Combination Mitigators, GS-Nitroxide JP4-039 and water Soluble Oxetanyl Sulfoxide MMS350 Improve Survival of Lethally Irradiated Mice

New small molecule radiation mitigator drugs are needed to administer 24 hr or later after a mass casualty radiation event. We have developed two small molecule mitigators with applicability as clinical radiation protectors. The mitochondrial targeted GS-nitroxide, JP4-039, acts as a superoxide dismutase mimic. A novel oxetanyl sulfoxide, MMS350, acts as a hydroxyl radical scavenger and mitigates both acute total body irradiation damage and prevents late radiation pulmonary fibrosis (Kalash, Epperly, Cao et. al., Radiation Research 180:474-490, 2013). We tested the effect of both compounds together in the C57BL/6NTac mouse TBI irradiation model. Each drug was a radiation protector and mitigator with murine hematopoietic progenitor cell line 32D cl3 in vitro. JP4-039 (10 μM) or MMS350 (100 μM) were incubated with the cells one hour before irradiation or added 30 min following irradiation. The irradiated cells were plated in methylcellulose, incubated for 7 days at 37^oC, and colonies of greater than 50 cells counted. The data was analyzed using linear quadratic or single hit, multi-target models. In vivo total body irradiated C57BL/6NTac mice received 9.25 Gy and were injected intravenously 24 hr later with JP4-039 (20 mg/kg) in F14 emulsion, MMS350 (20 mg/kg) or both JP4-039 (20 mg/kg) + MMS350 (20 mg/kg). Mice were followed for development of the hematopoietic syndrome (irradiation induced bone marrow failure). In vitro radiation survival curves demonstrated that both JP4-039 and MMS350 protected and mitigated 32D cl 3 cells by increasing the shoulder on the clonagenic survival curves. The shoulder on survival curve for 32D cl 3 cells was 2.0 ± 0.3 compared to 3.8 ± 0.5 when JP4-039 was administered pre irradiation (p = 0.0092), 5.4 ± 0.5 when JP4-039 was administered post irradiation (p = 0.0002), 3.5 ± 0.3 for cells incubated with MMS350 pre irradiation (p = 0.0104) or 5.3 ± 1.5 when MMS350 is administered post irradiation (p = 0.0096). Mice treated with JP4-039 had increased survival at 60 days following 9.25 Gy from 20% to 65% (p = 0.0092). Mice treated with MMS350 showed an increased survival of 55% (p = 0.0097). The combination of JP4-039 and MMS350 was additive and increased survival to 95% (p < 0.0001). The combination of JP4-039 and MMS350 increased mitigation of irradiation damage better than one drug alone.