Sensitive cytometry based system for enumeration, capture and analysis of gene mutations of circulating tumor cells.
Bottom Line: We compared the number of CTC detected using our system with those detected using the CellSearch system in 46 patients with various cancers.The percentage of samples with detected CTC was significantly higher with our system (65.2%) than with CellSearch (28.3%).The number of detected CTC per patient using our system was statistically higher than that using CellSearch (median 5, 0; P = 0.000172, Wilcoxon test).
Affiliation: Shien-Lab, National Cancer Center Hospital, Tokyo, Japan.Show MeSH
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Mentions: PC‐9 cells (5, 20 or 100 cells) and BT‐20 cells (10, 25, 50 or 100 cells) were spiked into 4 mL of peripheral blood, and the spiked cells were then recovered from the blood using On‐chip Sort. Recovered cells were visually distinguished from leukocytes (Fig. 4a) and were then counted under a fluorescent microscope. Regression analysis of the number of captured cells versus the number of spiked cells produced a correlation coefficient (R2) of 0.9029 (Fig. 4b). The mean capture rates were 101.35 ± 15.3% for PC‐9 and 77.0 ± 8.1% for BT‐20 (Fig. 4c). To estimate the minimal number of CTC that are required for detection of mutations using our method, we performed further spike‐in experiments. PC‐9 cells (5, 10 or 100 cells) and H1650 cells (5, 10 or 30 cells), which have the EGFR exon 19 deletion mutation, and BT20 cells (10, 20 or 50 cells) that express the PIK3CA variant H1047R mutation were spiked into 4 mL of peripheral blood. Following processing of each sample in a manner similar to that described for CTC enrichment (Fig. 2), the spiked cells were sorted from the peripheral blood using On‐chip Sort, which successfully sorted spiked cells from all samples. EGFR deletion mutation was detected in samples spiked with 5, 10 or 100 PC‐9 cells and 5, 10 or 30 H1650 cells using the method described in the Materials and Methods. The PIK3CA variant H1047R mutation was detected in samples spiked with 20 or 50 BT20 cells. The same results were obtained twice in independent experiments.
Affiliation: Shien-Lab, National Cancer Center Hospital, Tokyo, Japan.