Biphasic co-detection of melanoma aneuploid tumor cells and tumor endothelial cells in guidance of specifying the field cancerized surgical excision margin and administering immunotherapy.

An optimum safety excision margin (EM) delineated by precise demarcation of field cancerization along with reliable biomarkers that enable predicting and timely evaluating patients' response to immunotherapy significantly impact effective management of melanoma. In this study, optimized biphasic "immunofluorescence staining integrated with fluorescence insitu hybridization" (iFISH) was conducted along the diagnosis-metastasis-treatment-cellular MRD axis to longitudinally co-detect a full spectrum of intact CD31 aneuploid tumor cells (TCs), CD31 aneuploid tumor endothelial cells (TECs), viable and necrotic circulating TCs (CTCs) and circulating TECs (CTECs) expressing PD-L1, Ki67, p16 and Vimentin in unsliced specimens of the resected primary tumor, EM, dissected sentinel lymph nodes (SLNs) and peripheral blood in an early-stage melanoma patient. Numerous PD-L1 aneuploid TCs and TECs were detected at the conventional safety EM (2 cm), quantitatively indicating the existence of a field cancerized EM for the first time.

Case report: Post-therapeutic laryngeal carcinoma patient possessing a high ratio of aneuploid CTECs to CTCs rapidly developed malignancy in pancreas.

Effectively evaluating therapeutic efficacy, detecting minimal residual disease (MRD) after therapy completion, and predicting early occurrence of malignancy in cancer patients remain as unmet imperative clinical demands. This article presents a case of a laryngeal carcinoma patient who had a surgical resection and complete post-operative chemoradiotherapy in combination with the targeted therapy, then rapidly developed pancreatic adenocarcinoma. Detected by SE-iFISH, the patient had a substantial amount of 107 non-hematological aneuploid circulating rare cells including 14 circulating tumor cells (CTCs, CD31/CD45) and 93 circulating tumor endothelial cells (CTECs, CD31/CD45) with a high ratio of CTECs/CTCs > 5 upon finishing post-surgical combination regimens.

Nonhematogenic circulating aneuploid cells confer inferior prognosis and therapeutic resistance in gliomas.

Aneuploidy is the hallmark of malignancy. Our previous study successfully detected nonhematogenic circulating aneuploidy cells (CACs) in types of gliomas. The current prospective clinical study aims to further precisely subcategorize aneuploid CACs, including CD31 circulating tumor cells (CTCs) and CD31 circulating tumor endothelial cells, and thoroughly investigate the clinical utilities of these different subtypes of cells.

Analysis of the prognostic role and biological characteristics of circulating tumor cell-associated white blood cell clusters in non-small cell lung cancer.

Background: Recently, circulating tumor-cell-associated white blood cell (CTC-WBC) clusters have been reported to have prognostic value in some cancers. The prognostic role of CTC-WBC clusters in lung cancer has not yet been elucidated. Very little information is available about the biological characteristics of CTC-WBC clusters.

Aneuploid Circulating Tumor Cells as a Predictor of Response to Neoadjuvant Chemotherapy in Non-Small Cell Lung Cancer.

Purpose: This study aimed to explore the potential application of circulating tumor cells (CTCs) in predicting the therapeutic effect of neoadjuvant chemotherapy (NAC) in non-small-cell lung cancer (NSCLC).

Methods: Using integrated subtraction enrichment and immunostaining-fluorescence in situ hybridization, the serial CTCs of patients with NSCLC were detected in 7.5 mL of blood at baseline and after two cycles of cisplatin-based NAC, and all aneuploidies of chromosome 8 were examined in the enriched CTCs.

Identification and Comprehensive Co-Detection of Necrotic and Viable Aneuploid Cancer Cells in Peripheral Blood.

Aneuploid circulating tumor cells (CTCs, CD31) and circulating tumor endothelial cells (CTECs, CD31) exhibit an active interplay in peripheral blood, and play an essential role in tumorigenesis, neoangiogenesis, disease progression, therapy-resistant minimal residual disease (MRD), cancer metastasis and relapse. Currently, most CTC detection techniques are restricted to the indistinguishable quantification of circulating rare cells, including both necrotic and viable cells in cancer patients. Clinically imperative demands to distinguish and detect live and/or dead non-hematological aneuploid cancer cells in peripheral blood, which will assist in the rapid evaluation of therapeutic effects, real-time monitoring of treatment resistance longitudinally developed along with therapy and the effective detection of post-therapeutic MRD, have not yet been achieved.

Role of aneuploid circulating tumor cells and CD31 circulating tumor endothelial cells in predicting and monitoring anti-angiogenic therapy efficacy in advanced NSCLC.

Prognosticating the efficacy of anti-angiogenic therapy through longitudinal monitoring and early detection of treatment resistance in cancer patients remain highly challenging. In this study, co-detection and comprehensive phenotypic and karyotypic molecular characterization of aneuploid circulating tumor cells (CTCs) and circulating tumor endothelial cells (CTECs) were conducted on non-small cell lung cancer (NSCLC) patients receiving bevacizumab plus chemotherapy. Prognostic values of the cell-based significant univariate risk factors identified by Cox regression analyses were progressively investigated.

Dysregulated KRAS gene-signaling axis and abnormal chromatin remodeling drive therapeutic resistance in heterogeneous-sized circulating tumor cells in gastric cancer patients.

The mechanism by which heterogeneous-sized circulating tumor cells (CTCs) in gastric cancer (GC) patients are resistant to the targeted therapy and/or chemotherapy remains unclear. This study investigated prognostic value and genomic variations of size-heterogenous CTCs, in an attempt to unravel the molecular mechanisms underlying the therapeutic resistance, which is relevant to poor prognosis in GC. Aneuploid CTCs, detected in 111 advanced GC patients, were categorized into small (≤white blood cell [WBC], 25.

