title: Inhibition of autophagy as a strategy to augment radiosensitization by the dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor NVP-BEZ235.
abstract: We investigated the effect of 2-methyl-2-{4-[3-methyl-2-oxo-8-(quinolin-3-yl)-2,3-dihydro-1H-imidazo[4,5-c]quinolin-1-yl]phenyl} propanenitrile (NVP-BEZ235) (Novartis, Basel Switzerland), a dual phosphatidylinositol 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) inhibitor currently being tested in phase I clinical trials, in radiosensitization. NVP-BEZ235 radiosensitized a variety of cancer cell lines, including SQ20B head and neck carcinoma cells and U251 glioblastoma cells. NVP-BEZ235 also increased in vivo radiation response in SQ20B xenografts. Knockdown of Akt1, p110α, or mTOR resulted in radiosensitization, but not to the same degree as with NVP-BEZ235. NVP-BEZ235 interfered with DNA damage repair after radiation as measured by the CometAssay and resolution of phosphorylated H2A histone family member X foci. NVP-BEZ235 abrogated the radiation-induced phosphorylation of both DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and ataxia telangiectasia mutated. Knockdown of either p110α or mTOR failed to decrease the phosphorylation of DNA-PKcs, suggesting that the effect of the drug was direct rather than mediated via p110α or mTOR. The treatment of cells with NVP-BEZ235 also promoted autophagy. To assess the importance of this process in radiosensitization, we used the autophagy inhibitors 3-methyladenine and chloroquine and found that either drug increased cell killing after NVP-BEZ235 treatment and radiation. Knocking down the essential autophagy proteins autophagy related 5 (ATG5) and beclin1 increased NVP-BEZ235-mediated radiosensitization. Furthermore, NVP-BEZ235 radiosensitized autophagy-deficient ATG5(-/-) fibroblasts to a greater extent than ATG5(+/+) cells. We conclude that NVP-BEZ235 radiosensitizes cells and induces autophagy by apparently distinct mechanisms. Inhibiting autophagy via pharmacologic or genetic means increases radiation killing after NVP-BEZ235 treatment; hence, autophagy seems to be cytoprotective in this situation. Our data offer a rationale for combining NVP-BEZ235 along with an autophagy inhibitor (i.e., chloroquine) and radiation in future clinical trials.
authors.affiliation: Department of Radiation Oncology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors.forename: George J
authors.initials: GJ
authors.lastname: Cerniglia
authors.forename: Jayashree
authors.initials: J
authors.lastname: Karar
authors.forename: Sonia
authors.initials: S
authors.lastname: Tyagi
authors.forename: Melpo
authors.initials: M
authors.lastname: Christofidou-Solomidou
authors.forename: Ramesh
authors.initials: R
authors.lastname: Rengan
authors.forename: Constantinos
authors.initials: C
authors.lastname: Koumenis
authors.forename: Amit
authors.initials: A
authors.lastname: Maity
mesh_terms: D000818:Animals; D001343:Autophagy / Q000187:drug effects* / Q000235:genetics / Q000528:radiation effects*; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy / Q000473:pathology / Q000532:radiotherapy; D045744:Cell Line, Tumor; D016922:Cellular Senescence / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D004249:DNA Damage; D015536:Down-Regulation / Q000187:drug effects; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D005909:Glioblastoma / Q000188:drug therapy / Q000473:pathology / Q000532:radiotherapy; D006258:Head and Neck Neoplasms / Q000188:drug therapy / Q000473:pathology / Q000532:radiotherapy; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology*; D051379:Mice; D008819:Mice, Nude; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011804:Quinolines / Q000494:pharmacology*; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays
chemical_list: D007093:Imidazoles; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011804:Quinolines; D011838:Radiation-Sensitizing Agents; D058570:TOR Serine-Threonine Kinases; C531198:dactolisib