👤 Kazuaki Chikamatsu

The detection of circulating tumor cells (CTCs) using immunoaffinity-based methods often relies on epithelial-related markers, which may bias the selection of CTCs and limit the biological information obtained, depending on the targeted antigens. Herein, we compared the molecular profiles and clinical significance of CTCs based on the expression of epithelial-related markers ( Show less

Human papillomavirus (HPV)-positive head and neck squamous cell carcinoma (HNSCC) differs in terms of cellular and molecular biological characteristics from HPV-negative HNSCC. However, differences in circulating tumor cells (CTCs) between HPV-positive and -negative HNSCC remain unclear. We first analyzed eight epithelial-mesenchymal transition (EMT)-related genes (VIM, CDH1, CDH2, SNAI1, SNAI2, TWIST1, ZEB1, and ZEB2) using The Cancer Genome Atlas (TCGA) database. Next, we isolated CTCs from patients with HNSCC using CD45-negative selection and expression analysis of epithelial-related genes (EPCAM, EGFR, and MET) by RT-qPCR. CTC-positive samples were further analyzed for EMT-related genes. In addition, we investigated the proportion of circulating T cell subsets and CD38+ T cells using flow cytometry and their involvement in CTCs. Compared with HPV-negative HNSCC, expression of CDH1, SNAI1, SNAI2, TWIST1, and ZEB1 was downregulated in HPV-positive HNSCC, as determined by TCGA analysis. CTCs were detected in 19 (52.8 %) of 36 HPV-positive and 26 (68.4 %) of 38 HPV-negative patients with HNSCC. EPCAM-positive and MET-positive CTCs were significantly more frequent in patients with HPV-negative HNSCC. HPV-positive patients with HNSCC exhibited significantly high SNAI1 and ZEB2 expression in CTCs. Interestingly, differences in SNAI1 expression levels differed markedly between CTCs and TCGA based on HPV status. Moreover, HPV-positive patients with HNSCC exhibiting SNAI1-high CTCs showed a superior prognosis and a lower proportion of CD38+ T cells than those with SNAI1-low CTCs. Our results provide novel insights into the EMT-MET spectrum of CTCs and may contribute to the development of prognostic biomarkers for HPV-positive HNSCC. Show less

The emergence of immune checkpoint inhibitors (ICIs) has revolutionized the treatment of recurrent/metastatic (R/M) head and neck squamous cell carcinoma (HNSCC). Biomarkers of the therapeutic efficacy of ICIs have been extensively investigated. In this study, we aimed to analyze whether molecular phenotypes of circulating tumor cells (CTCs) are associated with treatment responses and clinical outcomes in patients with R/M HNSCC treated with nivolumab. Peripheral blood samples were collected before treatment initiation and after four infusions of nivolumab. CTCs isolated by depletion of CD45-positive cells were analyzed to determine the expression of EPCAM, MET, KRT19, and EGFR using real-time quantitative polymerase chain reaction. CTC-positive samples were analyzed to determine the expression of PIK3CA, CCND1, SNAI1, VIM, ZEB2, CD44, NANOG, ALDH1A1, CD47, CD274, and PDCD1LG2. Of 30 patients treated with nivolumab, 28 (93.3%) were positive for CTCs. In 20 CTC-positive patients, molecular alterations in CTCs before and after nivolumab treatment were investigated. Patients with MET-positive CTCs had significantly shorter overall survival than those with MET-negative CTCs (p = 0.027). The expression level of CCND1 in CTCs of disease-controlled patients was significantly higher than that of disease-progressed patients (p = 0.034). In disease-controlled patients, the expression level of CCND1 in CTCs significantly decreased after nivolumab treatment (p = 0.043). The NANOG expression in CTCs was significantly increased in disease-controlled patients after nivolumab treatment (p = 0.036). Our findings suggest that the molecular profiling of CTCs is a promising tool to predict the treatment efficacy of nivolumab. Show less

The relationship between the molecular profiling of circulating tumor cells (CTCs) and clinical factors is a challenge. In this study, we performed molecular detection and characterization of CTCs in patients with head and neck squamous cell carcinoma (HNSCC). CTCs captured by microfilter were analyzed for the expression of multiple epithelial markers (EPCAM, MET, KRT19, and EGFR) by RT-qPCR. The CTCs-positive samples were further analyzed for the expression of 10 genes (PIK3CA, CCND1, SNAI1, VIM, CD44, NANOG, ALDH1A1, CD47, CD274, and PDCD1LG2). Finally, we analyzed whether the molecular profiling of CTCs was associated with clinical factors. Twenty-eight (63.6%) of the 44 HNSCC patients were positive for at least one epithelial-related gene. CTC-positivity was significantly correlated with treatment resistance (p = 0.0363), locoregional recurrence (p = 0.0151), and a shorter progression-free survival (PFS) (p = 0.0107). Moreover, the expression of MET in CTCs was associated with a shorter PFS (p = 0.0426). Notably, patients with CD274-positive CTC showed prolonged PFS (p = 0.0346) and overall survival (p = 0.0378) compared to those with CD274-negative CTC. Our results suggest that molecular profiling characterized by the gene expression of CTCs influences clinical factors in patients with HNSCC. Show less