Objective:To explore the effects of Qidongning Formula(QDF) on lung cancer cells proliferation by regulating autophagy. Methods①The effects of different concentrations of QDF on the survival rate of A549 and NCI-H460 were detected by CCK8 assay, and the effects on the clone formation ability of A549 and NCI-H460 by clone formation assay. The effects of QDF on the expression of autophagy-associated proteins LC3B and p62 in A549 and NCI-H460 cells were detected by Western blot method, and the effect of QDF on the expression of LC3B was detected by immunofluorescence. The survival rates and colony formation ability of A549 and NCI-H460 cells under the inhibition of autophagy were detected by CCK8 assay and clone formation assay.②The subcutaneous transplanted tumor of NCI-H460 in nude mice was used as a model to observe the inhibitory effect of QDF on the tumor, and the expression of autophagy-related proteins LC3B and p62 in each group was detected by Western blot method.  Results: The experiment in vitro showed that QDF could inhibit the proliferation of A549 and NCI-H460 cells. Under the condition of autophagy inhibition, the ability decreased of QDF against A549 and NCI-H460 cells proliferation. Western blot results showed that QDF could up-regulate the expression of LC3B and down-regulate the expression of p62 in A549 and NCI-H460 cells. The immunofluorescence results showed that QDF could concentrate the dispersed LC3B in the form of speckled. Animal experiments showed that QDF could inhibit the growth of NCI-H460 subcutaneously transplanted tumor in nude mice, up-regulate the expression of LC3B and down-regulate the expression of p62.  Conclusion:sQDF could suppress lung cancer cells proliferation by regulating autophagy.