Objective To investigate the effect of butorphanol on the proliferation, apoptosis and angiogenesis of lung cancer cells by regulating the sonic hedgehog (SHH)/glioma‑associated oncogene homolog 1 (GLI1) signaling pathway. Methods The effect of 0.5, 1.0, 2.0, 4.0, 8.0, and 16.0 μmol/L butorphanol on the proliferation of lung cancer A549 cells was determined by 2,5‑diphenyl‑2H‑tetrazolium bromide (MTT) assay to determine the optimal intervention concentration. According to the completely random method, lung cancer A549 cells were divided into five groups (6 wells per group): a control group, a cyclopamine group, a butorphanol group, a butorphanol+no⁃loaded plasmid (pcDNA⁃NC) group, and a butorphanol+overexpressed SHH plasmid (pc⁃SHH) group. There was no treatment in the control group, while 10 µmol/L cyclopamine was added to the cyclopamine group, 8 µmol/L of butorphanol was added to the butorphanol group. Meanwhile, 8 µmol/L butorphanol was added to the pcDNA⁃NC group followed by transfection of pcDNA⁃NC; 8 µmol/L butorphanol was added to the butorphanol+pc⁃SHH group followed by transfection of pc⁃SHH. The optical densities at 490 nm D490 (24 h and 48 h), proliferation rate, apoptosis rate, and the levels of SHH messenger RNA (mRNA) and GLI1 mRNA were detected by MTT assay, 5 acetyli⁃2' deoxyuridine (Edu) staining, flow cytometry, and real‑time fluorescence quantitative polymerase chain reaction (FQ‑PCR), respectively. The cell lumen of each group was observed by three‑dimensional culture using a Matrigel matrix. The levels of vascular endothelial growth factor (VEGF⁃A), vascular endothelial cadherin (VE⁃cadherin), B‑cell lymphoma‑2 (Bcl‑2), B‑cell lymphoma‑2‑associated X protein (Bax), SHH, and GLI1 were measured by Western blot. Results Butorphanol at 0.5‒16.0 μmol/L were able to inhibit the proliferation activity of A549 cells in a concentration‑dependent manner, with a half maximal inhibitory concentration of about 8.0 μmol/L . The A549 cells in the control group presented a relatively complete lumen structure. In contrast, the lumen structures of A549 cells in the cyclophosphamide group, the butorphanol group, and the butorphanol+pcDNA‑NC group were incomplete and obviously damaged, while D490 (24 h and 48 h), the proliferation rate, the levels of SHH mRNA and GLI1 mRNA, VEGF‑A, VE cadherin, Bcl‑2, SHH, and GLI1 decreased (all P<0.05), but the apoptotic rate and Bax protein expression increased (all P<0.05). There was no statistical difference in each detection indicators of A549 cells between the cyclophosphamide group and the butorphanol group (all P>0.05). Compared with the butorphanol+pcDNA‑NC group, the luminal structure of A549 cells in the butorphanol+pc‑SHH group was relatively intact, with obviously less damage, while D490 (24 h and 48 h), the proliferation rate, the levels of SHH mRNA and GLI1 mRNA, VEGF‑A, VE cadherin, Bcl‑2, SHH, GLI1 protein increased (all P<0.05), but the apoptotic rate and Bax protein expression decreased (all P<0.05). Conclusion Butorphanol inhibits the proliferation and angiogenesis of lung cancer A549 cells and promotes their apoptosis by blocking the SHH/GLI1 signaling pathway.