Department of Translational Molecular Pathology, University of Texas MD Anderson Cancer Center, Houston, TX, USA.
Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut, Lebanon.
Methods Mol Biol. 2021;2279:187-198. doi: 10.1007/978-1-0716-1278-1_15.
Interrogation and characterization of lung cancer stem cells (CSCs) that are implicated in lung oncogenesis is crucial for our understanding of inter- and intra-tumor heterogeneity and the aggressive nature of the disease, including tumor resistance and relapse in response to conventional therapy. Here, we describe an in vitro surrogate model, namely the "sphere-forming assay," for the derivation, enrichment, and propagation of lung stem/progenitor cells with CSC properties, including self-renewal, tumor initiation capacity and propagation, and differentiation into cells of the tumor bulk, from a murine Kras-mutant lung adenocarcinoma cell line. Self-renewing cancer stem/progenitor cells, in the form of 3D in vitro spheres, can be phenotypically interrogated using downstream techniques such as gene expression analysis (e.g., whole transcriptome sequencing and quantitative real time PCR), which is the focus of this chapter.
研究与肺癌发生相关的肺癌干细胞（CSC）对于我们理解肿瘤内和肿瘤间异质性以及疾病的侵袭性，包括肿瘤耐药性和对常规治疗的复发至关重要。在这里，我们描述了一种体外替代模型，即“球体形成测定法”，用于从鼠 Kras 突变肺腺癌细胞系中衍生、富集和扩增具有 CSC 特性的肺干/祖细胞，包括自我更新、肿瘤起始能力和增殖，以及分化为肿瘤实质细胞。自我更新的癌症干细胞/祖细胞可形成 3D 体外球体，可使用下游技术（例如基因表达分析（例如，全转录组测序和定量实时 PCR））进行表型分析，这是本章的重点。
