Abstract
Size selection via filtration offers an antigen-independent approach for the enrichment of rare cell populations in blood of cancer patients. We evaluated the performance of a novel approach for multiplex rare cell detection in blood samples from metastatic breast (n = 19) and lung cancer patients (n = 21), and healthy controls (n = 30) using an automated microfluidic filtration and multiplex immunoassay strategy. Captured cells were enumerated after sequential staining for specific markers to identify circulating tumor cells (CTCs), circulating mesenchymal cells (CMCs), putative circulating stem cells (CSCs), and circulating endothelial cells (CECs). Preclinical validation experiments using cancer cells spiked into healthy blood demonstrated high recovery rate (mean = 85%) and reproducibility of the assay. In clinical studies, CTCs and CMCs were detected in 35% and 58% of cancer patients, respectively, and were largely absent from healthy controls (3%, p = 0.001). Mean levels of CTCs were significantly higher in breast than in lung cancer patients (p = 0.03). Fifty-three percent (53%) of cancer patients harbored putative CSCs, while none were detectable in healthy controls (p<0.0001). In contrast, CECs were observed in both cancer and control groups. Direct comparison of CellSearch® vs. our microfluidic filter method revealed moderate correlation (R2 = 0.46, kappa = 0.47). Serial blood analysis in breast cancer patients demonstrated the feasibility of monitoring circulating rare cell populations over time. Simultaneous assessment of CTCs, CMCs, CSCs and CECs may provide new tools to study mechanisms of disease progression and treatment response/resistance.
Highlights
Recent technological advances have enabled the reliable detection and characterization of circulating tumor cells (CTCs) in the blood of cancer patients [1, 2]
We evaluated the performance of a novel approach for detection and enumeration of multiple rare cell populations in the blood of metastatic breast and lung cancer patients using an automated microfluidic filtration and multiplex immunoassay strategy
We evaluated the clinical performance of our assay in detecting and enumerating CTCs, circulating mesenchymal cells (CMCs), circulating stem cells (CSCs), and circulating endothelial cells (CECs) in the blood of metastatic breast and lung cancer patients
Summary
Recent technological advances have enabled the reliable detection and characterization of circulating tumor cells (CTCs) in the blood of cancer patients [1, 2]. To quantify levels of CTCs, assays have been developed to facilitate the detection of epithelial cells in the blood by using cellular markers such as EPCAM and cytokeratins [3]. Recent studies have identified CTCs with low expression of epithelial markers, e.g., those undergoing epithelial-mesenchymal transition (EMT), which may not be detected by the epithelial-based assays [8, 9]. Filter-based methods facilitate the enrichment of non-hematologic rare cells (including CTCs) by exploiting the size differences between these cells and cells in peripheral blood [10,11,12,13,14]. Cells that are captured on the filter can be subjected to further assay for detection and enumeration of CTCs [17,18,19]
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