PD-L1 aneuploid circulating tumor endothelial cells (CTECs) exhibit resistance to the checkpoint blockade immunotherapy in advanced NSCLC patients.

Sustained angiogenesis and increased PD-L1 expression on endothelial and carcinoma cells contribute toward fostering an immunosuppressive microenvironment suitable for tumor growth. PD-L1 CTCs were reported to associate with poor prognosis in NSCLC patients. However, whether or not aneuploid circulating tumor endothelial cells (CTECs) express PD-L1, then serve as a surrogate biomarker to evaluate immunotherapy efficacy remains unknown.

Epithelial-type systemic breast carcinoma cells with a restricted mesenchymal transition are a major source of metastasis.

Carcinoma cells undergo epithelial-mesenchymal transition (EMT); however, contributions of EMT heterogeneity to disease progression remain a matter of debate. Here, we addressed the EMT status of ex vivo cultured circulating and disseminated tumor cells (CTCs/DTCs) in a syngeneic mouse model of metastatic breast cancer (MBC). Epithelial-type CTCs with a restricted mesenchymal transition had the strongest lung metastases formation ability, whereas mesenchymal-type CTCs showed limited metastatic ability.

Evolutionary Expression of HER2 Conferred by Chromosome Aneuploidy on Circulating Gastric Cancer Cells Contributes to Developing Targeted and Chemotherapeutic Resistance.

Previous human epidermal growth factor receptor-2 (HER2)-derived resistance studies were based on models, which could not mirror evolutionary expression of HER2 during therapy. To investigate dynamic expression of HER2 and its contribution to developing therapeutic resistance conferred by chromosome aneuploidy, both the HER2 phenotype and chromosome 8 (Chr 8) aneuploidy on circulating tumor cells (CTC) were coexamined in advanced gastric cancer (AGC) patients. A total of 115 AGC patients, including 56 of histopathologic HER2 (hHER2) subjects who received first-line HER2-targeted therapy plus chemotherapy, and 59 of hHER2 patients who received chemotherapy alone, were prospectively enrolled.

Quantified postsurgical small cell size CTCs and EpCAM circulating tumor stem cells with cytogenetic abnormalities in hepatocellular carcinoma patients determine cancer relapse.

Detection of hepatocellular carcinoma circulating tumor cells performed with conventional strategies, is significantly limited due to inherently heterogeneous and dynamic expression of EpCAM, as well as degradation of cytokeratins during epithelial-to-mesenchymal transition, which inevitably lead to non-negligible false negative detection of such "uncapturable and invisible" CTCs. A novel SE-iFISH strategy, improved for detection of HCC CTCs in this study, was applied to comprehensively detect, in situ phenotypically and karyotypically characterize hepatocellular and cholangiocarcinoma CTCs (CD45/CD31) in patients subjected to surgical resection. Clinical significance of diverse subtypes of CTC was systematically investigated.

Comprehensive in situ co-detection of aneuploid circulating endothelial and tumor cells.

Conventional circulating tumor cell (CTC) detection strategies rely on cell surface marker EpCAM and intracellular cytokeratins (CKs) for isolation and identification, respectively. Application of such methods is considerably limited by inherent heterogeneous and dynamic expression or absence of EpCAM and/or CKs in CTCs. Here, we report a novel strategy, integrating antigen-independent subtraction enrichment and immunostaining-FISH (SE-iFISH), to detect a variety of aneuploid circulating rare cells (CRCs), including CTCs and circulating tumor endothelial cells (CECs).

Aneuploidy of chromosome 8 in circulating tumor cells correlates with prognosis in patients with advanced gastric cancer.

Objective: Previous work indicated that aneuploidy of chromosome 8 in circulating tumor cells (CTCs) correlated with therapeutic efficacy for advanced gastric cancer (AGC) patients. In this follow-up study performed on the same population of AGC patients, we investigated whether and how aneuploidy of chromosome 8 in CTCs correlates with patients' clinical prognosis.

Methods: The prospective study was performed on 31 patients with newly diagnosed AGC.

Enhanced detection and comprehensive in situ phenotypic characterization of circulating and disseminated heteroploid epithelial and glioma tumor cells.

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.

Comprehensive characterization of chemotherapeutic efficacy on metastases in the established gastric neuroendocrine cancer patient derived xenograft model.

The HuPrime® human gastric neuroendocrine carcinoma derived xenograft model GA0087 was established in this study. GA0087 PDX model showed high gene expression of vascular endothelial growth factors (VEGF)-A and B, and high potential of lung metastasis. Circulating tumor cells (CTCs) with either large or small size, circulating tumor microemboli (CTM) and lung metastatic lesions were detected in GA0087 PDX mice.

WITHDRAWN: Enhanced Detection and Phenotypic and Karyotypic in Situ Characterization of Circulating Tumor Cells.

Available online October 16, 2014 This article has been withdrawn at the request of the author(s) and/or editor. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at http://www.

Clinical significance of phenotyping and karyotyping of circulating tumor cells in patients with advanced gastric cancer.

Background: Karyotyping and phenotyping of circulating tumor cells (CTCs) in therapeutic cancer patients is of particular clinical significance in terms of both identifying chemo-resistant CTC subtypes and understanding CTC evolution.

Methods: The integrated subtraction enrichment (SET) and immunostaining-fluorescence in situ hybridization (iFISH) platform was applied to detect and characterize CTCs in patients with advanced gastric cancer (AGC). Status of human epidermal growth factor receptor 2 (HER2) expressing and aneuploidy of chromosome 8 in CTCs enriched from the patients was examined by SET-iFISH following clinical chemotherapy or HER2-targeted therapy.