Cyclopiane Suppresses Gastric Cancer Invasion and Angiogenesis via the CYR61/αvβ3 Integrin/NF-κB Pathway.

backgroundCysteine-rich angiogenic inducer 61 (CYR61), a matricellular protein, has been implicated in gastric cancer aggressiveness through activation of integrin-mediated NF-κB signaling; however, effective pharmacological strategies targeting this axis remain limited. Cyclopiane, a bioactive small-molecule compound, has shown potential anti-tumor activity, but its role and underlying mechanisms in gastric cancer have not been fully elucidated. This study aimed to investigate whether Cyclopiane suppresses gastric cancer invasion and angiogenesis via modulation of the CYR61/αvβ3 integrin/NF-κB signaling pathway.

methodsHuman gastric cancer cell lines AGS and MKN45 were treated with increasing concentrations of Cyclopiane. Cell viability was assessed using MTT assays, while invasive capacity was evaluated using Matrigel-coated Transwell assays. TUNEL staining was used to assess changes in apoptosis in tumor cells. The expression of invasion-, angiogenesis-, and inflammation-related proteins was analyzed by Western blot (WB) and enzyme-linked immunosorbent assay (ELISA). Transcriptomic profiling of AGS cells treated with Cyclopiane versus vehicle control was performed to identify differentially expressed genes and enriched signaling pathways. CYR61 overexpression was achieved via adeno-associated virus (AAV-CYR61), with its pathway involvement further validated using integrin αvβ3 antagonists. The conventional anticancer drug 5-fluorouracil (5-FU) was employed to compare Cyclopiane's therapeutic effects. In vivo experiments utilized a nude mouse xenograft model to evaluate changes in tumor growth, angiogenesis, and inflammatory responses following Cyclopiane treatment.

resultsWe found that Cyclopiane significantly reduced cell viability, invasive capacity of AGS and MKN45 cells and promoted apoptosis in a dose-dependent manner, with its high-dose effects comparable to those of 5-FU. Transcriptomic analysis revealed that Cyclopiane downregulated CCN1/CYR61, ITGAV and ITGB3 expression, with pathway enrichment identifying integrin signaling, ECM-receptor interaction, focal adhesion and NF-κB pathways as the top modulated terms. In vivo experiments demonstrated that Cyclopiane significantly inhibited tumor growth and progression. Both in vivo and in vitro studies showed Cyclopiane significantly downregulated vascular endothelial growth factor (VEGF), matrix metalloproteinase-2 (MMP-2), and MMP-9 expression while inhibiting integrin αvβ3 expression and NF-κB activation, as evidenced by reduced phosphorylation levels of p65 and IκBα. Overexpression of CYR61 partially reversed the inhibition of invasion, angiogenic factor production, and NF-κB signaling mediated by Cyclopiane. The αvβ3/αvβ5 integrin antagonist Cilengitide also exhibited tumor-suppressive effects, and the co-treatment of Cyclopiane with Cilengitide achieved the strongest tumor-suppressive effect.

conclusionThese findings suggest that modulation of the CYR61/integrin/NF-κB signaling axis contributes, at least in part, to the anti-invasive and anti-angiogenic effects of Cyclopiane in preclinical gastric cancer models. Further studies are required to establish its molecular target specificity, safety, selectivity, and translational potential.