Abstract Background: Mitochondria are essential organelles responsible for ATP generation and play pivotal roles in diverse cellular processes. Mitochondrial dysfunction contributes to various pathological conditions, including cancer development. As the central organ for nutrient metabolism, the liver contains a particularly high density of mitochondria. Impaired mitochondrial function leads to excessive generation of reactive oxygen species (ROS) and oxidative stress, resulting in cellular damage closely associated with hepatocellular carcinoma (HCC). Cancer cells frequently undergo metabolic reprogramming to sustain their high energy demands during rapid proliferation. Given our previous findings identifying DDX3 as a tumor suppressor in HCC, this study aims to elucidate the effects of DDX3 on mitochondrial function and redox regulation, as well as its implications for HCC tumorigenesis. Methods: Mitochondrial morphology in DDX3-deregulated HCC cells was analyzed using super-resolution confocal microscopy. The expression of mitochondrial dynamics-related proteins was assessed by immunoblotting and quantitative real-time PCR. Metabolic activity was examined by MitoTracker Red/Green staining and Seahorse extracellular flux analysis. Redox homeostasis was evaluated by MitoSOX and DCFH-DA staining, along with measurements of the GSH/GSSG ratio and NADPH/NADP+ levels. DDX3-regulated cellular functions were investigated through proliferation, colony formation, and transwell migration/invasion assays, with or without metabolic inhibitors. Cancer stemness was assessed using sphere-formation, chemoresistance, and flow cytometry-based assays. Results: Reduced DDX3 expression in HCC cells led to marked alterations in mitochondrial morphology and increased mitochondrial oxidative phosphorylation (OXPHOS), resulting in elevated mitochondrial superoxide accumulation. Unexpectedly, the total cellular ROS levels were decreased in DDX3-knockdown cells. Further analyses revealed that low DDX3 expression enhances the cellular capacity to maintain redox balance, conferring a more robust antioxidant defense. Additionally, HCC cells with reduced DDX3 expression exhibited greater proliferation and self-renewal potential under varying nutrient conditions, effects that were attenuated upon treatment with metabolic inhibitors. Conclusion: Our results suggested that DDX3 may suppress HCC progression by inhibiting mitochondrial activity, thereby reinforcing its role as a tumor suppressor in HCC. Citation Format: Jun-Huan Huang, Heng-Yu Lin, Yi-Yuan Su, RU-TSUN MAI. DDX3 modulates mitochondrial function to inhibit HCC progression abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 596.
Huang et al. (Fri,) studied this question.