PO.MCB09.01 · 分子与细胞生物学
色氨酸代谢物驱动三阴性乳腺癌细胞的铁死亡抵抗
Tryptophan metabolites drive ferroptosis resistance in triple-negative breast cancer cells
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
引言
由于色氨酸双加氧酶TDO2、IDO1和IDO2的表达升高，色氨酸（Trp）代谢物在多种癌症类型中增加。虽然近期研究强调了这些代谢物的免疫抑制功能，但其对癌细胞存活的内在意义仍知之甚少。本研究探究了色氨酸分解代谢的前两种代谢物——甲酰犬尿氨酸（formylkynurenine）和犬尿氨酸（kynurenine）在三阴性乳腺癌（TNBC）铁死亡抵抗中的作用。
方法
我们使用来自SRA数据库的46例ER+、103例TNBC肿瘤和21例邻近正常组织样本进行了全面的基因表达分析。分析BRCA-TCGA的RNA-seq和DNA-seq数据，以鉴定TNBC患者中三种色氨酸双加氧酶的改变。对以50 μM甲酰犬尿氨酸和犬尿氨酸培养的BT549细胞系进行RNA-seq。使用蛋白质印迹和实时qPCR验证基因表达变化。通过LC-MS和MS/MS方法定量细胞内谷胱甘肽（GSH）和氧化型谷胱甘肽（GSSG）水平。通过用300 nM RSL3处理细胞评估细胞对铁死亡的敏感性。
结果
我们的分析显示，与管腔型和邻近正常组织相比，TNBC肿瘤中三种色氨酸双加氧酶TDO2、IDO1和IDO2的表达均增加。在IDO1和IDO2中检测到DNA扩增，且与管腔型亚型相比，TNBC样本中IDO1启动子甲基化降低。甲酰犬尿氨酸和犬尿氨酸在50-100 μM浓度下均显著增强BT549细胞系的增殖。转录组分析表明，这些代谢物调控参与ROS代谢和铁转运的基因。代谢组学分析进一步证实细胞内GSH/GSSG比值发生改变。值得注意的是，我们首次报道犬尿氨酸上调GPX4蛋白表达，而甲酰犬尿氨酸则不然。然而，两种代谢物均增加了细胞对RSL3诱导铁死亡的抵抗。
结论
甲酰犬尿氨酸和犬尿氨酸均增加GSH/GSSG比值。犬尿氨酸上调GPX4表达，而甲酰犬尿氨酸不改变其水平。这些发现确立了一条将色氨酸分解代谢与TNBC铁死亡抵抗相联系的新型代谢轴，提示靶向该通路可能代表针对这一侵袭性乳腺癌亚型的有前景的治疗策略。
查看英文原文 English abstract
Introduction
Tryptophan (Trp) metabolites are increased in various cancer types due to elevated expression of Trp dioxygenases, TDO2, IDO1, and IDO2. While recent studies have highlighted the immunosuppressive functions of these metabolites, their intrinsic significance to cancer cell survival remains poorly understood. This study explores the role of formylkynurenine and kynurenine, the first two metabolites in Trp catabolism on triple-negative breast cancer (TNBC) resistance to ferroptosis.
Methods
We performed comprehensive gene expression analysis using 46 ER+, 103 TNBC tumours, and 21 adjacent normal tissue samples from the SRA database. BRCA-TCGA RNA-seq and DNA-seq data were analysed to identify alterations in three Trp dioxygenases in TNBC patients. RNA-seq was conducted on BT549 cell line cultured with 50 μM formylkynurenine and kynurenine. Gene expression changes were validated using western blotting and real-time qPCR. Intracellular glutathione (GSH) and oxidized glutathione (GSSG) levels were quantified by LC-MS and MS/MS approach. Cell sensitivity to ferroptosis was evaluated by treating cells with 300 nM RSL3.
Results
Our analysis revealed increased expression of all three Trp dioxygenases, TDO2, IDO1, and IDO2, in TNBC tumours compared to luminal and adjacent normal tissues. DNA amplification was detected in IDO1 and IDO2, and reduced IDO1 promoter methylation in TNBC samples compared to luminal subtypes. Formylkynurenine and kynurenine both at concentrations of 50-100 μM significantly enhanced BT549 cell line proliferation. Transcriptomic analysis indicated that these metabolites regulate genes involved in ROS metabolism and iron transport. Metabolomics analysis further demonstrated alterations in the intracellular GSH/GSSG ratio. Notably, we report for the first time that kynurenine upregulates GPX4 protein expression, while formylkynurenine does not. However, both metabolites increased cellular resistance to RSL3-induced ferroptosis.
Conclusion
Formylkynurenine and kynurenine both increase GSH/GSSG ratio. While kynurenine upregulates GPX4 expression, formylkynurenine does not alter its levels. These findings establish a novel metabolic axis linking Trp catabolism to ferroptosis resistance in TNBC, suggesting that targeting this pathway could represent a promising therapeutic strategy for this aggressive subtype of breast cancer.
利益披露 Disclosure
J. Motalebzadeh, None..
H. Anani, None..
C. Thompson-Peach, None..
T. Hickey, None..
L. Butler, None..
N. Robinson, None..
D. Thomas, None.