Recently, several miRNAs have been revealed to play critical roles in oncogenesis and tumor progression of many cancers. Thioredoxin-1 (Trx-1) binding protein-2 (TBP-2) is an internal inhibitor of Trx-1, which plays the role in regulating oxidative stress, inhibiting cell growth, and promoting apoptosis. The expression of TBP-2 is usually decreased in cancer tissues. However, whether the miRNAs regulate the TBP-2 expression in lung cancer is still unclear. In this study, we examined the levels of TBP-2, miR-93, miR-373, and miR-17-5p in lung cancer tissues and their adjacent normal lung tissues of 36 patients. We found that the expressions of miR-93, miR-373, and miR-17-5p were higher, whereas the expression of TBP-2 mRNA and protein was significantly lower in lung cancer tissues compared with adjacent normal lung tissues. After the three miRNA mimics were transfected in the lung cancer cells, NCI-H460, the level of TBP-2 mRNA and TBP-2 protein was decreased. Then, the anti-cancer drug 5-fluorouracil was used to stimulate the NCI-H460 cells; the mRNA levels of miR-93, miR-373, and miR-17-5p were decreased, and the level of TBP-2 mRNA and protein was increased. Collectively, the above results suggest that miR-93, miR-373, and miR-17-5p negatively regulate the TBP-2 expression in lung cancer. This study may provide therapeutic targets with lung cancer. © Copyright © 2020 Li, Liang, Zhang, Yuan, Gao, Shi and Bai.