Radiation Mitigator Research Articles

Amelioration of radiation-induced pulmonary fibrosis by a water-soluble bifunctional sulfoxide radiation mitigator (MMS350).

A water-soluble ionizing radiation mitigator would have considerable advantages for the management of acute and chronic effects of ionizing radiation. We report that a novel oxetanyl sulfoxide (MMS350) is effective both as a protector and a mitigator of clonal mouse bone marrow stromal cell lines in vitro, and is an effective in vivo mitigator when administered 24 h after 9.5 Gy (LD100/30) total-body irradiation of C57BL/6NHsd mice, significantly improving survival (P = 0.0097). Furthermore, MMS350 (400 μM) added weekly to drinking water after 20 Gy thoracic irradiation significantly decreased: expression of pulmonary inflammatory and profibrotic gene transcripts and proteins; migration into the lungs of bone marrow origin luciferase+/GFP+ (luc+/GFP+) fibroblast progenitors (in both luc+ marrow chimeric and luc+ stromal cell line injected mouse models) and decreased radiation-induced pulmonary fibrosis (P < 0.0001). This nontoxic and orally administered small molecule may be an effective therapeutic in clinical radiotherapy and as a counter measure against the acute and chronic effects of ionizing radiation.

MMS350: A Water Soluble Radiation Protector and Mitigator

We have developed a novel water soluble oxetanyl sulfoxide (MMS350) which is a DMSO analog with radiation protector and mitigator properties. We sought to demonstrate that MMS350 protects and mitigates radiation damage in vivo.

Reduction of Late Pulmonary Fibrosis in Thoracic-Irradiated C57bl/6hnsd Mice by a Water Soluble Radiation Mitigator MMS-350

Radiation induced pulmonary fibrosis is a major complication of radiation therapy and identification of therapeutic drugs to ameliorate this complication remains a challenge. MMS-350, a novel water-soluble sulfoxide that has been developed as a second generation, selective analog of dimethyl sulfoxide (DMSO) has been shown to be a radiation mitigator against total body irradiation of C57Bl/6Hnsd mice. To determine whether the MMS-350 ameliorated irradiation fibrosis, we placed thoracic irradiated mice on MMS-350 in the drinking water.

Human Induced Pluripotent Stem Cell (iPSC) Derived Intestinal Organoids (iHIOs) to Screen Radiation Mitigators

Human Induced Pluripotent Stem Cell (iPSC) Derived Intestinal Organoids (iHIOs) to Screen Radiation Mitigators

Evaluation of potential ionizing irradiation protectors and mitigators using clonogenic survival of human umbilical cord blood hematopoietic progenitor cells

We evaluated the use of colony formation (colony-forming unit-granulocyte macrophage [CFU-GM], burst-forming unit erythroid [BFU-E], and colony-forming unit-granulocyte-erythroid-megakaryocyte-monocytes [CFU-GEMM]) by human umbilical cord blood (CB) hematopoietic progenitor cells for testing novel small molecule ionizing irradiation protectors and mitigators. The following compounds were added before (protection) or after (mitigation) ionizing irradiation: GS-nitroxides (JP4-039 and XJB-5-131), the bifunctional sulfoxide MMS-350, the phosphoinositol-3-kinase inhibitor LY29400, triphenylphosphonium-imidazole fatty acid, the nitric oxide synthase inhibitor (MCF-201-89), the p53/mdm2/mdm4 inhibitor (BEB55), methoxamine, isoproterenol, propranolol, and the adenosine triphosphate-sensitive potassium channel blocker (glyburide). The drugs XJB-5-131, JP4-039, and MMS-350 were radiation protectors for CFU-GM. JP4-039 was also a radiation protector for CFU-GEMM. The drugs XJB-5-131, JP4-039, and MMS-350 were radiation mitigators for BFU-E, MMS-350 and JP4-039 were mitigators for CFU-GM, and MMS350 was a mitigator for CFU-GEMM. In contrast, other drugs were effective in murine assays; TTP-IOA, LY294002, MCF201-89, BEB55, propranolol, isoproterenol, methoxamine, and glyburide but showed no significant protection or mitigation in human CB assays. These data support the testing of new candidate clinical radiation protectors and mitigators using human CB clonogenic assays early in the drug discovery process, thus reducing the need for animal experiments.

