Abstract 5808: Diclofenac inhibits esophageal cancer cell growth by depleting mitochondrial functions

Abstract

Abstract Esophageal cancer is one of the most aggressive forms of human malignancy with a 5-yearsurvival rate of less than 20%. Mitochondria are dynamic organelles that play essential roles in various cellular processes, including energy metabolism, redox homeostasis, and apoptotic cell death. Alterations in mitochondrial biology are associated with esophageal carcinogenesis and esophageal cancer cell response to therapy. As the anti-inflammatory drug diclofenac (DCF)induces mitochondrial dysfunction, we hypothesized that DCF may inhibit esophageal carcinogenesis by affecting mitochondrial processes. In the human esophageal squamous cell carcinoma (ESCC) cell line TE11, we performed MTT assay and Annexin-V/Propidium Iodide (PI) to evaluate the impact of DCF on proliferation and cell death, respectively. RNA sequencing and Ingenuity Pathway Analysis (IPA) identified differentially expressed genes (DEGs) and altered canonical pathways in DCF-treated TE11 cells. To assess metabolic function, we measured the levels of pyruvate, lactate, and ATP. Mitochondrial membrane potential and ROS production were assessed by flow cytometry for MitoTracker red/green and MitoSox red, respectively. Mice bearing subcutaneous syngeneic ESCC tumors were treated with DCF.DCF inhibited proliferation of TE11 in dose dependent manner with an IC50 concentration of 76.7µM. By contrast, the IC50 concentration of DCF in normal esophageal keratinocytes was 876.2µM, supporting selectivity of DCF for esophageal cancer cells. In TE11, 200 µM DCF induced apoptosis in 60% of cells. RNA-sequencing identified that DCF significantly altered expression of3287 genes (FC ≥ 1.50) in TE11, including the top cancer-associated genes: PLK1, MCM2, MCM3,MCM7, MCM10 and SKP2. IPA analysis revealed that DCF activated p53 signaling while inhibited Gluconeogenesis I, Glycolysis I and Oxidative Phosphorylation. DCF significantly inhibited glycolysis as demonstrated by reduction in concentrations of pyruvate (1.76-fold) and lactate (7.30-fold) and downregulation of the glycolysis- associated genes PKM2, PFKM, LDHA, and PKM1. A decreased production of ATP (5-fold) was also noted in DCF-treated TE11 cells. In addition, DCF depleted mitochondrial membrane potential (2.5-fold) and increased mitochondrial ROS production (5-fold). A reduction in ROS by mitochondrial antioxidant MitoTempo increased viability of the DCF-treated TE11 cells, indicating that ROS contributes to the anticancer activity of DCF. Consistent with antitumor activity of DCF in TE11, the drug significantly decreased the tumor volume in syngeneic ESCC tumors in vivo. Our preclinical findings indicate that DCF may limit esophageal cancer cell growth through the inhibition of mitochondrial functions, identifying a novel experimental therapeutic for ESCC. Future studies will define the precise molecular mechanisms through which DCF promotes cell death in vitro and in vivo. Citation Format: Mohammad Faujul Kabir, Adam Karami, Anbin Mu, Don-Gerard Conde, Kelly A. Whelan. Diclofenac inhibits esophageal cancer cell growth by depleting mitochondrial functions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5808.