Fernandes Glaucia Maria de Mendonça, Galbiatti-Dias Ana Lívia Silva, Ferreira Leticia Antunes Muniz, Serafim Junior Vilson, Rodrigues-Fleming Gabriela Helena, de Oliveira-Cucolo Juliana Garcia, Biselli-Chicote Patrícia Matos, Kawasaki-Oyama Rosa Sayoko, Maniglia José Victor, Pavarino Érika Cristina, Goloni-Bertollo Eny Maria
Genetics and Molecular Biology Research Unit (UPGEM), São José do Rio Preto Medical School (FAMERP) São José do Rio Preto, São Paulo, Brazil.
Department of Otolaryngology and Head and Neck Surgery, São José do Rio Preto Medical School (FAMERP) São José do Rio Preto, São Paulo, Brazil.
Am J Transl Res. 2021 Jan 15;13(1):143-155. eCollection 2021.
Laryngeal cancer (LC) is one of the common head and neck neoplasms and is characterized by resistance to conventional therapy and poor prognosis. This may result from the presence of cancer stem cells (CSCs), which form a small population in tumors with metastatic potential, high invasive capacity, self-renewal, and differentiation. This study aimed to evaluate the effectiveness of 5-fluorouracil and cisplatin individually, as well as the combination of cetuximab and paclitaxel in a CSC subpopulation separated with biomarkers related to tumoral growth (CD44, CD117, and CD133). In addition, expression of and -A genes and proteins related to cell proliferation were evaluated in this subpopulation. The CD44, CD133, and CD117 biomarkers were used to analyze the identification and separation of both subpopulations using FACSAria Fusion. Subpopulations positive for CD44, CD133, and CD117 or lacking these biomarkers were classified as laryngeal cancer stem cells (LCSCs) or laryngeal cancer non-stem cells (non-LCSCs), respectively. Matrigel invasion and colony forming assays were performed to confirm CSC presence. Subpopulations were cultured and exposed to 5-fluorouracil, cisplatin, and cetuximab/paclitaxel drugs for 24 h. Cell proliferation was determined using MTS assay. and gene expression levels were evaluated using quantitative real time PCR with TaqMan Assay in both subpopulations. The non-LCSC subpopulation was considered as the control for relative expression. We found that the LCSC subpopulation demonstrated more resistance to cetuximab and paclitaxel combination chemotherapy when compared with the non-LCSC subpopulation of the cell line. These LCSC subpopulations presented up-regulated expression of and genes and proteins and no gene expression, but TrkB protein expression was up-regulated in the LC cell line when compared to the non-CSC subpopulation. "In conclusion, the combination of CD44, CD133, and CD117 biomarkers has stem cell properties. Moreover, LCSCs, are capable of resisting treatment and present high and gene expression".
喉癌（LC）是常见的头颈部肿瘤之一，其特点是对传统治疗有抗性且预后较差。这可能是由于癌症干细胞（CSC）的存在，它们在具有转移潜能、高侵袭能力、自我更新和分化能力的肿瘤中占一小部分。本研究旨在评估5-氟尿嘧啶和顺铂单独使用的有效性，以及西妥昔单抗和紫杉醇联合使用对通过与肿瘤生长相关的生物标志物（CD44、CD117和CD133）分离出的CSC亚群的有效性。此外，还评估了该亚群中与细胞增殖相关的 和 -A基因及蛋白的表达。使用FACSAria Fusion通过CD44、CD133和CD117生物标志物分析这两个亚群的鉴定和分离。CD44、CD133和CD117呈阳性或缺乏这些生物标志物的亚群分别被分类为喉癌干细胞（LCSC）或喉癌非干细胞（非LCSC）。进行基质胶侵袭和集落形成试验以确认CSC的存在。将亚群培养并暴露于5-氟尿嘧啶、顺铂和西妥昔单抗/紫杉醇药物24小时。使用MTS试验测定细胞增殖。在两个亚群中使用TaqMan分析的定量实时PCR评估 和 基因表达水平。将非LCSC亚群视为相对表达的对照。我们发现，与细胞系的非LCSC亚群相比，LCSC亚群对西妥昔单抗和紫杉醇联合化疗表现出更强的抗性。这些LCSC亚群呈现 和 基因及蛋白的上调表达且无 基因表达，但与非CSC亚群相比，LC细胞系中的TrkB蛋白表达上调。“总之，CD44、CD133和CD117生物标志物的组合具有干细胞特性。此外，LCSC能够抵抗治疗并呈现高 和 基因表达”。
