Amelioration of Head and Neck Radiation-Induced Mucositis and Distant Marrow Suppression in Fanca-/- and Fancg-/- Mice by Intraoral Administration of GS-Nitroxide (JP4-039).

Squamous cell carcinomas of the head and neck are appearing with increased frequency in both marrow transplanted and non-transplanted Fanconi anemia (FA) patients. FA patients commonly display radiosensitivity of epithelial tissues, complicating effective radiotherapy. Fancd2-/- mice (C57BL/6J and 129/Sv background) demonstrate epithelial tissue sensitivity to single-fraction or fractionated irradiation to the head and neck and distant marrow suppression (abscopal effect), both ameliorated by intraoral administration of the mitochondrial-targeted antioxidant, GS-nitroxide, JP4-039. We now report that mice of two other FA genotypes, Fancg-/- (B6) and the most prevalent human genotype Fanca-/- (129/Sv), also demonstrate: 1. reduced longevity of hematopoiesis in long-term bone marrow cultures; 2. radiosensitivity of bone marrow stromal cell lines; and 3. head and neck radiation-induced severe mucositis and abscopal suppression of distant marrow hematopoiesis. Intraoral administration of JP4-039/F15, but not non-mitochondrial-targeted 4-amino-Tempo/F15 or F15 alone, prior to each radiation treatment ameliorated both local and abscopal radiation effects. Head and neck irradiated TGF-ß-resistant SMAD3-/- (129/Sv) mice and double-knockout SMAD3-/- Fancd2-/- (129/Sv) mice treated daily with TGF-ß receptor antagonist, LY364947, still displayed abscopal bone marrow suppression, implicating a non-TGF-ß mechanism. Thus, amelioration of both local normal tissue radiosensitivity and distant marrow suppression by intraoral administration of JP4-039 in Fancg-/- and Fanca-/- mice supports a clinical trial of this locally administered normal tissue radioprotector and mitigator during head and neck irradiation in FA patients.

The GS-nitroxide JP4-039 improves intestinal barrier and stem cell recovery in irradiated mice.

Total body irradiation (TBI) leads to dose- and tissue-specific lethality. In the current study, we demonstrate that a mitochondrion-targeted nitroxide JP4-039 given once 24 hours after 9-10 Gy TBI significantly improves mouse survival, and the recovery of intestinal barrier, differentiation and stem cell functions. The GI-protective effects are associated with rapid and selective induction of tight junction proteins and cytokines including TGF-ß, IL-10, IL-17a, IL-22 and Notch signaling long before bone marrow depletion. However, no change was observed in crypt death or the expression of prototypic pro-inflammatory cytokines such as TNF-α, IL-6 or IL-1ß. Surprisingly, bone marrow transplantation (BMT) performed 24 hours after TBI improves intestinal barrier and stem cell recovery with induction of IL-10, IL-17a, IL-22, and Notch signaling. Further, BMT-rescued TBI survivors display increased intestinal permeability, impaired ISC function and proliferation, but not obvious intestinal inflammation or increased epithelial death. These findings identify intestinal epithelium as a novel target of radiation mitigation, and potential strategies to enhance ISC recovery and regeneration after accidental or medical exposures.

Improved Total-Body Irradiation Survival by Delivery of Two Radiation Mitigators that Target Distinct Cell Death Pathways.

The acute lethality of total-body irradiation (TBI) involves damage to multiple organs, including bone marrow and intestine. Ionizing radiation mitigators that are effective when delivered 24 h or later after TBI include the anti-apoptotic drug, JP4-039 and the anti-necroptotic drug, necrostatin-1. In contrast to effective delivery of JP4-039 at 24 h after TBI, necrostatin-1 is most effective when delivery is delayed until 48 h, a time that correlates with the elevation of necroptosis-inducing inflammatory cytokines and necroptosis-induced serine phosphorylation of receptor-interacting serine/threonine-protein kinase-3 (RIP3) in tissues. The goal of this work was to determine whether administration of JP4-039 influenced the optimal delivery time for necrostatin-1. We measured daily levels of 33 proteins in plasma compared to intestine and bone marrow of C57BL/6NTac female mice over a 7-day time period after 9.25 Gy TBI (LD50/30). Protein responses to TBI in plasma were different from those measured in intestine or bone marrow. In mice that were given JP4-039 at 24 h after TBI, we delayed necrostatin-1 delivery for 72 h after TBI based on measured delay in RIP-3 kinase elevation in marrow and intestine. Sequential delivery of these two radiation mitigator drugs significantly increased survival compared to single drug administration.