Rationale: Lung cancer is the most common cause of cancer-related death in the United States and worldwide. Despite decades of efforts on research and smoking cessation, the 5-year survival rate of lung cancer patients remains poor at 15%, and the population of adenocarcinoma cases in nonsmokers is rising. PDLIM5, a member of the Enigma subfamily of PDZ and LIM domain protein family, contains an N-terminal PDZ domain and 3 LIM domains at its C-terminus. We have previously shown that PDLIM5 regulates TGF-β/Smad family, which are known to participate in the pathogenesis of lung cancer. In this study, we aim to investigate whether PDLIM5 plays a role in lung cancer and establish a high throughput screening (HTS) platform for PDLIM5-targeted drug discovery. Methods: We searched the Oncomine data base for PDLIM5 gene expression. We suppressed PDLIM5 in A549 cells and determined the expression levels of epithelial-mesenchymal transition markers. We exposed A549 cells to hypoxia or treated them with TGF-β1 and measured the expression levels of PDLIM5. We generated a stable mink lung epithelial cell line (MLEC) containing a TGFβ/Smad luciferase reporter with lentivirus-mediated suppression of PDLIM5 (MLEC-shPDLIM5) and measured levels of Smad2/3 and pSmad2/3. We used MLEC-shPDLIM5 and a control cell line (MLEC-shCTL) to screen the Prestwick library (1,200 compounds). We treated MLEC and A549 cells with paclitaxel and determined levels of Smad2/3 and pSmad2/3. Results: We found that PDLIM5 was overexpressed in non-small-cell lung carcinoma (NSCLC) patients and that the expression levels of PDLIM5 inversely correlated with the survival rate of lung cancer patients. Hypoxia but not TGF-β1 induced PDLIM5 expression levels. Suppression of PDLIM5 induced E-cadherin levels while decreased levels of vimentin and α-SMA. In MLEC, suppression of PDLIM5 decreased Smad-dependent luciferase activity, smad3, and pSmad2. More importantly, we identified and validated that paclitaxel as a PDLIM5 inhibitor in MLEC. Furthermore, we showed that paclitaxel inhibited Smad2 expression and Smad3 phosphorylation in A549 cells. Conclusions: We have shown that PDLIM5 is overexpressed in lung cancer cells and its upregulation is hypoxia-dependent but TGF-β independent. Loss of PDLIM5 inhibits EMT. We have identified paclitaxel as a PDLIM5 inhibitor. This system is robust and suitable for PDLIM5 targeted drug discovery. Support: a UIC UICentre Arm1 Inception Grant, a Gilead Sciences Research Scholars Program in Pulmonary Arterial Hypertension award and R01 HL123804 (G. Zhou). Citation Format: Deborah Park, Han Cheng, Tianji Chen, Merve Tor, Nour Khatib, Jason Huang, Qiyuan Zhou, Lijun Rong, Guofei Zhou. Targeting PDLIM5 for lung cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3785.