AMPK-independent autophagy promotes radioresistance of human tumor cells under clinical relevant hypoxia in vitro

作者：Chaachouay Hassan; Fehrenbacher Birgit; Toulany Mahrnoud; Schaller Martin; Multhoff Gabriele; Rodemann H Peter^*

来源：Radiotherapy and Oncology, 2015, 116(3): 409-416.

DOI：10.1016/j.radonc.2015.08.012

• Clinical relevant hypoxia
• Autophagy
• Ionizing radiation
• Solid tumor cells

Background and purpose: Blocking of the autophagy-signaling has the potential to improve cancer therapy. In the present study, the role of autophagy for radioresistance of human tumor cells was tested under clinically relevant hypoxia (1% O-2).
Materials and methods: Non-small cell lung cancer cell lines A549 and H460, head and neck squamous cell carcinoma FaDu, colon carcinoma cell line HCT116 and mouse-embryo-fibroblasts were analyzed under normoxic (21% O-2) and hypoxic (0.01% and 1% O-2) conditions with respect to clonogenic cell survival and hypoxia-induced autophagy. Immunofluorescence and electron microscopy were used to monitor the autophagy process and Western blotting of LC3, AMPK, and BNIP3 was applied to analyze autophagy signaling.
Results: Clinically relevant hypoxia stimulated autophagy in tumor cells as indicated by enhanced LC3-I to LC3-II conversion. Furthermore, hypoxia stimulated autophagy was approved by Immunofluorescence staining and electron-microscopy analysis of autophagosome vacuoles. Preconditioning of tumor cells to moderate-hypoxia increased their radioresistance that was significantly reversed following pretreatment with autophagy inhibitor, chloroquine. Using siRNA against AMPK as well as AMPK deficient cells, autophagy stimulation by 1% O-2 was shown to be AMPK-independent. However, a correlation between the expression of BNIP3 and autophagy-stimulation was observed under this condition.
Conclusion: Under clinically relevant hypoxia (1% O-2) the stimulation of autophagy mediates resistance of hypoxic tumor cells to ionizing radiation, which is independent of AMPK signaling.

• 出版日期2015-9