title: Ubenimex enhances the radiosensitivity of renal cell carcinoma cells by inducing autophagic cell death.
abstract: Renal cell carcinoma (RCC) is resistant to standard radiotherapy. Ubenimex, an aminopeptidase N inhibitor, is widely used as an adjunct therapy after surgery to enhance the function of immunocompetent cells and confer antitumor effects. Our previous study demonstrated that ubenimex induces autophagic cell death in RCC cells. Recently, the molecular mechanism of autophagy induction has been associated with radiosensitivity in RCC cells. In the present study, the ability of ubenimex to enhance RCC cell sensitivity to radiation via the induction of autophagic cell death was determined, and the mechanism of action of this effect was investigated. The 786-O and OS-RC-2 human RCC cell lines were treated with 0.5 mg/ml ubenimex and different doses of irradiation (IR). The cell viability was measured using a colony-formation assay and flow cytometry. Acridine orange (AO)-ethidium bromide (EB) staining was assessed by fluorescence microscopy as an indicator of autophagic cell death. Protein expression was assessed by western blotting. Autophagosomes were evaluated using transmission electron microscopy. RCC cells were used to evaluate the sensitivity to radiation using clonogenic survival and lactate dehydrogenase assays. Furthermore, these parameters were also tested at physiological oxygen levels. The AO-EB staining and flow cytometry of the OS-RC-2 cells indicated that the combined treatment significantly enhanced autophagic cell death compared with ubenimex or IR alone. Therefore, treatment with ubenimex did not significantly alter cell cycle progression but increased cell death when combined with radiation. An Akt agonist could significantly weaken this effect, indicating that ubenimex may act as an Akt inhibitor. Furthermore, the western blot analysis indicated that the combined treatment inhibited the Akt signaling pathway compared with ubenimex treatment or IR alone. Ubenimex may enhance RCC cell sensitivity to radiation by inducing cell autophagy. This induction changes the role of autophagy from protective to lethalin vitro, and this switch is associated with the inhibition of the Akt signaling pathway.
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Liu
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Xiaoqing
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Jiaju
authors.initials: J
authors.lastname: Lu
authors.affiliation: Department of Neurology, Shandong Police Hospital, Jinan, Shandong 250021, P.R. China.
authors.forename: Liping
authors.initials: L
authors.lastname: Han
authors.affiliation: Department of Dermatology, Shandong University, Jinan, Shandong 250000, P.R. China.
authors.forename: Yongfei
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Sentai
authors.initials: S
authors.lastname: Ding
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Zhao
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Dongbin
authors.initials: D
authors.lastname: Bi
authors.affiliation: Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
authors.forename: Zhihong
authors.initials: Z
authors.lastname: Niu
keywords: Akt; RCC; autophagic cell death; radiosensitivity; ubenimex