tRF-17-9L5FZU3 is a novel prognostic biomarker and therapeutic target for esophageal squamous cell carcinoma: mechanisms involving RhoB suppression.

tRNA-derived fragments (tRFs) have arisen as critical regulators in cancer development, however, their functions in esophageal squamous cell carcinoma (ESCC) remain poorly understood. Here, we reported that tRF-17-9L5FZU3 was significantly upregulated in ESCC cells and tissues, and correlated with poor patient outcome. Through integrated small RNA sequencing and qRT-PCR validation in 40 paired clinical samples, functional analyses, including colony formation, CCK-8, Transwell migration/invasion assays, and xenograft mouse models, demonstrated that tRF-17-9L5FZU3-knockdown suppresses ESCC cell proliferation, invasion, migration, and tumor growth in vivo. Furthermore, tRF-17-9L5FZU3 was shown to directly bind the 3'UTR of RhoB (Ras Homolog Family Member B) mRNA via dual-luciferase reporter assays, RNA pull-down, and AGO2 co-immunoprecipitation, leading to repression of RhoB expression at both mRNA and protein levels. RhoB was consistently downregulated in ESCC tumors and exhibited a significant inverse correlation with tRF-17-9L5FZU3 expression. Rescue experiments confirmed that oncogenic effects of tRF-17-9L5FZU3 were dependent on RhoB suppression. Moreover, tRF-17-9L5FZU3-knockdown decreased TGF-β secretion, increased IL-2 production, and inhibited PI3K/AKT and NF-κB signaling, which were reversed by co-silencing of RhoB. Collectively, these findings found that tRF-17-9L5FZU3 promoted ESCC malignancy by directly repressing RhoB, thereby activating oncogenic signaling pathways, positioning the tRF-17-9L5FZU3-RhoB axis as a critical regulator and potential therapeutic target in ESCC.