Using single-cell sequencing to elucidate the mechanism by which immune cell-to-cell communication regulates bone metastasis in multi- organ cancers

To investigate the regulatory mechanisms of immune cell-cell communication in bone metastasis across multiple cancer types, this study employed single-cell RNA sequencing (scRNA-seq) to analyze normal bone marrow and bone metastasis tissue samples from multiple cancer types, including breast, lung, cervical, and renal cancers, from the GEO database (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE266330).First, all cells were classified into 16 populations, including B cells, cycling cells, and macrophages, based on gene expression profiles. The authors found that the proportion of T cells and B cells decreased in the bone metastasis group compared with the normal control group (ctrl). Furthermore, the cell type composition of bone metastasis tissues varied across cancer types. For example, T cells were more abundant in the renal cancer (KC) and lung cancer (LC) bone metastasis groups, monocytes were more abundant in the breast cancer (BC) bone metastasis group, and epithelial cells were more abundant in the colon cancer (CC) bone metastasis group. Furthermore, multiple signaling pathways were upregulated in the BC, CC, and LC groups. Secondly, focusing on macrophages, they divided them into nine subpopulations and found that the proportion of pro-oncogenic macrophage subpopulations (such as C4_Macro_PhagoLysosome and C0_Macro_ComplementHigh) in bone metastasis samples was significantly increased. Furthermore, the expression of tumor suppressor genes such as RUNX3 and SYK was decreased, while the expression of pro-oncogenic genes such as MSR1 and TREM2 was increased. Further analysis screened out 12 key signaling pathways that regulate bone metastasis, including oxidative phosphorylation, glycolysis, hypoxia, and inflammatory response. These pathways work synergistically to promote bone metastasis through metabolic adaptation, immune escape, bone remodeling, and cell invasion. Furthermore, the regulatory patterns of macrophage subpopulations in bone metastasis tissues of different cancer types vary. Subsequent analysis of NK cells and T cells revealed that NK cell subtypes in bone metastasis tissues were more complex, with increased proportions of NK_C1_CD56dim (strong cytotoxic function) and NK_C0_CD56bright (immunomodulatory function), and increased expression of immune-activating genes such as XCL1 and XCL2. T cells were divided into 13 subpopulations, with significant changes in the proportion of immunosuppressive T cells (such as Tregs) in the bone metastasis group. Tumor suppressor genes such as SPOCK2 and RASA3 were downregulated, while immune-related genes such as GZMB and SPP1 were upregulated. These genes regulate T cell differentiation and function through the aforementioned 12 key signaling pathways. Finally, intercellular communication analysis revealed that communication between immune cells and other cells increases explosively after bone metastasis. Osteoclasts serve as key signaling sources in BC, CC, and LC, extensively communicating with immune cells through signaling molecules such as SPP1, whereas osteoclast communication is weaker in KC. Osteoclasts also drive bone metastasis through four mechanisms: a "vicious cycle," bone remodeling, immunosuppression, and promotion of tumor invasion and angiogenesis.This study reveals the differentiation patterns of immune cell subtypes and intercellular communication mechanisms in bone metastasis across multiple cancer types, providing a theoretical basis for clinically inhibiting tumor bone metastasis and secondary metastasis.