PHGDH knockdown activates autophagic flux to suppress migration and invasion of gastric cancer cells.

Background: Phosphoglycerate dehydrogenase (PHGDH), the key rate-limiting enzyme responsible for controlling the serine biosynthetic pathway, has been implicated in metabolic reprogramming in numerous cancers. Despite this, its significance and regulatory mechanistic relevance in gastric cancer (GC) have yet to be fully elucidated. Here, we sought to elucidate the influence of PHGDH on autophagy and malignant phenotypes in GC and to elucidate its mechanistic relevance in GC progression. Methods: GC tissues and adjacent noncancerous tissues (n=7 pairs) were assessed for PHGDH expression by Western blotting, quantitative real-time polymerase chain reaction (qRT-PCR), and immunohistochemistry. GC cell lines with differential PHGDH expression were screened, and stable knockdown and overexpression models were established. Functional assays, including MTS assays, colony formation, wound healing, and Transwell migration assays, were performed to evaluate proliferation and motility, respectively. Western blotting and autophagic flux assays, as well as analyses of live-cells expressing mRFP-GFP-LC3, were used to evaluate ROS production and autophagy. Results: PHGDH was upregulated in GC tissues (n=7 pairs) with a stage-dependent trend, though larger cohorts are needed to confirm this pattern. Downregulation of PHGDH curtailed MGC803 cell growth, migratory and invasive abilities, and triggered apoptosis, while overexpression enhanced cell motility. Mechanistically, PHGDH silencing increased ROS and HIF-1α levels, reduced mTOR phosphorylation, and facilitated autophagic flux, as demonstrated by elevated LC3-II and reduced P62 expression. These changes were further confirmed by 3-MA/CQ treatment and mRFP-GFP-LC3 imaging. Notably, PHGDH downregulation may indirectly promote the assembly of the ULK1 complex through the concomitant upregulation of FIP200 and ATG101, thereby initiating autophagy. Conclusions: These data suggest that PHGDH plays a role in the progression of GC and may be considered a potential therapeutic target upon further confirmation in larger clinical studies.