<br />
with GCA, and it has been highly correlated to poor outcomes with more<br />
aggressive disease. The ability to accurately determine HER2 status by<br />
testing circulating tumor cells (CTCs) may improve patient treatment by<br />
allowing ongoing assessment <strong>of</strong> HER2 status during treatment and/or<br />
identifying additional patients who could potentially benefit from HER2targeted<br />
therapy. Methods: The Collaborative Enzyme Enhanced Reactiveimmunoassay<br />
(CEER) was utilized to determine the expression and activation<br />
(phosphorylation) levels <strong>of</strong> HER2 in CTCs isolated from blood specimens<br />
obtained from 105 metastatic GCA patients. Results: Utilizing the CEER<br />
platform, the levels <strong>of</strong> HER2 expression and phosphorylation were determined<br />
for CTCs isolated from metastatic GCA patients. Evaluable CTCs<br />
were found in 33% (35/105) <strong>of</strong> enrolled patients. Out <strong>of</strong> 35 patients, 7<br />
patients (20%) have high HER2 over expression, 6 patients (17%) have<br />
moderate HER2 expression and 11 patients (31%) have HER2 activation<br />
(phospho positive) with no HER2 over-expression. Conclusions: When CTCs<br />
were present, the CEER assay identified varying levels <strong>of</strong> HER2 involvements<br />
in 68% <strong>of</strong> metastatic GCA patients. HER2 positive CTCs could serve<br />
as a prognostic and/or predictive marker in patients with advanced GCA and<br />
CTC-HER2 pr<strong>of</strong>ile shifts can be utilized to monitor the treatment efficacy.<br />
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