Single-cell transcriptomic analyses provide insights into SPP1+ TAM-mediated immune suppression and CD8+ T cell dysfunction in lung cancer
Rahma Taher Almgrami, Tengyue Zhang, Qitai Zhao, M. James You, Jinyan Liu, Yi Zhang
Abstract
Metastatic lung carcinoma poses considerable treatment difficulties. It exhibits cellular characteristics that differ from those of early-stage cancer. Immunotherapy demonstrates enhanced hope for patients with advanced and metastatic lung cancer. Tumor-associated macrophages (TAMs) may be a contributing factor that diminishes the effectiveness of immunotherapy. Secreted phosphoprotein 1 (SPP1) + TAMs are considered to possess immunosuppressive characteristics, as their interaction with CD8 + T lymphocytes results in the exhaustion of these cells. We analyzed single-cell RNA sequencing datasets of lung cancer metastases in order to examine the development of brain metastatic tumors. Furthermore, the cell–cell interactions between SPP1 + TAM cells and CD8 + T cells within primary and brain metastatic cancers were systematically compared. Additionally, we utilized a flow cytometer and immunofluorescence to demonstrate how SPP1 affects the function of CD8 + T cells. In vitro, we generated SPP1-overexpressing macrophages and performed qPCR, Western blot, and co-culture assays with or without anti-SPP1 or anti-A2AR treatment to evaluate immunosuppressive effects. The results demonstrated that the proportions of immune cells in metastatic brain tissues are lower, while the infiltration of macrophages is higher. SPP1 + TAMs contribute to immune suppression in lung cancer by limiting the activation of CD8 + T cells and cytokine production. Anti-SPP1 treatment positively impacts CD8 + T cell function, counteracting SPP1-induced dysfunction and facilitating the production of cytokines. SPP1 overexpression in macrophages enhanced their immunosuppressive phenotype by upregulating CD73 and cytokines such as IL-10 and TGF-β, leading to impaired CD8⁺ T cell function via A2AR signaling; notably, neutralization of SPP1 or A2AR successfully restored CD8⁺ T cell activity. Our findings characterize the immunological environment of both primary and metastatic lung cancer, highlighting SPP1-mediated immune suppression as a potential therapeutic target to restore T cell responses. Preclinical data demonstrate that Anti-SPP1 antibodies can reverse T cell exhaustion and enhance immune responses in lung cancer.