Zhan Yizhi, Xu Jinsong, Zhang Zhanqiao, Hu Yating, Li Yongsheng, Qian Junying, Ling Yunyan, Wu Dehua, Deng Haijun, Li Guoxin, Shen Zhiyong, Fang Yuan
Department of Pathology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong, P. R. China.
Department of Pathology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong, P. R. China.
Cancer Commun (Lond). 2025 Jul 16. doi: 10.1002/cac2.70047.
Colorectal cancer liver metastasis (CRLM) is characterized by an immunosuppressive microenvironment and a blunted response to immunotherapy. Notably, tumor-associated macrophages (TAMs) play a critical role in modulating immune responses and exhibit significant heterogeneity in CRLM. Sphingosine kinase 1 (SPHK1) serves as a pivotal kinase in maintaining the balance between ceramide and sphingosine-1-phosphate (S1P) levels. However, the effects of SPHK1 within TAMs on tumor immune evasion during CRLM remain elusive. This study aimed at investigating the role of TAM-intrinsic SPHK1 in tumor immunosuppressive microenvironment in CRLM.
SPHK1 expression levels in TAMs were estimated by immunofluorescence and bioinformatics analysis. Several animal models were established to elucidate the role of SPHK1 in tumor immunity reprogramming in vivo. Flow cytometry, cytokine assay, and transwell assay were conducted to investigate the effects of SPHK1 in TAMs in cell-cell communication in vitro. RNA-sequencing, Western blotting, and quantitative real-time polymerase chain reaction were used to explore the molecular mechanism by which SPHK1 activated NLR family pyrin domain containing 3 (NLRP3) inflammasome in TAMs.
We found that SPHK1 was mainly expressed in TAMs and identified SPHK1 TAMs as associated with CRLM and diminished efficacy of immunotherapy in human patients. These SPHK1 TAMs exhibited strong immunosuppressive activities by inducing CD8 T cell exhaustion with high programmed cell death 1 (PD-1) expression via the interaction between TAMs and CRC cells. Mechanistically, SPHK1-produced S1P exerted an autocrine effect to activate NLRP3 inflammasome and interleukin 1 beta (IL-1β) release via nuclear factor-kappa B (NF-κB) and hypoxia inducible factor 1 subunit alpha (HIF-1α) signaling in TAMs. Paracrine IL-1β then upregulated the expression of monocyte chemoattractants and ADAM metallopeptidase domain 17 (ADAM17) sheddase in CRC cells, resulting in TAM infiltration and CD8 T cell dysfunction in the liver microenvironment. Furthermore, combining SPHK1-targeting treatments with anti-PD-1 therapy or radioimmunotherapy largely stalled liver metastasis and caused a significant extension of lifespan in preclinical mouse models.
Our findings highlighted the role of SPHK1 of TAMs in facilitating CRLM by promoting CD8 T cell dysfunction and immunosuppressive microenvironment. Combining SPHK1 blockade with anti-PD-1 therapy may be a promising treatment regimen for patients with CRLM.
结直肠癌肝转移（CRLM）的特征是免疫抑制微环境以及对免疫疗法反应迟钝。值得注意的是，肿瘤相关巨噬细胞（TAM）在调节免疫反应中起关键作用，并且在CRLM中表现出显著的异质性。鞘氨醇激酶1（SPHK1）是维持神经酰胺和1-磷酸鞘氨醇（S1P）水平平衡的关键激酶。然而，SPHK1在TAM中对CRLM期间肿瘤免疫逃逸的影响仍不清楚。本研究旨在探讨TAM内在的SPHK1在CRLM肿瘤免疫抑制微环境中的作用。
通过免疫荧光和生物信息学分析评估TAM中SPHK1的表达水平。建立了几种动物模型以阐明SPHK1在体内肿瘤免疫重编程中的作用。进行流式细胞术、细胞因子测定和Transwell测定以研究SPHK1在TAM中对体外细胞间通讯的影响。采用RNA测序、蛋白质免疫印迹和定量实时聚合酶链反应来探索SPHK1激活TAM中含NLR家族pyrin结构域3（NLRP3）炎性小体的分子机制。
我们发现SPHK1主要在TAM中表达，并确定SPHK1+ TAM与人类患者的CRLM和免疫治疗疗效降低相关。这些SPHK1+ TAM通过TAM与结直肠癌细胞之间的相互作用诱导高程序性细胞死亡蛋白1（PD-1）表达的CD8 + T细胞耗竭，从而表现出强大的免疫抑制活性。机制上，SPHK1产生的S1P通过TAM中的核因子-κB（NF-κB）和缺氧诱导因子1α亚基（HIF-1α）信号传导发挥自分泌作用，激活NLRP3炎性小体并释放白细胞介素1β（IL-1β）。旁分泌的IL-1β随后上调结直肠癌细胞中单核细胞趋化因子和ADAM金属肽酶结构域17（ADAM17）裂解酶的表达，导致肝微环境中TAM浸润和CD8 + T细胞功能障碍。此外，在临床前小鼠模型中，将靶向SPHK1的治疗与抗PD-1治疗或放射免疫治疗相结合在很大程度上阻止了肝转移并显著延长了生存期。
我们的研究结果突出了TAM中的SPHK1通过促进CD8 + T细胞功能障碍和免疫抑制微环境在促进CRLM中的作用。将SPHK1阻断与抗PD-1治疗相结合可能是CRLM患者有前景的治疗方案。
