Chen Hui, Zhang Xuemei, Wang Ziyi, Luo Jing, Liu Yingbin, Shao Rong
Shanghai Key Laboratory of Biliary Tract Disease Research, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200092, China.
Department of Pharmacology and Chemical Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China.
Cell Mol Life Sci. 2024 Dec 24;82(1):11. doi: 10.1007/s00018-024-05497-5.
Glioblastoma (GB) is the stage IV of glioma and mesenchymal GB represents the most common and malignant subtype characterized with elevated expression of a mesenchymal marker YKL-40 and resistance to immune drug therapy. Here, we determined if YKL-40 regulates kynurenine (Kyn) pathway (KP) metabolism that contributes to establishing an immune suppressive microenvironment in GB.
Tumor cells expressing YKL-40 from GB patients were isolated and activated cellular metabolisms were identified via gene microarray analysis. KP metabolism was determined by LC/MS/MS system. Indoleamine 2,3-dioxygenase 1 (IDO1), tryptophan 2,3-dioxygenase (TDO2), their regulatory transcription factors AhR and SRF were evaluated using WB. AhR and SRF transactivity was measured by luciferase reporter gene assays with binding motif mutation, while m6A-mediated AhR and SRF mRNA stability was determined in the presence of an METTL3inhibitor. YKL-40 and Kyn-induced tumor cell migration and CD8+ cytotoxic T cell (CTL) apoptosis were measured in cultured cells. Tumors cells expressing YKL-40 were injected to mouse brains to establish orthotpic tumor models. In GB, YKL-40, IDO1 and TDO2 expression was analyzed for correlation with patient survival.
KP metabolism was activated in YKL-40-expressing tumor cells. YKL-40 divergently regulated IDO1 and TDO2 via induction of AhR and SRF, respectively. mRNA levels of AhR and SRF were stabilized by decreased METTL3 and YTHDF2. YKL-40 and Kyn secreted from tumor cells and infiltrating M2 macrophages cooperated to enhance tumor cell migration and inhibit CTL immunity. In xenografts, tumors expressing YKL-40 displayed the elevated KP metabolism and macrophage infiltration, but decreased CTLs. Treatment with an anti-PD-1 antibody Tislelizumab significantly increased YKL-40+ mouse survival. In GB, YKL-40 was positively correlated with IDO1 expression and both were associated with decreased survival, whereas IDO1 was negatively correlated with TDO2.
YKL-40 upregulates IDO1 or TDO2 to activate KP metabolism, and coordinates with Kyn to establish an inhibitory tumor immune microenvironment, leading to tumor immune evasion.
胶质母细胞瘤（GB）是胶质瘤的IV期，间充质GB是最常见且恶性程度最高的亚型，其特征是间充质标志物YKL-40表达升高以及对免疫药物治疗耐药。在此，我们确定YKL-40是否调节犬尿氨酸（Kyn）途径（KP）代谢，该代谢有助于在GB中建立免疫抑制微环境。
从GB患者中分离出表达YKL-40的肿瘤细胞，并通过基因微阵列分析鉴定激活的细胞代谢。通过液相色谱/串联质谱系统测定KP代谢。使用蛋白质免疫印迹法评估吲哚胺2,3-双加氧酶1（IDO1）、色氨酸2,3-双加氧酶（TDO2）及其调节转录因子芳烃受体（AhR）和血清反应因子（SRF）。通过结合基序突变的荧光素酶报告基因测定法测量AhR和SRF的转录活性，而在存在甲基转移酶样蛋白3（METTL3）抑制剂的情况下测定m6A介导的AhR和SRF mRNA稳定性。在培养细胞中测量YKL-40和Kyn诱导的肿瘤细胞迁移以及CD8 + 细胞毒性T细胞（CTL）凋亡。将表达YKL-40的肿瘤细胞注射到小鼠脑内以建立原位肿瘤模型。在GB中，分析YKL-40、IDO1和TDO2的表达与患者生存率的相关性。
在表达YKL-40的肿瘤细胞中KP代谢被激活。YKL-40分别通过诱导AhR和SRF对IDO1和TDO2进行不同调节。AhR和SRF的mRNA水平通过降低METTL3和YTHDF2而稳定。肿瘤细胞和浸润的M2巨噬细胞分泌的YKL-40和Kyn协同作用以增强肿瘤细胞迁移并抑制CTL免疫。在异种移植中，表达YKL-40的肿瘤显示出升高的KP代谢和巨噬细胞浸润，但CTL减少。用抗程序性死亡蛋白1（PD-1）抗体替雷利珠单抗治疗显著提高了YKL-40 + 小鼠的生存率。在GB中，YKL-40与IDO1表达呈正相关，且两者均与生存率降低相关，而IDO1与TDO2呈负相关。
YKL-40上调IDO1或TDO2以激活KP代谢，并与Kyn协同作用建立抑制性肿瘤免疫微环境，导致肿瘤免疫逃逸。
