title: GS-nitroxide (JP4-039)-mediated radioprotection of human Fanconi anemia cell lines.
abstract: Fanconi anemia (FA) is an inherited disorder characterized by defective DNA repair and cellular sensitivity to DNA crosslinking agents. Clinically, FA is associated with high risk for marrow failure, leukemia and head and neck squamous cell carcinoma (HNSCC). Radiosensitivity in FA patients compromises the use of total-body irradiation for hematopoietic stem cell transplantation and radiation therapy for HNSCC. A radioprotector for the surrounding tissue would therefore be very valuable during radiotherapy for HNSCC. Clonogenic radiation survival curves were determined for pre- or postirradiation treatment with the parent nitroxide Tempol or JP4-039 in cells of four FA patient-derived cell lines and two transgene-corrected subclonal lines. FancG(-/-) (PD326) and FancD2(-/-) (PD20F) patient lines were more sensitive to the DNA crosslinking agent mitomycin C (MMC) than their transgene-restored subclonal cell lines (both P < 0.0001). FancD2(-/-) cells were more radiosensitive than the transgene restored subclonal cell line (ñ = 2.0 ± 0.7 and 4.7 ± 2.2, respectively, P = 0.03). In contrast, FancG(-/-) cells were radioresistant relative to the transgene-restored subclonal cell line (ñ = 9.4 ± 1.5 and 2.2 ± 05, respectively, P = 0.001). DNA strand breaks measured by the comet assay correlated with radiosensitivity. Cell lines from a Fanc-C and Fanc-A patients showed radiosensitivity similar to that of Fanc-D2(-/-) cells. A fluorophore-tagged JP4-039 (BODIPY-FL) analog targeted the mitochondria of the cell lines. Preirradiation or postirradiation treatment with JP4-039 at a lower concentration than Tempol significantly increased the radioresistance and stabilized the antioxidant stores of all cell lines. Tempol increased the toxicity of MMC in FancD2(-/-) cells. These data provide support for the potential clinical use of JP4-039 for normal tissue radioprotection during chemoradiotherapy in FA patients.
authors.affiliation: Department of Radiation Oncology, University of Pittsburgh Cancer Institute, Pittsburgh, Pennsylvania 15232, USA.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Bernard
authors.forename: Hyun
authors.initials: H
authors.lastname: Kim
authors.forename: Hebist
authors.initials: H
authors.lastname: Berhane
authors.forename: Michael W
authors.initials: MW
authors.lastname: Epperly
authors.forename: Darcy
authors.initials: D
authors.lastname: Franicola
authors.forename: Xichen
authors.initials: X
authors.lastname: Zhang
authors.forename: Frank
authors.initials: F
authors.lastname: Houghton
authors.forename: Donna
authors.initials: D
authors.lastname: Shields
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Bakkenist
authors.forename: Marie-Celine
authors.initials: MC
authors.lastname: Frantz
authors.forename: Erin M
authors.initials: EM
authors.lastname: Forbeck
authors.forename: Julie P
authors.initials: JP
authors.lastname: Goff
authors.forename: Peter
authors.initials: P
authors.lastname: Wipf
authors.forename: Joel S
authors.initials: JS
authors.lastname: Greenberger
mesh_terms: D001692:Biological Transport; D002460:Cell Line; D002999:Clone Cells; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects / Q000528:radiation effects; D005199:Fanconi Anemia / Q000473:pathology*; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000172:deficiency; D052241:Fanconi Anemia Complementation Group G Protein / Q000172:deficiency; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D016685:Mitomycin / Q000494:pharmacology; D009589:Nitrogen Oxides / Q000378:metabolism / Q000494:pharmacology*; D011837:Radiation-Protective Agents / Q000378:metabolism / Q000494:pharmacology*; D019076:Transgenes / Q000235:genetics
chemical_list: D052236:Fanconi Anemia Complementation Group D2 Protein; D052241:Fanconi Anemia Complementation Group G Protein; C544775:JP4-039; D009589:Nitrogen Oxides; D011837:Radiation-Protective Agents; D016685:Mitomycin; D005978:Glutathione