Abstract C098: Overexpression of glycoprotein non-metastatic melanoma protein B (GPNMB) in esophageal cancer cells is mediated through a miR-29a-3p-TFEB axis

Abstract Introduction: Glycoprotein non-metastatic melanoma protein B (GPNMB) is a transmembrane protein that is expressed at low levels in normal tissues but is highly expressed and associated with poor prognoses in aggressive cancers. Recent reports have suggested that GPNMB expression is increased in esophageal squamous cell cancer cells, but its expression in esophageal adenocarcinoma cells is not well defined. The GPNMB promoter contains highly conserved binding sequences for the microphthalmia (MiT/TFE) family of transcription factors including TFEB, which is a potent driver of GPNMB expression. Based on sequence analysis, miR-29a-3p is predicted to bind TFEB mRNA with high affinity. The purpose of this study is to characterize the expression of GPNMB, TFEB, and miR-29a-3p in esophageal cancer cell lines and define the regulation of GPNMB by miR-29a-3p and TFEB in these cells. Methods: Studies were conducted in human esophageal epithelial cells (Het1A) and in TE7, TE10, OE21 esophageal squamous cancer cell lines, as well as in the esophageal adenocarcinoma cell lines OE33, OE-19, and SKGT4. Global microRNA expression in these cell lines was determined by microarray analysis and confirmed by real-time PCR. MiR-29a-3p function was tested by transient overexpression. Protein expression was determined by Western blot and immunohistochemistry. Levels of mRNA were measured by real-time and droplet digital PCR. Results: Endogenous GPNMB mRNA and protein expression were increased in all three esophageal squamous cancer cell lines TE7, TE10, OE21, and in an esophageal adenocarcinoma cell line, OE33, compared to Het1A cells. GPNMB expression in these cells was preferentially localized to the cell membrane. MiR-29a-3p expression in these cells was markedly downregulated in comparison to Het1A cells. Basal levels of TFEB mRNA and protein were also increased in these esophageal cancer cell lines compared to Het1A cells. In addition, TFEB mRNA expression was preferentially localized to the nuclear fraction in these cells. TFEB protein and mRNA levels were reduced in a time-dependent manner following miR-29a-3p overexpression these cells. GPNMB mRNA and protein showed similarly decreased expression in these cells following miR-29a-3p overexpression. Conclusion: GPNMB and TFEB expression are increased in several esophageal squamous cell cancer cell lines, as well as in an esophageal adenocarcinoma cell line, compared to esophageal epithelial cells. Conversely, miR-29a-3p expression is markedly reduced in these esophageal cancer cells relative to Het1A cells. Overexpression of miR-29a-3p in the esophageal cancer cells results in decreased expression of TFEB and GPNMB. Further studies on the function of GPNMB in esophageal cancer cells and the effects of modulating it expression in these cells is warranted. Citation Format: Pornima Phatak, Kaushal Asrani, Tamara Lotan, James M. Donahue. Overexpression of glycoprotein non-metastatic melanoma protein B (GPNMB) in esophageal cancer cells is mediated through a miR-29a-3p-TFEB axis [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr C098.