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Conventional strategy of anti-EpCAM capture and immunostaining of cytokeratins (CKs) to detect circulating tumor cells (CTCs) is limited by highly heterogeneous and dynamic expression or absence of EpCAM and/or CKs in CTCs. In this study, a novel integrated cellular and molecular approach of subtraction enrichment (SE) and immunostaining-FISH (iFISH) was successfully developed. Both large or small size CTCs and circulating tumor microemboli (CTM) in various biofluid samples including cerebrospinal fluid (CSF) of cancer patients and patient-derived-xenograft (PDX) mouse models were efficiently enriched and comprehensively identified and characterized by SE-iFISH. Non-hematopoietic CTCs with heteroploid chromosome 8 were detected in 87-92% of lung, esophageal and gastric cancer patients. Characterization of CTCs performed by CK18-iFISH showed that CK18, the dual epithelial marker and tumor biomarker, was strong positive in only 14% of lung and 24% of esophageal CTCs, respectively. Unlike conventional methodologies restricted only to the large and/or both EpCAM and CK positive CTCs, SE-iFISH enables efficient enrichment and performing in situ phenotypic and karyotypic identification and characterization of the highly heterogeneous CTC subtypes classified by both chromosome ploidy and the expression of various tumor biomarkers. Each CTC subtype may possess distinct clinical significance relative to tumor metastasis, relapse, therapeutic drug sensitivity or resistance, etc.

oncotarget

Enhanced detection and comprehensive<i>in situ</i>phenotypic characterization of circulating and disseminated heteroploid epithelial and glioma tumor cells

Enhanced detection and comprehensivein situphenotypic characterization of circulating and disseminated heteroploid epithelial and glioma tumor cells