Chronic stress–induced ANPEP drives liver cancer progression by increasing glutathione synthesis and inhibiting ferroptosis
Yongkang Wu, Yankun Zhang, Xiaojia Shi, Mengting Wu, Min Sun, Ying Feng, Wenmeng Ma, Xiule Jiang, Dingqi Fei, Mingjian Zhao, Zhuanchang Wu, Chunyang Li, Xiaohong Liang, Lifen Gao, Chunhong Ma, Xuetian Yue
Yongkang Wu, Yankun Zhang, Xiaojia Shi, Mengting Wu, Min Sun, Ying Feng, Wenmeng Ma, Xiule Jiang, Dingqi Fei, Mingjian Zhao, Zhuanchang Wu, Chunyang Li, Xiaohong Liang, Lifen Gao, Chunhong Ma, Xuetian Yue
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Research Article Cell biology Gastroenterology Hepatology

Chronic stress–induced ANPEP drives liver cancer progression by increasing glutathione synthesis and inhibiting ferroptosis

Abstract

Emerging evidence demonstrates that chronic stress alters immunological, neurochemical, and endocrinological functions, thereby promoting tumor progression. However, the underlying metabolic mechanism of chronic stress in tumor progression is still elusive. Using multiomics analysis, we found that aminopeptidase N (ANPEP) was upregulated in tumors with chronic restraint, associating with the reprogramming of amino acid metabolism. Functional assays revealed that ANPEP promoted liver cancer growth and metastasis. Knockdown of ANPEP blocked chronic stress–induced liver cancer progression. Chronic stress–induced glucocorticoids promoted nuclear receptor subfamily 3 group C member 1 nuclear translocation to activate ANPEP transcription by directly binding to its promoter. Furthermore, ANPEP promotes glutathione synthesis, subsequently inhibiting ROS-induced ferroptosis. Mechanistically, ANPEP interacted with solute carrier family 3 member 2 (SLC3A2) to block membrane associated ring-CH-type finger 8–mediated lysosome-dependent degradation of SLC3A2, promoting intracellular l-cystine transport, thereby increasing glutathione synthesis. The combination of ANPEP silencing and sorafenib treatment showed a synergistic effect in inhibiting liver cancer progression. Finally, clinical data and mouse models demonstrated that chronic stress drove liver tumor progression via ANPEP-regulated SLC3A2. These findings reveal unanticipated communication between chronic stress and metabolic reprogramming during liver cancer progression, providing potential therapeutic implications for liver cancer.

Authors

Yongkang Wu, Yankun Zhang, Xiaojia Shi, Mengting Wu, Min Sun, Ying Feng, Wenmeng Ma, Xiule Jiang, Dingqi Fei, Mingjian Zhao, Zhuanchang Wu, Chunyang Li, Xiaohong Liang, Lifen Gao, Chunhong Ma, Xuetian Yue

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Figure 4

Chronic stress promotes ANPEP transcription through NR3C1 activation.

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Chronic stress promotes ANPEP transcription through NR3C1 activation. (A...
(A and B) ANPEP mRNA and protein levels in Huh7 and HepG2 cells with indicated manipulations after 48 hours of DEX treatment (A, n = 3). (C and D) ANPEP mRNA and protein levels in control and NR3C1-knockdown Huh7 and HepG2 cells after 48 hours of DEX treatment (C, n = 3). (E) ANPEP promoter activity in HEK293 with or without NR3C1 knockdown after DEX treatment (n = 3). (F) A schematic of NR3C1 binding regions on the ANPEP promoter based on the ENCODE database. Corresponding truncations of the ANPEP promoter were constructed, and biotin-labeled DNA probes were designed. (G) Luciferase activity of ANPEP promoter truncations in HEK293 cells with or without NR3C1 overexpression, followed by 48 hours of DEX treatment (n = 3). (H) Biotin-labeled DNA probes were incubated with nuclear extracts from HepG2 cells with or without DEX treatment, followed by streptavidin pull-down and Western blotting. (I and J) Cell proliferation (I) and Transwell assays (J) in Huh7 and HepG2 cells with indicated manipulations, after 48 hours of DEX treatment (I, n = 5; J, n = 6). Data are presented as mean ± SD. Statistical significance was determined by 2-way ANOVA with Šidák’s multiple comparisons test (I), 1-way ANOVA with Tukey’s multiple comparisons test (C, E, and J) and 2-tailed unpaired Student’s t test (A and G). *P < 0.05; **P < 0.01; ***P < 0.001.