Circulating Tumor Cell Clusters Are Frequently Detected in Women with Early-Stage Breast Cancer

Abstract

The clinical relevance of circulating tumor cell clusters (CTC-clusters) in breast cancer (BC) has been mostly studied using the CellSearch®, a marker-dependent method detecting only epithelial-enriched clusters. However, due to epithelial-to-mesenchymal transition, resorting to marker-independent approaches can improve CTC-cluster detection. Blood samples collected from healthy donors and spiked-in with tumor mammospheres, or from BC patients, were processed for CTC-cluster detection with 3 technologies: CellSearch®, CellSieve™ filters, and ScreenCell® filters. In spiked-in samples, the 3 technologies showed similar recovery capability, whereas, in 19 clinical samples processed in parallel with CellSearch® and CellSieve™ filters, filtration allowed us to detect more CTC-clusters than CellSearch® (median number = 7 versus 1, p = 0.0038). Next, samples from 37 early BC (EBC) and 23 metastatic BC (MBC) patients were processed using ScreenCell® filters for attaining both unbiased enrichment and marker-independent identification (based on cytomorphological criteria). At baseline, CTC-clusters were detected in 70% of EBC cases and in 20% of MBC patients (median number = 2, range 0–20, versus 0, range 0–15, p = 0.0015). Marker-independent approaches for CTC-cluster assessment improve detection and show that CTC-clusters are more frequent in EBC than in MBC patients, a novel finding suggesting that dissemination of CTC-clusters is an early event in BC natural history.

Better together: circulating tumor cell clustering in metastatic cancer

“…Moreover, the detection of CTC clusters in patients with metastatic cancer showed an additional prognostic value confirming the increased aggressiveness of clusters compared with single CTCs observed in mouse models [ 3 , 14 – 17 ]. Studies on the clinical significance of CTC clusters in early-stage breast cancer are indeed lacking, but recent studies showing the detection of CTC clusters in patients with nonmetastatic breast cancer open a new opportunity to investigate the role of CTC clusters in the early steps of tumor dissemination and further emphasize the need to understand the molecular drivers across all subtypes and stages of breast cancer [ 18 – 20 ].…”

“…The detection of clusters in early-stage patients by using blood filtration was also recently reported. CTC clusters were detected in 26 of 37 patients before starting neoadjuvant treatment [ 20 ], and the malignancy of clusters detected by filtration before neoadjuvant chemotherapy was confirmed for 46 of 48 clusters isolated from six patients by single-cluster isolation and genomic profiling [ 18 ]. These results further support the ability of filtration techniques to detect CTC clusters, as already reported in metastatic breast cancer [ 32 ] and other malignancies at different disease stages [ 24 , 26 , 31 – 35 ].…”

“…Size exclusion might in fact be the best strategy to enrich CTC clusters, considering that they are considerably larger than white blood cells (WBCs). Indeed, by direct comparison of filtration and the CellSearch platform with clinical samples, two studies demonstrated increased CTC cluster detection attained by using filters [ 20 , 24 ]. However, on the one hand, some concerns derive from the shear stress during filtration, which might damage CTC clusters, and from the presence of clusters of normal cells in the circulation, which could reduce the specificity of this approach if not coupled with additional phenotypic and/or genotypic analyses.…”

Heterogeneity of Circulating Tumor Cell Neoplastic Subpopulations Outlined by Single-Cell Transcriptomics

“…Extensive data obtained from clinical and preclinical models and cancer types have demonstrated the relevance of blood-borne cancer dissemination, and asserted that cell dissemination occurs early during tumor development [ 8 , 9 ]. Notably, circulating tumor cells (CTCs) directly captured from the peripheral blood of patients have emerged as a powerful biomarker for monitoring cancer progression [ 7 , 10 , 11 , 12 ]. Because CTCs express distinct and specific transcriptional profiles compared to primary or metastatic tumors, they offer the unique opportunity to analyze transcriptional information, which can be therapeutically useful [ 7 , 13 , 14 , 15 ].…”

“…This definition has been applied by the US Food and Drug Administration (FDA)-cleared CellSearch TM platform for the capture and enumeration of CTCs in clinical settings [ 16 , 17 , 18 ]. Quantification of CTCs has proven to be an independent prognostic indicator of progression-free/overall survival of metastatic cancer patients, including mBC [ 6 , 10 , 12 ]. Importantly, recent studies have identified new populations of breast cancer CTCs, which display phenotypic and gene expression profiles, which are distinct from “classic” CTCs [ 19 , 20 ].…”