Identifying Circulating Breast Cancer Cells in Women With Metastatic Breast Cancer
RATIONALE: Studying samples of blood from patients with metastatic breast cancer in the laboratory may help doctors identify biomarkers related to breast cancer and learn more about how breast cancer begins and spreads in the body.
PURPOSE: This research study is looking at a new way of identifying circulating breast cancer cells in women with metastatic breast cancer.
- To compare identification of circulating breast cancer cells (CBCCs) by a novel technique using stem cell marker retinaldehyde dehydrogenase (ALDH) and surface antigen expression (CD44+, CD24-) to the standard technique using the CellSearch® system in women with metastatic breast cancer.
- To determine whether CBCCs have the potential to grow into metastatic lesions.
OUTLINE: Patients undergo blood sample collection to help develop a new technique using stem cell marker retinaldehyde dehydrogenase (ALDH) and surface antigen expression (CD44+, CD24-) in isolating circulating breast cancer cells (CBCCs). Blood is also drawn to measure the number of circulating tumor cells using the standard CellSearch® system.
Mononuclear cells are isolated by density centrifugation. Cells are stained against surface antigens that provide specific expression patterns for CBCCs (CD44, CD24). Cells are analyzed on a fluorescence activated cell sorting (FACS) Calibur flow cytometer and sequentially gated (ALDHhigh→CD44+ vs CD24-/low or CD44+ vs CD24-/low→ALDHhigh) for detection of CBCCs. For further confirmation of epithelial origin, ALDHhighCD44+CD24-/low cells are isolated using a FACSAria flow sorter, cytocentrifuged onto glass slides then stained for the expression of epithelial-specific cytokeratins 5, 8, 14, 18 and 19 by standard immunohistochemical techniques. Using the phenotype that is found to most highly enrich for epithelial cells, cells are isolated by FACS and assayed for clonogenic growth.
flow cytometry, fluorescence activated cell sorting, immunohistochemistry staining method, immunologic technique, laboratory biomarker analysis
Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins
National Cancer Institute (NCI)
Results (where available)
- Source: http://clinicaltrials.gov/show/NCT00897338
- Information obtained from ClinicalTrials.gov on July 15, 2010
Medical and Biotech [MESH] Definitions
Technique using an instrument system for making, processing, and displaying one or more measurements on individual cells obtained from a cell suspension. Cells are usually stained with one or more fluorescent dyes specific to cell components of interest, e.g., DNA, and fluorescence of each cell is measured as it rapidly transverses the excitation beam (laser or mercury arc lamp). Fluorescence provides a quantitative measure of various biochemical and biophysical properties of the cell, as well as a basis for cell sorting. Other measurable optical parameters include light absorption and light scattering, the latter being applicable to the measurement of cell size, shape, density, granularity, and stain uptake.
A technique encompassing morphometry, densitometry, neural networks, and expert systems that has numerous clinical and research applications and is particularly useful in anatomic pathology for the study of malignant lesions. The most common current application of image cytometry is for DNA analysis, followed by quantitation of immunohistochemical staining.
Laser Scanning Cytometry
A scanning microscope-based, cytofluorimetry technique for making fluorescence measurements and topographic analysis on individual cells. Lasers are used to excite fluorochromes in labeled cellular specimens. Fluorescence is detected in multiple discrete wavelengths and the locational data is processed to quantitatively assess APOPTOSIS; PLOIDIES; cell proliferation; GENE EXPRESSION; PROTEIN TRANSPORT; and other cellular processes.
Cd4 Lymphocyte Count
The number of CD4-POSITIVE T-LYMPHOCYTES per unit volume of BLOOD. Determination requires the use of a fluorescence-activated flow cytometer.
In Situ Hybridization, Fluorescence
A type of IN SITU HYBRIDIZATION in which target sequences are stained with fluorescent dye so their location and size can be determined using fluorescence microscopy. This staining is sufficiently distinct that the hybridization signal can be seen both in metaphase spreads and in interphase nuclei.
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