Single-Cell RNA Sequencing Reveals the Critical Role of SEC16B in Lung Metastasis of Osteosarcoma

Osteosarcoma (OS) is highly malignant and easily prone to lung metastasis. The mechanisms of lung metastasis in OS remain unclear. The single-cell RNA sequencing (scRNA-seq) samples in this study included six primary osteosarcoma samples (published in-house data), two lung metastasis samples (GSE152048), and four normal bone tissue samples (GSE169396). To identify potential targets for metastasis, bulk RNA sequencing data from four primary tumors and four lung metastases (in-house data) were also analyzed. scRNA-seq identified five tumor cell subpopulations. CytoTRACE and lung metastasis scores indicated that the C1 subpopulation was most closely associated with lung metastasis. By intersecting lung metastasis-related genes identified via hdWGCNA analysis with differentially expressed genes from bulk RNA sequencing, SEC16B was identified as the key gene influencing lung metastasis. qRT-PCR results revealed that SEC16B expression was significantly downregulated in OS cell lines. Transwell assay demonstrated that overexpression of SEC16B significantly inhibited the invasion and migration capabilities of OS cells. Additionally, analyses using Scissor, CellphoneDB, and CSOmap suggested that fibroblasts, endothelial cells, and OS cells in the tumor microenvironment formed a pre-metastatic niche through mechanisms involving angiogenesis and extracellular matrix remodeling. Overall, this study identifies a new population that may promote lung metastasis by downregulating SEC16B in OS. Moreover, fibroblasts and endothelial cells in the tumor microenvironment play a critical role in OS lung metastasis.