2,778 research outputs found

    A lab-on-a-disc platform enables serial monitoring of individual CTCs associated with tumor progression during EGFR-targeted therapy for patients with NSCLC

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    Rationale: Unlike traditional biopsy, liquid biopsy, which is a largely non-invasive diagnostic and monitoring tool, can be performed more frequently to better track tumors and mutations over time and to validate the efficiency of a cancer treatment. Circulating tumor cells (CTCs) are considered promising liquid biopsy biomarkers; however, their use in clinical settings is limited by high costs and a low throughput of standard platforms for CTC enumeration and analysis. In this study, we used a label-free, high-throughput method for CTC isolation directly from whole blood of patients using a standalone, clinical setting-friendly platform. Methods: A CTC-based liquid biopsy approach was used to examine the efficacy of therapy and emergent drug resistance via longitudinal monitoring of CTC counts, DNA mutations, and single-cell-level gene expression in a prospective cohort of 40 patients with epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer. Results: The change ratio of the CTC counts was associated with tumor response, detected by CT scan, while the baseline CTC counts did not show association with progression-free survival or overall survival. We achieved a 100% concordance rate for the detection of EGFR mutation, including emergence of T790M, between tumor tissue and CTCs. More importantly, our data revealed the importance of the analysis of the epithelial/mesenchymal signature of individual pretreatment CTCs to predict drug responsiveness in patients. Conclusion: The fluid-assisted separation technology disc platform enables serial monitoring of CTC counts, DNA mutations, as well as unbiased molecular characterization of individual CTCs associated with tumor progression during targeted therapy

    The rationale for liquid biopsy in colorectal cancer: focus on circulating tumor cells

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    Capturing circulating tumor cells (CTCs) and/or circulating tumor DNA from blood, which represents a precious source of biological material derived from both primary and metastatic tumors, has been named a 'liquid biopsy'. While the circulating tumor DNA might be more representative of the bulk of the metastatic tumor, CTCs are thought to reflect more of the metastases-initiating cells. Consequently, a liquid biopsy made of tumor cells and tumor DNA that is able to track cancer evolution, as a fingerprint of the patient's individual tumor, and is easy to perform at every stage of the disease course, sounds attractive. This article mainly focuses on the applications of CTCs to track tumor dynamics in real time using colorectal cancer as a model system. The analysis of viable CTCs at DNA, RNA and protein levels, as well as their expansion in vitro, may allow deep investigation of the features of metastases-initiating cells

    Circulating tumor cells isolation: The “post-EpCAM era”

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    Circulating tumor cells (CTCs) represent a submicroscopic fraction detached from a primary tumor and in transit to a secondary site. The prognostic significance of CTCs in metastatic cancer patients was demonstrated for the first time more than ten years ago. To date, it seems clear enough that CTCs are highly heterogeneous and dynamically change their shape. Thus, the inadequacy of epithelial cell adhesion molecule (EpCAM) as universal marker for CTCs detection seems unquestionable and alternative methods able to recognize a broader spectrum of phenotypes are definitely needed. In this review the pleiotropic functions of EpCAM are discussed in detail and the role of the molecule in the biology of CTCs is critically dissected

    The potential for liquid biopsies in the precision medical treatment of breast cancer.

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    Currently the clinical management of breast cancer relies on relatively few prognostic/predictive clinical markers (estrogen receptor, progesterone receptor, HER2), based on primary tumor biology. Circulating biomarkers, such as circulating tumor DNA (ctDNA) or circulating tumor cells (CTCs) may enhance our treatment options by focusing on the very cells that are the direct precursors of distant metastatic disease, and probably inherently different than the primary tumor's biology. To shift the current clinical paradigm, assessing tumor biology in real time by molecularly profiling CTCs or ctDNA may serve to discover therapeutic targets, detect minimal residual disease and predict response to treatment. This review serves to elucidate the detection, characterization, and clinical application of CTCs and ctDNA with the goal of precision treatment of breast cancer

    Impact of chronic exposure to bevacizumab on EpCAM-based detection of circulating tumor cells

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    BACKGROUND: Circulating tumor cells (CTCs) are often undetected through the immunomagnetic epithelial cell adhesion molecule (EpCAM)-based CellSearch(®) System in breast and colorectal cancer (CRC) patients treated with bevacizumab (BEV), where low CTC numbers have been reported even in patients with evidence of progression of disease. To date, the reasons for this discrepancy have not been clarified. This study was carried out to investigate the molecular and phenotypic changes in CRC cells after chronic exposure to BEV in vitro. METHODS: The human CRC cell line WiDr was exposed to a clinically relevant dose of BEV for 3 months in vitro. The expression of epithelial and mesenchymal markers and EpCAM isoforms was determined by western blotting and immunofluorescence. To evaluate the impact of EpCAM variant isoforms expression on CTC enumeration by CellSearch(®), untreated and treated colon cancer cells were spiked into 7.5 mL of blood from a healthy donor and enumerated by CellSearch(®). RESULTS: Chronic exposure of CRC cell line to BEV induced decreased expression of EpCAM 40 kDa isoform and increased expression EpCAM 42 kDa isoform, together with a decreased expression of cytokeratins (CK), while no evidence of epithelial to mesenchymal transition (EMT) in treated cells was observed. The recovery rate of cells through CellSearch(®) was gradually reduced in course of treatment with BEV, being 84%, 70% and 40% at 1, 2 and 3 months, respectively. CONCLUSIONS: We hypothesize that BEV may prevent CellSearch(®) from capturing CTCs through altering EpCAM isoforms

