SPP1 <sup>high</sup> macrophage-induced T-cell stress promotes colon cancer liver metastasis through SPP1/CD44/PI3K/AKT signaling
Dongbing Ding, Weibo Li, Jiannan Ren, Yiquan Li, Guangchun Jiang, Rongpu Liang, He Huang, Jiarong You, Yuan Wang, Bo Wei
Abstract
Background Patients with colon cancer liver metastases (CCLM) frequently exhibit poor responses to immunotherapy, a phenomenon attributed in part to an immune desert tumor microenvironment. This study aimed to comprehensively characterize the immune landscape in primary colon cancers and their matched liver metastases via single-cell transcriptome analysis, with the goal of identifying potential immunotherapeutic targets. Methods Tumor specimens from patients with CCLM were subjected to single-cell RNA sequencing. Immune subpopulations were profiled with emphasis on exhausted T cells (Tex)—including both CD8 + and CD4 + ANK3 + subsets—as well as on a distinct stress response T-cell subset (TSTR) defined by high HSPA1A/HSPA1B expression. In parallel, we performed assessments of the phenotype and prognostic impact of SPP1 high myeloid cells, along with in vitro assays to examine their role in modulating T-cell number and function. Results Liver metastatic lesions exhibited a significantly elevated infiltration of Tex compared with primary tumors. Notably, Tex cells exhibited upregulated expression of exhaustion-related marker genes such as ANK3, ZBTB20, ETV6, and CAMK4, which were markedly downregulated in TSTR cells. TSTR was identified as an intermediate developmental state between effector and exhausted T cells in patients with CCLM, suggesting that TSTR cells represent a distinct state from exhausted T cells. Furthermore, myeloid cells expressing high levels of secreted phosphoprotein 1 (SPP1), along with apolipoprotein C-I and apolipoprotein E, were associated with poor prognosis in patients with CCLM. In vitro studies revealed that Macro_SPP1 high cells diminished T-cell populations and triggered a stress response state in both CD4 + and CD8 + T cells via the SPP1/CD44/PI3K/AKT signaling pathway in a CD44-dependent manner. Importantly, combination treatment with anti-SPP1 and anti-programmed cell death protein-1 antibodies significantly inhibited liver metastasis growth, enhanced dendritic cell maturation, decreased M2-polarized macrophages, and restored T-cell infiltration and function. Conclusions These findings reveal a previously unrecognized relationship between Macro_SPP1 high cells and HSPA1A high /HSPA1B high T cells in driving CCLM progression, suggesting a potential synergistic therapeutic approach that could boost immune checkpoint treatment efficacy in patients with CCLM.