Abstract
BackgroundThe molecular profiles of tumors may inform the selection of appropriate targeted therapies. Circulating tumor cells (CTCs) reflect the real-time status of tumor genotypes. CTCs exhibit high genetic heterogeneity within a patient; accordingly, the analysis of individual CTCs, including their heterogeneity, may enable more precise treatments. We analyzed KRAS mutations in single CTCs from patients with metastatic colorectal cancer (mCRC) using a new single-cell picking system.MethodsBlood samples were obtained from 61 patients with mCRC. CTCs were enriched and fluorescently labeled using the CellSearch® System. They were recovered using the single-cell picking system based on the fluorescence intensity of marker dyes. Single CTCs and tumor tissue samples were examined for mutations in codons 12 and 13 of the KRAS gene.ResultsCTCs were detected in 27 of 61 patients with mCRC. We isolated at least two CTCs from 15 of 27 patients. KRAS genotype was evaluated in a total of 284 CTCs from 11 patients, and 15 cells with mutations were identified in four patients. In 10 of 11 patients, the KRAS status was the same in the primary tumor and CTCs. In one patient, the KRAS status was discordant between the primary tumor and CTCs. In two patients, different KRAS mutations were found among individual CTCs.ConclusionsWe successfully isolated single CTCs and detected KRAS mutations in individual cells from clinical samples using a novel application of single-cell isolation system. Using the system, we detected CTC heterozygosity and heterogeneity in KRAS status among CTCs within a patient and between CTCs and tumor tissues.
Highlights
The molecular profiles of tumors may inform the selection of appropriate targeted therapies
Evaluation of single-cell collection using the single-cell picking system To quantify the rate of tumor cell recovery using the single-cell picking system, fluorescently labeled H1975 cells were loaded onto the single-cell picking system and collected individually in wells of a 96-well microplate
In this study, we evaluated the feasibility of detecting KRAS mutations in single Circulating tumor cell (CTC) isolated from metastatic colorectal cancer (mCRC) patients using the ASONECell Picking System
Summary
The molecular profiles of tumors may inform the selection of appropriate targeted therapies. We analyzed KRAS mutations in single CTCs from patients with metastatic colorectal cancer (mCRC) using a new single-cell picking system. The use of new antitumor agents for metastatic CRC (mCRC), such as epidermal growth factor receptor-targeted monoclonal antibodies (anti-EGFR), has significantly improved the treatment of colorectal disease [1, 2]. KRAS mutations are present in 30–40% of CRC patients [3]. Activating mutations in KRAS are responsible for antiEGFR therapy resistance in mCRC; KRAS. Acquired resistance is partly achieved by the selection of preexisting minor subclones harboring mutations that confer resistance to targeted therapy [7, 8].
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