Sensitive and direct detection of circulating tumor cells by multimarker µ-nuclear magnetic resonance - PubMed (original) (raw)

Sensitive and direct detection of circulating tumor cells by multimarker µ-nuclear magnetic resonance

Arezou A Ghazani et al. Neoplasia. 2012 May.

Abstract

Identifying circulating tumor cells (CTCs) with greater sensitivity could facilitate early detection of cancer and rapid assessment of treatment response. Most current technologies use EpCAM expression as a CTC identifier. However, given that a significant fraction of cancer patients have low or even absent EpCAM levels, there is a need for better detection methods. Here, we hypothesize that a multimarker strategy combined with direct sensing of CTC in whole blood would increase the detection of CTC in patients. Accordingly, molecular profiling of biopsies from a patient cohort revealed a four-marker set (EpCAM, HER-2, EGFR, and MUC-1) capable of effectively differentiating cancer cells from normal host cells. Using a point-of-care micro-nuclear magnetic resonance (µNMR) system, we consequently show that this multimarker combination readily detects individual CTC directly in whole blood without the need for primary purification. We also confirm these results in a comparative trial of patients with ovarian cancer. This platform could potentially benefit a broad range of applications in clinical oncology.

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Figures

Figure 1

Clinical rationale and quad-_µ_NMR schematic. (A) Biomarker expression, as assessed by _µ_NMR, showed that a significant percentage of cancer patients (n = 58) were negative for EpCAM (34.4%), HER-2 (32.7%), MUC-1 (32.7%), or EGFR (31.0%). Combining these markers, however, enabled identification of nearly all cancer patients (99.2%). (B) Schematic of the quad-_µ_NMR system. TCO-labeled antibodies are added to whole blood. RBC are then lysed and the cells spun down before reaction with Tz-containing magnetic nanoparticles. The process of labeling antibodies and targeting nanoparticles requires less than 30 minutes. Biomarker measurements are then taken using the _µ_NMR device (shown).

Figure 2

Comparison of single and quadmarker detection using _µ_NMR. Quad-_µ_NMR CTC targeting (red bars) in whole blood showed higher NMR signals than single-marker CTC targeting (bluebars). For comparative purposes, the _µ_NMR values are displayed as relative ratios. All measurements were obtained intriplicate (note narrow error bars). In all cell lines assessed, quad-_µ_NMR consistently out-performed all single-_µ_NMR detections. Most notably, quad-_µ_NMR even outperformed EpCAM-_µ_NMR in high- (A), intermediate- (B), and low/negative- (C) EpCAM-expressing cell lines.

Figure 3

Detection sensitivity of quad-_µ_NMR in whole blood compared with the clinical standard. Varying numbers of cancer cells (i.e., 200, 100, 50, and 25 cells) were spiked separately into 7-ml samples of blood from a healthy donor. Quad-_µ_NMR measurements are shown as mean values. Quad-_µ_NMR was able to detect 86/200 spiked cells, 41/100 spiked cells, 18/50 spiked cells, and 8/25 spiked cells. All _µ_NMR measurements were done intriplicate for both test and control samples. (A) Quad-_µ_NMR outperformed single marker EpCAM-_µ_NMR for all cell concentrations assessed. (B) Observed values are plotted against theoretical values (grayline).

Figure 4

Comparison of quad-_µ_NMR to the clinical standard. (A) Using quad-_µ_NMR detection, the average recovery rate was 38% across the various cell concentrations assessed (i.e., 200, 100, 50 and 25 spiked cells). Identical experiments performed using the CellSearch detection system showed an average recovery rate of just 9.1% across all concentrations, ranging from 14% at the 200-cell spike (∼30 cells/ml) to 4.3% at the 25-cell spike (∼3 cells/ml, i.e., below the five-cell detection threshold recommended by CellSearch). (B) Compared to the clinically used CellSearch system, quad-_µ_NMR showed higher CTC detection sensitivity (P < .05) for all cell concentrations assessed.

Figure 5

Detection of CTC levels in patient samples using quad-_µ_NMR. (A) CTC levels in the peripheral blood of 15 advanced-stage (IIIC or IV) ovarian cancer patients compared with those of 10 healthy individuals, as assessed by quad-_µ_NMR. (B) A comparison of CTC levels in samples determined by either quad-_µ_NMR or CellSearch. A positive result was obtained for 13 of 15 cancer patients using quad-NMR, whereas only 1 of 15 patients (patient 13) was clearly positive with CellSearch. The dynamic range of CTC levels obtained using quad-_µ_NMR was 0 to 170.

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Sensitive and direct detection of circulating tumor cells by multimarker µ-nuclear magnetic resonance - PubMed (original) (raw)