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Background/Aim: We tested JP4-039, a GS-nitroxide radiation damage mitigator in proton therapy of Fanconi anemia (FA) mice. Materials and Methods: Fanca(–/–) and Fanca(+/+) bone marrow stromal cells were pre-treated with JP4-039 and irradiated with either protons or photons (0-10 GyRBE) followed by clonogenic survival and β-Galactosidase senescence analysis. Fanca(–/–) and Fanca(+/+) mice were pretreated with JP4-039 for 10 min prior to oropharyngeal irradiation with either protons or photons (0 or 30 GyRBE) followed by sacrifice and measurement of oral cavity ulceration, distant hematopoietic suppression, and real-time polymerase chain reaction analysis. Results: JP4-039 reduced oral cavity ulceration in Fanca(–/–) mice, transcripts Nfkb, Ap1, Sp1, and Nrf2, and proton therapy induced distant marrow suppression. Conclusion: JP4-039 protected Fanca(–/–) and Fanca(+/+) cells and mouse oral cavity from both proton and photon radiation.

We quantitated age-related accumulation of senescent cells in irradiated Fanconi anemia (FA) (Fanca(−/−)) mouse cell lines in vitro and monitored the effect of continuous administration (via drinking water) of the water-soluble radiation mitigator, MMS350, on tissues in vivo over one year after 7.5 Gy total-body irradiation (TBI). Irradiated Fanca(−/−) mouse bone marrow stromal cell lines showed increased numbers of beta-galactosidase- and p21-positive senescent cells compared to Fanca(+/+) cell lines, which was reduced by MMS350. One week after 7.5 Gy TBI, Fanca(−/−) mice showed increased numbers of senescent cells in spleen compared to Fanca(+/+) controls, decreased bone marrow cellularity, failure of explanted bone marrow to proliferate in vitro to form a hematopoietic microenvironment and no detectable single stromal cell cloning capacity. There was no detectable amelioration by MMS350 administration at one week. In contrast, one year post-TBI, Fanca(−/−) mice demonstrated fewer senescent cells in brain and spleen compared to Fanca(+/+) controls. While Fanca(−/−) mouse bone marrow stromal cells explanted one year post-TBI still failed to proliferate in vitro, continuous oral administration of 400 μM, MMS350 in drinking water restored explanted stromal cell proliferation. The data indicate that continuous administration of MMS350 modulated several properties of TBI-accelerated aging in Fanca(−/−) mice as well as control mice, and support further study of MMS350 as a modulator of radiation late effects.

Senescence is an irreversible cell cycle arrest induced by various triggers, including aging and irradiation. Fanconi Anemia (FA) is a genetic disorder associated with defects in 22 proteins in the scaffold DNA double strand break repair. FA mice develop anemia and marrow failure and have been considered as a potential model for accelerated aging. Fancd2-/- murine bone marrow (BM) stromal cell lines are radiosensitive in clonogenic survival curves compared to wild type (Berhane et al, Radiat Res, 2014). We tested the Fanca-/- mouse model for irradiation induced senescence. 129Sv Fanca+/+ and Fanca-/- BM stromal cell lines were irradiated to 5Gy, then assayed for senescence at various timepoints. Cells were stained with a chromogenic beta-galactosidase assay. Cells were also stained by immunofluorescence (IF) to determine percent of cells positive for p16 and p21. For total body irradiation (TBI) studies, 129Sv Fanca+/+ and Fanca-/- mice were given 7Gy. Organs were harvested at Day 7, then assayed for senescence. Statistical analysis was carried out using student's t-test. Baseline levels of beta-gal positive Fanca-/- BM stromal cells were not significantly different from Fanca+/+ (respectively, 0.88% ± 0.22% vs 0.74% ± 0.23%). Fanca-/- and Fanca+/+ IF also showed no significant differences in baseline p16 levels (4.98%±2.5 vs 14.37%± 4.06%) and p21 levels (14.57% ± 7.61% vs. 11.7% ± 3.48%). In contrast, Fanca-/- cells showed significantly higher beta-gal staining compared to Fanca+/+ at days 1, 3, and 5 after irradiation (p=0.0042, p Citation Format: Aranee P. Sivananthan, Xichen Zhang, Wen Hou, Donna Shields, Renee Fisher, Michael Epperly, Joel S. Greenberger. Increased irradiation-induced senescence in Fanconi anemia (FA) mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 842.