Analysis of the Single-Cell Heterogeneity of Adenocarcinoma Cell Lines and the Investigation of Intratumor Heterogeneity Reveals the Expression of Transmembrane Protein 45A (TMEM45A) in Lung Adenocarcinoma Cancer Patients

Simple Summary Non-small cell lung cancer (NSCLC) is one of the main causes of cancer-related deaths worldwide. Intratumoral heterogeneity (ITH) is responsible for the majority of difficulties encountered in the treatment of lung-cancer patients. Therefore, the heterogeneity of NSCLC cell lines and primary lung adenocarcinoma was investigated by single-cell mass cytometry (CyTOF). Human NSCLC adenocarcinoma cells A549, H1975, and H1650 were studied at single-cell resolution for the expression pattern of 13 markers: GLUT1, MCT4, CA9, TMEM45A, CD66, CD274, CD24, CD326, pan-keratin, TRA-1-60, galectin-3, galectin-1, and EGFR. The intra- and inter-cell-line heterogeneity of A549, H1975, and H1650 cells were demonstrated through hypoxic modeling. Additionally, human primary lung adenocarcinoma, and non-involved healthy lung tissue were homogenized to prepare a single-cell suspension for CyTOF analysis. The single-cell heterogeneity was confirmed using unsupervised viSNE and FlowSOM analysis. Our results also show, for the first time, that TMEM45A is expressed in lung adenocarcinoma. Intratumoral heterogeneity (ITH) is responsible for the majority of difficulties encountered in the treatment of lung-cancer patients. Therefore, the heterogeneity of NSCLC cell lines and primary lung adenocarcinoma was investigated by single-cell mass cytometry (CyTOF). First, we studied the single-cell heterogeneity of frequent NSCLC adenocarcinoma models, such as A549, H1975, and H1650. The intra- and inter-cell-line single-cell heterogeneity is represented in the expression patterns of 13 markers-namely GLUT1, MCT4, CA9, TMEM45A, CD66, CD274 (PD-L1), CD24, CD326 (EpCAM), pan-keratin, TRA-1-60, galectin-3, galectin-1, and EGFR. The qRT-PCR and CyTOF analyses revealed that a hypoxic microenvironment and altered metabolism may influence cell-line heterogeneity. Additionally, human primary lung adenocarcinoma and non-involved healthy lung tissue biopsies were homogenized to prepare a single-cell suspension for CyTOF analysis. The CyTOF showed the ITH of human primary lung adenocarcinoma for 14 markers; particularly, the higher expressions of GLUT1, MCT4, CA9, TMEM45A, and CD66 were associated with the lung-tumor tissue. Our single-cell results are the first to demonstrate TMEM45A expression in human lung adenocarcinoma, which was verified by immunohistochemistry.