"Sentinel" circulating tumor cells allow early diagnosis of lung cancer in patients with chronic obstructive pulmonary disease

Abstract

Chronic obstructive pulmonary disease (COPD) is a risk factor for lung cancer. Migration of circulating tumor cells (CTCs) into the blood stream is an early event that occurs during carcinogenesis. We aimed to examine the presence of CTCs in complement to CT-scan in COPD patients without clinically detectable lung cancer as a first step to identify a new marker for early lung cancer diagnosis. The presence of CTCs was examined by an ISET filtration-enrichment technique, for 245 subjects without cancer, including 168 (68.6%) COPD patients, and 77 subjects without COPD (31.4%), including 42 control smokers and 35 non-smoking healthy individuals. CTCs were identified by cytomorphological analysis and characterized by studying their expression of epithelial and mesenchymal markers. COPD patients were monitored annually by low-dose spiral CT. CTCs were detected in 3% of COPD patients (5 out of 168 patients). The annual surveillance of the CTC-positive COPD patients by CT-scan screening detected lung nodules 1 to 4 years after CTC detection, leading to prompt surgical resection and histopathological diagnosis of early-stage lung cancer. Follow-up of the 5 patients by CT-scan and ISET 12 month after surgery showed no tumor recurrence. CTCs detected in COPD patients had a heterogeneous expression of epithelial and mesenchymal markers, which was similar to the corresponding lung tumor phenotype. No CTCs were detected in control smoking and non-smoking healthy individuals. CTCs can be detected in patients with COPD without clinically detectable lung cancer. Monitoring "sentinel" CTC-positive COPD patients may allow early diagnosis of lung cancer.

Figure 1. Cytomorphological and immunocytochemical analysis of Circulating Tumor Cells (CTCs) detected by the ISET technique in patients with COPD.

(A) and (B) CTCs isolated by the ISET method and identified by MGG staining from Patient 1. (A) An isolated CTC with malignant cytomorphological features (Double arrows: pores of the filter). (B) A cluster (CTM) composed of 20 CTCs with malignant cytomorphological features (Original magnification×1000; bars: 8 µm; double arrows: pore containing a lymphocyte). (C) and (D) Immuno-stained CTMs observed in the blood filtered using the ISET method from Patient 2. (C) CTM strongly expressing the pan-cytokeratin antigen only (Double arrows: pore containing a lymphocyte). (D) CTM co-expressing pan-cytokeratin and vimentin antigens [Double arrows: pores of the filter, (Original magnification×400; bars: 16 µm; immuno-peroxidase staining with a pan-cytokeratin antibody (KL1), and an immuno-phosphatase staining with an anti-vimentin antibody)]. (E) A549 epithelial tumor cell line and K562 leukemic cell line having large vimentin aggregates were spiked in human blood, further filtered by ISET, and were used as positive controls for the double immunolabeling assays with KL1 (brown immuno-peroxidase staining, arrows) and with vimentin (reddish immuno-phosphatase staining, double arrows; Original magnification×1000; bars: 40 µm).