Objective: To explore the effects and mechanism of pillyrin on apoptosis of nasopharyngeal carcinoma (NPC) CNE-1 and CNE-2 cells by regulating miR-34c-5p. Methods: CNE-1 and CNE-2 cells were cultured. MTT assay was used to screen the effect of pillyrin on the proliferation activity of NPC cells. CNE-1 and CNE-2 cells were assigned into control group, pillyrin group, miR-NC, miR-34c-5p group, pillyrin+in-miR-NC group, and pillyrin+in-miR-34c-5p group, respectively. Colony formation, EdU, TUNEL, flow cytometry, and Western blot were used to detect the effects of pillyrin on the proliferation and apoptosis of NPC cells. Results: The proliferation activity of CNE-1 and CNE-2 cells decreased with the increase of pillyrin concentration (P<0.05). At 8 μmol/L pillyrin, the inhibitory ability of CNE-1 and CNE-2 cell proliferation activity was close to 50%. For the control group, the pillyrin group showed an increase in miR-34c-5p, TUNEL positive cell rate, apoptosis rate, p53, and FasL in CNE-1 and CNE-2 cells, and a decrease in colony formation, EdU positive cell ratio, c-Myc, PCNA, and Survivin (P<0.05). For the miR-NC group, the miR-34c-5p group showed an increase in miR-34c-5p, TUNEL positive cell rate, apoptosis rate, p53, and FasL in CNE-1 and CNE-2 cells, and a decrease in colony formation, EdU positive cell ratio, c-Myc, PCNA, and Survivin (P<0.05). For the pillyrin+in-miR-NC group, the pillyrin+in-miR-34c-5p group showed a decrease in miR-34c-5p, TUNEL positive cell rate, apoptosis rate, p53, and FasL in CNE-1 and CNE-2 cells, and an increase in colony formation, EdU positive cell ratio, c-Myc, PCNA, and Survivin (P<0.05). Conclusion: Pillyrin upregulates miR-34c-5p to inhibit the apoptosis of CNE-1 and CNE-2 in NPC cells.