Fluid biopsy for circulating tumor cell identification in patients with early-and late-stage non-small cell lung cancer: a glimpse into lung cancer biology

Abstract

Circulating tumor cell (CTC) counts are an established prognostic marker in metastatic prostate, breast, and colorectal cancer, and recent data suggests a similar role in late stage non-small cell lung cancer (NSCLC). However, due to sensitivity constraints in current enrichment-based CTC detection technologies, there is little published data about CTC prevalence rates and morphologic heterogeneity in early stage NSCLC, or the correlation of CTCs with disease progression and their usability for clinical staging. We investigated CTC counts, morphology, and aggregation in early stage, locally advanced, and metastatic NSCLC patients by using a fluid phase biopsy approach that identifies CTCs without relying on surface receptor-based enrichment and presents them in sufficiently high definition (HD) to satisfy diagnostic pathology image quality requirements. HD-CTCs were analyzed in blood samples from 78 chemotherapy-naïve NSCLC patients. 73% of the total population had a positive HD-CTC count (> 0 CTC in 1 mL of blood) with a median of 4.4 HD-CTCs/mL (range 0–515.6) and a mean of 44.7 (±95.2) HD-CTCs/mL. No significant difference in the medians of HD-CTC counts was detected between stage IV (n=31, range 0–178.2), stage III (n=34, range 0–515.6) and stages I/II (n=13, range 0–442.3). Furthermore, HD-CTCs exhibited a uniformity in terms of molecular and physical characteristics such as fluorescent cytokeratin intensity, nuclear size, frequency of apoptosis and aggregate formation across the spectrum of staging. Our results demonstrate that, despite stringent morphologic inclusion criteria for the definition of HD-CTCs, the HD-CTC assay shows high sensitivity in the detection and characterization of both early and late stage lung cancer CTCs. Larger studies are warranted to investigate the prognostic value of CTC profiling in early stage lung cancer. This finding has implications for the design of larger studies examining screening, therapy, and surveillance in lung cancer patients.

Biophysical technologies for understanding circulating tumor cell biology and metastasis

“…There was a high HD-CTC detection rate of 73% across the whole population, with a median of 4.4 HD-CTCs per 1 mL blood and a mean of 44.1 HDCTCs per 1 mL blood (40). Notably, HD-CTCs were detected in 85% of patients with stage I and II disease and 79% in stage III (40). When examined against FDG-PET imaging in NSCLC, HD-CTC number did not correlate with tumor diameter, and increasing HD-CTC counts only weakly correlated with increasing SUV values.…”

“…In comparison to CellSearch, the HD-CTC assay found significantly higher numbers of CTCs in significantly more patients with metastatic breast, prostate, and lung cancer (19,41). Furthermore, tumor staging did not seem to make a difference in the detection of CTCs in NSCLC (40). The major disadvantage with the HD-CTC assay, however, is the inability to analyze live cells due to the fixing of cells during processing (13).…”

“…Cells need to meet specific criteria to be classified as HD-CTC, namely to have the following features: an intact DAPI nucleus without identifiable apoptotic changes or a disrupted appearance, express cytokeratin, lack expression of CD45, and be morphologically distinct from surrounding nonmalignant leukocytes (19,40). The advance of this platform is that images not meeting HD-CTC inclusion criteria were not discarded, but rather, were retained and digitally catalogued for subsequent re-analysis of HD-CTCs (40). This retention of all cells is touted by Epic Sciences, the primary licensee of this technology, as having the ability to maintain a "no cell left behind" approach.…”

Tumor type-specific and skin region-selective metastasis of human cancers: another example of the “seed and soil” hypothesis

“…Importantly, they can be detected with a relatively high incidence even in early stage disease [34]. Reports on the comparison of early and late stage disease CTC numbers found limited correlation [36,37]. With advances of cell isolation technologies, the detection threshold has become lower, reaching 1-5 CTC/10 ml blood [32].…”

Diagnostics: Playing detective

“…The team recently showed that their technique could detect circulating tumour cells in blood samples from patients with nonsmall-cell lung cancer, even in the early stages of disease 1 .…”