title: Enhanced radiation damage of tumor vasculature by mTOR inhibitors.
abstract: It is known that radiation activates the phosphoinositol-3 kinase (PI3K)/Akt pathway and that inhibition of PI3K or Akt sensitizes tumor vasculature to radiotherapy. Mammalian target of rapamycin (mTOR) is a downstream target of Akt, and we hypothesized that irradiation activates mTOR signaling in both glioma and endothelial cells (ECs) and that radiosensitization results from inhibiting mTOR signaling. mTOR inhibitors, rapamycin and RAD001 (everolimus) were found to radiosensitize vascular ECs, but failed to sensitize glioma cells as determined by clonogenic assay. Therefore, we investigated the anti-angiogenic effects of mTOR inhibitors. Increased phospho-mTOR protein was detected in irradiated human umbilical vein endothelial cells (HUVEC), but not in GL261 glioma cells. Phospho-S6, a biomarker for mTOR signaling, was also found to be induced following irradiation in HUVEC and this effect was inhibited by PI3K or mTOR inhibitors. Significant increase in cleaved caspase 3 was detected when Rad001 was combined with radiation. Endothelial tube formation was significantly diminished following treatment with rapamycin and 3 Gy of radiation. Histological sections of GL261 tumors from mice showed a greatly reduced vascular density when treated with RAD001 and radiation. Power Weighted Doppler of glioma xenografts in mice showed a significant reduction in vasculature and blood flow compared with mice treated with 3 Gy or RAD001 alone. We conclude that irradiation activates mTOR signaling in vascular endothelium and that rapamycin and RAD001 increased apoptosis of ECs in response to radiation. To the authors' best knowledge this is the first study which demonstrates that mTOR inhibitors may be a way to target the vasculature by radiosensitizing the vascular endothelium resulting in better tumor control as seen in experiments demonstrating increased tumor growth delay in mice treated with rapamycin with radiation compared with mice treat with either treatment alone. We conclude that mTOR inhibitors have increased efficacy as antiangiogenics when combined with radiation.
authors.affiliation: Department of Radiation Oncology, Vanderbilt Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN 37232-5671, USA.
authors.forename: Eric T
authors.initials: ET
authors.lastname: Shinohara
authors.forename: Carolyn
authors.initials: C
authors.lastname: Cao
authors.forename: Ken
authors.initials: K
authors.lastname: Niermann
authors.forename: Yi
authors.initials: Y
authors.lastname: Mu
authors.forename: Fenghua
authors.initials: F
authors.lastname: Zeng
authors.forename: Dennis E
authors.initials: DE
authors.lastname: Hallahan
authors.forename: Bo
authors.initials: B
authors.lastname: Lu
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D015153:Blotting, Western; D045744:Cell Line, Tumor; D042783:Endothelial Cells / Q000187:drug effects / Q000528:radiation effects; D000068338:Everolimus; D005910:Glioma / Q000098:blood supply* / Q000188:drug therapy / Q000532:radiotherapy; D006801:Humans; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D011494:Protein Kinases / Q000187:drug effects / Q000378:metabolism*; D011830:Radiation Effects; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D020123:Sirolimus / Q000031:analogs & derivatives* / Q000494:pharmacology*; D058570:TOR Serine-Threonine Kinases; D018608:Ultrasonography, Doppler; D014841:von Willebrand Factor / Q000378:metabolism
chemical_list: D020533:Angiogenesis Inhibitors; D000068338:Everolimus; D019869:Phosphatidylinositol 3-Kinases; D011494:Protein Kinases; D011838:Radiation-Sensitizing Agents; D020123:Sirolimus; D058570:TOR Serine-Threonine Kinases; D014841:von Willebrand Factor; C546842:MTOR protein, human; C546843:mTOR protein, mouse