Mitigation of Radiation-Induced Damage by Targeting EGFR in Noncancerous Human Epithelial Cells

Methyl-2-cyano-3,12 dioxoolean-1,9 diene-28-oate (CDDO-Me) is an antioxidative, anti-inflammatory modulator, which activates the nuclear factor-erythroid 2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway. While CDDO-Me has radioprotective activity through Nrf2 activation in vitro and in vivo, its ability to mitigate radiation-induced damage when provided after irradiation has not been studied. Here we investigated whether CDDO-Me mitigates ionizing radiation (IR)-induced DNA damage in immortalized normal human colonic epithelial cells (HCECs) and bronchial epithelial cells (HBECs). DNA damage and clonogenic survival were assessed after treatment with CDDO-Me postirradiation. We observed that treatment with CDDO-Me within 30 min after irradiation improved both DNA damage repair and clonogenic survival independently of Nrf2. CDDO-Me activates the epidermal growth factor receptor (EGFR) related DNA repair responses. In the presence of CDDO-Me, EGFR is phosphorylated and translocates into the nucleus where it interacts with DNA-PKcs. CDDO-Me-mediated mitigation activity can be abrogated through depletion of EGFR, ectopic overexpression of mutant EGFR or inhibition of DNA-PKcs. While post-treatment of CDDO-Me protected noncancerous HCECs and HBECs against IR, cancer cells (HCT116 and MCF7) were not protected by CDDO-Me. These results suggest that targeting EGFR using CDDO-Me is a promising radiation mitigator with potential utility for first responders to nuclear accidents.

Gamma-tocopherol-N,N-dimethylglycine ester as a potent post-irradiation mitigator against whole body X-irradiation-induced bone marrow death in mice

We examined the radioprotective and mitigative effects of gamma-tocopherol-N,N-dimethylglycine ester (GTDMG), a novel water-soluble gamma-tocopherol derivative, against X-irradiation-induced bone marrow death in mice. Mice (C3H, 10 weeks, male) were injected intraperitoneally with GTDMG suspended in a 0.5% methyl cellulose solution before or after receiving of 7.5-Gy whole body X-irradiation. GTDMG significantly enhanced the 30-day survival rate when given 30 min before or immediately after the irradiation. Its mitigative activity (administered after exposure) was examined further in detail. The optimal concentration of GTDMG given immediately after irradiation was around 100 mg/kg body weight (bw) and the 30-day survival rate was 97.6 ± 2.4%. When GTDMG was administered 1, 10 and 24 h post-irradiation, the survival rate was 85.7 ± 7.6, 75.0 ± 9.7 and 36.7 ± 8.8%, respectively, showing significant mitigation even at 24 h after irradiation (P < 0.05). The value of the dose reduction factor (100 mg/kg bw, given intraperitoneally (i.p.) immediately after irradiation) was 1.25. GTDMG enhanced the recovery of red blood cell-, white blood cell-, and platelet-counts after irradiation and significantly increased the number of endogenous spleen colonies (P < 0.05). Subcutaneous (s.c.) administration also had mitigative effects. In conclusion, GTDMG is a potent radiation mitigator.

Abstract 70: GS nitroxide (JP4-039) induces radiation resistance of conditional MnSOD tet/tet murine bone marrow stromal cells.

Abstract Background/Objectives: Manganese superoxide dismutase (MnSOD) is a mitochondrial targeted enzyme which dismutates irradiation induced superoxide. Transgenic MnSOD -/- mice die two to three days after birth, and embryo cell lines are radiosensitive in vitro (Epperly et al, Rad Res 154:365-374,2000). Transgene mediated increased expression of MnSOD increases cell line and tissue radioresistance in heterozygote MnSOD +/- mice (Epperly,et al, Rad Res 154:365-374,2000). To regulate MnSOD expression, we developed a mouse line in which expression of one or both endogenous MnSOD alleles is activated by doxycycline (DOX) administration. Homozygous (tet/tet) mice displayed very low levels of MnSOD in the absence of DOX. Both tet/tet and tet/+ cell lines increased MnSOD expression in the presence of DOX. We evaluated the effect of JP4-039 a Gramicidin S (GS) derived-nitroxide as a mitochondrial targeted radiation protector and mitigator on Mnsod tet/tet and MnSOD tet/+cell lines in vitro. Materials/Methods: Bone marrow stromal cell lines were established from MnSOD tet/tet, tet/+ and +/+ mice and compared to embryonic fibroblast cell line from unconditional MnSOD -/- mice. Stress response and cytokine gene expression was measured using real time PCR, and clonagenic survival curves were generated relative to MnSOD biochemical activity. Cells were grown in DOX, JP4-039, or both, 24 hours prior to, or immediately following, irradiation. Results: Incubation of MnSOD tet/tet cells in DOX increased MnSOD biological activity to control levels within 24 hrs. Control C57BL/6NHsd cells were radio-resistant (ñ = 7.90 ± 1.36) relative to MnSOD tet/tet in no DOX (ñ = 2.79 + 1.04, p = 0.0175). Radioresistance was restored in DOX treated MnSOD tet/tet cells (ñ = 8.69 ± 1.09 (p = 0.0060) .There was no significant DOX induced change in radiosensitivity of MnSOD +/+ or MnSOD -/- cells. A synergistic increase in cell survival followed addition of JP4-039 to DOX in MnSOD tet/tet and MnSODtet/+ cell lines. RT-PCR showed an elevation in MnSOD expression, as well as an elevation in expression of stress response genes TGFb,NF-KB, NFE212 at 1 and 6 hours following irradiation in +/+ cells. In tet/+, tet/tet, and -/- cell lines decreased levels of MnSOD were seen at baseline and at both 1 and 6 hours after irradiation without the addition of DOX. DOX added to tet/+ and tet/tet cell lines increased MnSOD levels 1 and 6 hours following irradiation. The addition of DOX to both tet/+ and tet/tet cells caused an increase in TGFb and NF-KB post irradiation in comparison to the same cells without DOX post irradiation. A decrease in NFE212 was found in tet/+ and tet/tet cells post irradiation in comparison to the same cells without DOX post-irradiation. Conclusions: MnSOD tet/tet cells should be valuable to study the cellular mechanisms involved in the response to ionizing irradiation. Acknowledgements: This project supported by NIAID U191A168021. Citation Format: Ronny Kalash, Frank Houghton, Hebist Berhane, Peter Wipf, Donna Shields, Michael W. Epperly, Richard Chaillet, Shaonan Cao, Xichen Zhang, Joel S. Greenberger. GS nitroxide (JP4-039) induces radiation resistance of conditional MnSOD tet/tet murine bone marrow stromal cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 70. doi:10.1158/1538-7445.AM2013-70

