PO-074 Metformin-induced apoptosis facilitates degradation of the C-FLIPL protein through a caspase-dependent pathway in human renal cell carcinoma A498 cells

Abstract

IntroductionGastric cancer (GC) is the fourth most common cancer and the third-leading cause of cancer-related deaths worldwide. Despite certain advances in chemotherapy regimens and targeted therapies, the 5 year survival of patients with advanced GC is disappointing. The positive results of the apatinib phase III trial have cast new light on treatment for patients with advanced gastric cancer (GC) refractory to two or more lines of prior systemic chemotherapies. However, a dose reduction or treatment interruption is required to prevent apatinib toxicity such as hand-foot skin reaction (HFSR), proteinuria, and hypertension. In addition, suitable biomarkers are not accessible for identifying the patients who are most likely to benefit from apatinib. Therefore, predictive biomarkers are urgently needed to reduce toxicity and identify patients that will benefit from apatinib treatment.Material and methodsClonogenic assay, cytotoxicity assay, apoptosis assay, qRT-PCR, western blot were conducted in GC cells to evaluate the effectiveness of apatinib in vitro. Confocal microscopy assay was peformed to examine apatinib-induced autophagic flux in GC cells. Transmission electron microscopy assay was conducted to show the formation of autophagic vesicles. Xenograft models in mice were conducted and six tumour tissues were sent for miRNA-seq and circRNA-seq. Sanger sequencing was applied to determine the full length of the amplification products.Results and discussionsIn this study, we found that apatinib inhibited the growth of GC cells and xenograft tumours. Apatinib promoted autophagy activation via upregulation of ATG7 expression, and autophagy inhibition enhanced apatinib-induced apoptosis. With miRNA-seq and circRNA-seq analyses of GC xenograft models, we demonstrated that circRNA004582 functioned as an endogenous sponge for miR-3657 to inhibit its activity and further upregulate ATG7 expression. Silencing of circRNA004582 inhibited apatinib-induced autophagy, which was rescued by miR-3657. Moreover, knockdown of circRNA004582 promoted apatinib-induced apoptosis in GC cells. These findings provided the first evidence that the circRNA004582-miR-3657-ATG7 axis mediates a regulatory pathway critical for the regulation of autophagy and apatinib sensitivity in GC cells and xenografts.ConclusionIn conclusion, specific blockage of circRNA004582 may be a potential therapeutic target for autophagy inhibition in the context of apatinib treatment in human GC. Prospective clinical studies are warranted to examine these biomarkers.