The specific pathogenesis of acute lung injury (ALI) is complex and not yet clear, and the clinical treatment methods are relatively limited. It is of great clinical significance to explore its pathogenesis and effective molecular targets. Here, we identified an ALI biomarker (UPP1) associated with uridine metabolism by a systematic bioinformatics approach. It was also confirmed to be associated with the glycolytic pathway in the mouse ALI model. In addition, drug sensitivity analysis based on the CMAP database identified three UPP1-associated drugs (CAY10585, XL147 and IOX2) that may be useful in the treatment of ALI. Molecular docking and molecular dynamics simulations further confirmed the stability of the binding between UPP1 and the three drugs. In conclusion, this study confirms that the uridine metabolism gene UPP1 associated with glycolysis is a key biomarker of ALI and provides valuable insights into the potential application of CAY10585, XL147 and IOX2 in ALI.