Physciosporin targeting of cellular myelocytomatosis oncogene in gastric cancer suppresses energy metabolism and metastasis via lysosomal degradation

Abstract

Background: Gastric cancer (GCa), the fourth leading cause of cancer-related death worldwide, remains a major global health burden due to its cellular and molecular heterogeneity, poor prognosis, and high incidence. These challenges highlight the importance of finding effective preventive and therapeutic strategies. Previous studies identified Physciosporin (PHY) as a promising anticancer candidate; however, its effects on GCa have not yet been explored.

Purpose: We examined the impact of PHY on energy metabolism and metastasis in human GCa cells.

Methods: The effect of PHY on cell viability in GCa was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, while its ability to suppress metastasis was evaluated through scratch wound migration and invasion assays at non-toxic concentrations. The impact of PHY on energy metabolism was tested via glycolytic proton efflux rate and oxygen consumption rate measurements. The results of the phenotypic assays were confirmed by evaluating protein levels through Western blot and mRNA expression levels using qRT-PCR. Immunofluorescence staining was performed to determine the subcellular localization of cellular myelocytomatosis oncogene (c-MYC). These in vitro effects of PHY were validated in vivo assay by the peritoneal metastasis of GCa.

Results: Here, we show that PHY reduced migration and invasion of GCa cell lines AGS and TMK1 significantly. At 5 μM, PHY decreased cell motility, energy metabolism via glycolysis, oxidative phosphorylation, epithelial-mesenchymal transition (EMT), as well as inhibiting the Wnt/β-catenin signaling pathway. PHY did not affect motility or energy metabolism in cells transfected with si-MYC, but did in cells overexpressing MYC. Co-treatment with cycloheximide and PHY accelerated the degradation of c-MYC protein. In addition, treatment with bafilomycin A1 or chloroquine plus PHY prevented degradation of c-MYC, while co-treatment with 3-methyl adenine or MG132 plus PHY did not. Furthermore, co-localization of LAMP1 and c-MYC was increased by PHY. These findings suggest that targeting MYC with PHY may suppress energy metabolism and motility in GCa cells by lysosomal degradation. In addition, co-treatment with 5 μM of PHY and 5 μM of 5-fluorouracil (5-FU) has shown a slightly stronger inhibitory effect on cell metastasis and energy metabolism compared to 5 μM of 5-FU treatment alone in GCa cells. These in vitro effects of PHY were validated in vivo. The results showed that 2.5 mg/kg PHY decreased peritoneal metastasis of GCa significantly by suppressing the number of tumor nodules, fluorescence intensity of nodules, target proteins, and altering the composition of metabolites generated by the glycolysis, fatty acid, and tricarboxylic acid cycles.

Conclusions: PHY downregulates the metastasis, mitochondrial respiration, aerobic glycolysis, EMT, and the Wnt/β-catenin signaling pathway of GCa by suppression of c-MYC.

Keywords: Energy metabolism; Gastric cancer; Lysosomal degradation; Physciosporin; c-MYC.