    What information could the main actors of liquid biopsy provide? A representative case of non-small cell lung cancer (NSCLC)

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    In non-small cell lung cancer (NSCLC), there is a consensus regarding the use of liquid biopsy, generally, to detect "druggable" mutations and, in particular, to monitor tyrosine kinase inhibitor (TKI) treatments. However, whether circulating tumor cells (CTCs) are better tools than cell-free DNA (cfDNA), is still a matter of debate, mainly concerning which antigen(s) we should use to investigating simultaneously both epithelial and epithelial-to-mesenchymal transient (EMT) phenotype in the same sample of CTCs. To address this item, we exploited here a single-tube liquid biopsy, to detect both epithelial cell adhesion molecule (EpCAM)-positive CTCs and EpCAM-low/negative CTCs, because down-modulation of EpCAM is considered the first step in EMT. Furthermore, we analyzed the DNA from CTCs of four different phenotypes (ctcDNA), according to their EpCAM expression and cytokeratin pattern, and circulating tumor DNA (ctDNA) by droplet digital PCR (ddPCR), in order to disclose activating and resistancedriving mutations. Liquid biopsy reflected spatial and temporal heterogeneity of the tumor under treatment pressure. We provide the proof-of-concept that the complementary use of ctDNA and ctcDNA represents a reliable, minimally invasive and dynamic tool for a more comprehensive view of tumor evolution

    Classification of large circulating tumor cells isolated with ultra-high throughput microfluidic Vortex technology.

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    Circulating tumor cells (CTCs) are emerging as rare but clinically significant non-invasive cellular biomarkers for cancer patient prognosis, treatment selection, and treatment monitoring. Current CTC isolation approaches, such as immunoaffinity, filtration, or size-based techniques, are often limited by throughput, purity, large output volumes, or inability to obtain viable cells for downstream analysis. For all technologies, traditional immunofluorescent staining alone has been employed to distinguish and confirm the presence of isolated CTCs among contaminating blood cells, although cells isolated by size may express vastly different phenotypes. Consequently, CTC definitions have been non-trivial, researcher-dependent, and evolving. Here we describe a complete set of objective criteria, leveraging well-established cytomorphological features of malignancy, by which we identify large CTCs. We apply the criteria to CTCs enriched from stage IV lung and breast cancer patient blood samples using the High Throughput Vortex Chip (Vortex HT), an improved microfluidic technology for the label-free, size-based enrichment and concentration of rare cells. We achieve improved capture efficiency (up to 83%), high speed of processing (8 mL/min of 10x diluted blood, or 800 μL/min of whole blood), and high purity (avg. background of 28.8±23.6 white blood cells per mL of whole blood). We show markedly improved performance of CTC capture (84% positive test rate) in comparison to previous Vortex designs and the current FDA-approved gold standard CellSearch assay. The results demonstrate the ability to quickly collect viable and pure populations of abnormal large circulating cells unbiased by molecular characteristics, which helps uncover further heterogeneity in these cells

    Liquid Biopsy in Rare Cancers: Lessons from Hemangiopericytoma

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    Hemangiopericytoma (HPT) is a rare mesenchymal tumor of fibroblastic type and for its rarity is poorly studied. The most common sites of metastatic disease in patients with intracranial HPT are the bone, liver, and lung, suggestive for an hematogenous dissemination; for this reason, we investigated, for the first time, the presence of circulating tumor cells (CTCs) in hemangiopericytoma patient by CellSearch® and SceenCell® devices. Peripheral blood samples were drawn and processed by CellSearch, an EpCAM-dependent device, and ScreenCell®, a device size based. We found nontypical CTCs by CellSearch system and the immunofluorescence analysis performed on CTCs isolate by ScreenCell demonstrated the presence of single CTCs and CTC clusters. The molecular characterization of single CTCs and CTC clusters, using antibodies directed against EpCAM, CD34, cytokeratins (8, 18, and 19), and CD45, showed a great heterogeneity in CTC clusters. We believe that the present study may open a new scenario in the rare tumors: the introduction of the liquid biopsy and the molecular characterization of circulating tumor cells could lead to personalized targeted treatments and also for rare tumors
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