Tumor cells are dislodged into the pulmonary vein during lobectomy

Abstract

Objective Intraoperative tumor shedding may facilitate tumor dissemination. Earlier studies defined shed tumor cells primarily by cytomorphological examination, and could not always distinguish normal epithelial cells from tumor cells. We sought to accurately identify tumor cells using single-cell sequencing and determine whether these cells were mobilized into the circulation during pulmonary lobectomy. Methods We analyzed 42 blood samples collected from the tumor-draining pulmonary vein at the end of lobectomy procedures. We used arrays of nanowells to enumerate and retrieve single EpCAM+ cells. Targeted sequencing of 10–15 cells and nested PCR of single cells detected somatic mutations in shed epithelial cells consistent with patient-matched tumor but not normal tissue. Results The mean of EpCAM+ cells in VATS lobectomy (no wedge) specimens (n=16) was 165 (median 115; range 0–509) but sampling cells from three patients indicated that only 0–38% of the EpCAM+ cells were tumor cells. The mean of VATS lobectomy (wedge) specimens (n=12) was 1128 (median 197; range 47– 9406) and all of the EpCAM+ cells were normal epithelial cells in two patients sampled. The mean of EpCAM+ cells in thoracotomy specimens (n=14) was 238 (median 22; range 9–2920) and 0–50% of total EpCAM+ cells were tumor cells based on four patients sampled. Conclusion Surgery mobilizes tumor cells into the pulmonary vein, along with many normal epithelial cells. EpCAM alone cannot differentiate between normal and tumor cells. On the other hand, single-cell genetic approaches with patient-matched normal and tumor tissue can accurately quantify the number of shed tumor cells.

Prevalence and number of circulating tumour cells and microemboli at diagnosis of advanced NSCLC

“…Manipulation of tumour in animal models (Juratli et al 2014) and in human lung cancer (Sawabata et al 2007;Yao et al 2014) is associated with delivery of CTC in the bloodstream. Since the same phenomenon might occur in lung tumours undergoing FNAB, in each patient 15 mL of blood was collected in two EDTA tubes (7.5 mL each) soon before CT-FNAB through the antecubital vein access routinely prepared before CT-FNAB.…”

Lung cancer biopsy dislodges tumor cells into circulating blood

“…These results suggest that a tumor biopsy of a lung cancer lesion has the potential to dislodge tumor cells into the peripheral circulating blood, as can occur with a surgical procedure. [12][13][14][15] Initially, tumor cell dislodgement to the peripheral circulating blood by surgical manipulation was demonstrated using a cytological technique, but the sensitivity was very low. [17] Later, polymerase chain reaction [18,19] and flow-cytometry [20] methods were introduced to improve sensitivity and specificity, but results were not predictable.…”

“…Morphological detection of CTC dislodgement was shown using the EpCAM positive selection method [CellSearch ® system (Janssen Diagnostics, Raritan, NJ)], but the sensitivity was still low. [15] As such, isolated tumor cells (ITC), surrogates of CTC, were extracted from pulmonary vein blood, revealing that detecting ITC/CTC was an indicator of early recurrence; the presence of cluster ITC/CTC indicated wrong prognosis, using the EpCAM positive selection method [21] or the CD45 negativedepression gravity method [RosettSep ® (Stemcell Technologies, Vancouver, Canada)]. [22][23][24] As the sensitivity of such CTC detecting methods was not greater, the sample used pulmonary vein blood, because it contains more ITC/CTC than the peripheral circulating blood.…”

“…[12][13][14][15] Above all, detecting cluster circulating tumor cell (CTC) is speculated the best a strong predictor of early recurrence. [13] FBB also manipulates the area of lung cancer during biopsy; thus, CTCs might be dislodged from the lesion to the circulating blood as the same manner as surgical manipulation.…”

Automated rare single cell picking with the ALS cellcelector™

“…The system is fully agnostic regarding upstream enrichment methods handling large numbers of contaminating cells (e.g., 50,000 per sample) and does not require sample volume reduction. CellCelector™ has been successfully used downstream of various CTC enrichment techniques: (i) positive immune‐magnetic: CellSearch® , Isoflux™ , Illumina MagSweeper™ ; (ii) negative immune‐magnetic: Dynal® MPC‐S , RosetteSep CD45 ; (iii) density gradient centrifugation ; or (iv) size‐based separation: Parsortix™ .…”