Rev-erbα is a nuclear receptor, which regulates circadian rhythm, inflammatory responses and lipid metabolism. We previously showed Rev-erbα reduction in human gastric cancer, which is associated with TMN stages and poor prognosis. We hypothesized that Rev-erbα modulates proliferation via glycolytic flux and the pentose phosphate pathway (PPP) in gastric cancer. Knockdown of Rev-erbα significantly increased proliferation as well as glycolytic flux and the PPP in human gastric cancer cells. These effects were reduced by a Rev-erbα agonist GSK4112 in a dose-dependent manner. Furthermore, Rev-erbα was recruited on the promoters of PFKFB3 and G6PD genes, thereby inhibiting their gene transcription. GSK4112 treatment reduced PFKFB3 and G6PD gene expression, which was not affected by BMAL1 knockdown. Pharmacological inhibition of glycolysis and the PPP using corresponding PFKFB3 and G6PD inhibitors attenuated Rev-erbα knockdown-induced proliferation in gastric cancer cells. GSK4112 treatment was not able to reduce proliferation in SGC-7901 overexpressing both PFKFB3 and G6PD genes. Both PFKFB3 and G6PD were overexpressed in patients with gastric cancer, and positively correlated with the TMN stages. The PPP and glycolysis were enhanced in gastric cancer tissues of patients with low expression of Rev-erbα compared to the patients with high expression of Rev-erbα. In conclusion, Rev-erbα reduction causes gastric cancer progression by augmenting the PPP and glycolysis.

Gastric cancer is the fifth most diagnosed malignant tumor worldwide with limited effective chemotherapy. Ferroptosis is a new type of programmed cell death, which is becoming as a novel therapeutic target for tumors. Arenobufagin (ArBu) is a bufadienolide isolated from toad skin and venom, which exhibits broad-spectrum anti-tumor activity. It is unclear whether ArBu causes ferroptosis, thereby exhibiting anti-tumor activity in gastric cancer. We aimed to determine whether ArBu causes ferroptosis in cultured human gastric cancer cells. Different human gastric cancer cells were treated with ArBu (5-20 μM, 48 h). Indicators of apoptosis and ferroptosis were measured. CRISPR/Cas-9 system was employed to delete Nr1d1 gene. ArBu incubation reduced cell viability in a concentration-dependent manner. ArBu caused ferroptosis but not apoptosis at a lower concentration (10 μM), despite it caused both of them at a higher concentration (20 μM). Cotreatment with a selective ferroptosis inhibitor ferrostatin-1 protected against ArBu (10 μM)-induced reduction in cell viability. ArBu-mediated ferroptosis was associated with abnormal expression of genes involved in iron uptake, lipid peroxidation, and antioxidants. Particularly, Nr1d1 gene expression was most significantly increased after ArBu treatment. Furthermore, activating Rev-erbα encoded by Nr1d1 by a selective agonist GSK4112 (1 and 2 μM, 48 h) caused ferroptosis. In contrast, Rev-erbα knockout using the CRISPR/Cas-9 system diminished ArBu-induced ferroptosis in cultured human gastric cancer cells. ArBu causes ferroptosis by increasing Rev-erbα expression in human gastric cancer cells. This has implications of ArBu as a promising therapy for gastric cancer. 1. Natural Products. Traditional medicine, pharmacology, gastric cancer, signal pathway.

Gastric cancer is a serious threat to human health. Nuclear receptor subfamily 1 group D member 1 (REV‑ERBα) is a member of the nuclear hormone receptor family that regulates lipid metabolism, inflammatory responses and circadian rhythms. However, the role of REV‑ERBα in the pathogenesis of human gastric cancer is unclear. The present study employed gastric cancer tissues from 74 patients and determined the association between REV‑ERBα expression with clinicopathological variables and prognosis. Furthermore, the association between REV‑ERBα and apoptosis in undifferentiated and moderately differentiated human gastric cancer cells was determined. It was identified that REV‑ERBα expression was decreased in gastric cancer, which was positively associated with poor differentiation (P=0.009), T stage (P=0.001), Tumor‑Node‑Metastasis (TMN) stage (P=0.001) and lymph node metastasis (P=0.007). In the survival analysis, the 3‑ and 5‑year survival times of patients were significantly associated with REV‑ERBα expression (P=0.009 and P=0.002, respectively). Low REV‑ERBα expression was associated with poor prognosis (P<0.05). Concurrently, cleaved caspase‑3 expression was downregulated, whereas expression levels of Bcl‑2 and the Bcl‑2/Bax ratio were upregulated in gastric cancer tissues compared with normal tissues. REV‑ERBα activator GSK4112 caused apoptosis in SGC‑7901 and BGC‑823 cell lines. REV‑ERBα levels were decreased in human gastric cancer, which was associated with poor differentiation, TMN stages and poor prognosis. REV‑ERBα is a potential biomarker for tumor development and prognosis, and a potential therapeutic target for gastric cancer.

Nuclear receptor retinoid-related orphan receptor alpha promotes apoptosis but is reduced in human gastric cancer