Comparison of RNA Marker Panels for Circulating Tumor Cells and Evaluation of Their Prognostic Relevance in Breast Cancer

SIMPLE SUMMARY: Circulating tumor cells are precursors of distant metastasis in many cancer types. These cells circulate in the peripheral blood, which can be analyzed in a non-invasive procedure to identify patients at risk to develop metastases. In order to improve non-invasive diagnostic procedures for breast cancer, we aimed at investigating gene transcripts as liquid biopsy markers in the blood employing density gradient centrifugation as a cost- and time-saving method for the isolation of the target cells. We were able to detect the selected biomarkers in 86.1% of metastatic and 31.1% of early breast cancer patients. The presence of some markers were significantly related with shorter survival of the patients. Our data suggest that these transcripts have the potential to identify patients with poor prognosis who might benefit from further clinical intervention. ABSTRACT: Liquid biopsy is a promising tool for therapy monitoring of cancer patients, but a need for further research in this field exists in order to improve sensitivity, specificity, standardization and minimize costs. In our present study, we evaluated two panels of transcripts related with the presence of circulating tumor cells (CTCs) (Panel 1: CK19, EpCAM, SCGB2A2 and Panel 2: EMP2, SLC6A8, HJURP, MAL2, PPIC and CCNE2) in two cohorts of breast cancer patients (metastatic and early). A blood cell fraction possibly containing CTCs was isolated with density gradient centrifugation, followed by RNA isolation and qPCR using TaqMan(®) or RT-qPCR using hybridization probes. The positivity rates of the investigated panels were similar, albeit higher in metastatic (69.4% Panel 1, 75.0% Panel 2; total 86.1%) compared to early (18.9% Panel 1, 23.3% Panel 2; total 31.1%) breast cancer patients. CK19, SCGB2A2, EMP2, HJURP, MAL2, and CCNE2 individually correlated with shorter overall survival in the metastatic patient cohort. The findings highlight the additional value of Panel 2 markers, which are in contrast to CK19 and EpCAM not solely linked to an epithelial phenotype.