Disruption of the PI3K axis abrogates ionizing radiation‐induced cell death

Effective and safe therapies are lacking to abrogate the lethal effects of accidental or intentional public exposure to toxic doses of ionizing radiation. Our recent unbiased phenotypic screen using a sentinel human pluripotent cell line and a small interfering RNA (siRNA) library targeting druggable genes implicated the phosphoinositide‐3‐kinase (PI3K) pathway in cellular death after ionizing radiation. Using a model of embryonic stem cells, we now demonstrate that the prototype PI3K inhibitor LY294002 as well a new PI3K inhibitor class EMF388–021 mitigated the lethal effects of ionizing radiation including the induction of apoptosis. Moreover, a single intraperitoneal dose of LY294002 (30 mg/kg) given 4 or 24 h after 9.25 Gy of total body irradiation significantly reduced death. Similarly, a single intraperitoneal dose of EMF388–021 (10 mg/kg) given 4 h after 9.25 Gy of ionizing radiation markedly reduced lethality. We found PI3K inhibition stimulated radiation‐induced autophagic flux and reduced mitochondrial respiration providing potential mechanisms for the radiation mitigation. This study documents that an unbiased siRNA assay can assist in the identification of pharmacologically useful radiation mitigators and implicates the PI3K pathway in the human radiation response.

Synthesis of analogs of the radiation mitigator JP4-039 and visualization of BODIPY derivatives in mitochondria

JP4-039 is a lead structure in a series of nitroxide conjugates that are capable of accumulating in mitochondria and scavenging reactive oxygen species (ROS). To explore structure-activity relationships (SAR), new analogs with variable nitroxide moieties were prepared. Furthermore, fluorophore-tagged analogs were synthesized and provided the opportunity for visualization in mitochondria. All analogs were tested for radioprotective and radiomitigative effects in 32Dcl3 cells.

MMS350: A Novel Bifunctional Sulfoxide With Radiation Protection and Mitigation Properties

New small molecules for radiation protection and mitigation, applied in either radiation counterterrorism or clinical radiation therapy, must demonstrate safety and effectiveness in both animal models and in humans. MMS350 is a novel water soluble oxetane-substituted sulfoxide which has been developed as a more selective and more potent alternative to dimethyl sulfoxide (DMSO) which is a radioprotector in mice. We evaluated MMS350 as a radioprotector or mitigator. Irradiation survival curves were first performed on bone marrow stromal cells derived from C57BL/6NHsd mice. MMS350 was dissolved in tissue culture medium and administered at a final concentration of 100 μM one hour before irradiation or immediately after irradiation. Cells were irradiated to doses from 0 to 8 Gy , then plated in 4 well tissue culture plates, incubated at 37°C in a high humidity CO2 incubator for 7 days, stained with crystal violet, and colonies of greater than 50 cells counted. The data was analyzed using linear quadratic or single-hit, multiple-target models. To determine if MMS350 was a radiation mitigator in vivo, C57BL/6NTac mice were irradiated to the LD50/30 total body irradiation dose of 9.5 Gy and injected IP with 20 mg/kg of MMS350 dissolved in phosphate buffered saline (PBS) at either 4 or 24 hr after irradiation. MMS350 was protective when administered before irradiation, as shown by the increased shoulder on the survival curve with a ñ of 15.8 ± 2.9 compared to 5.8 ± 1.1 (p = 0.0039) for control irradiated cells. MMS350 was also mitigative when given after irradiation, as shown by the increase in the D0 to 2.4 ± 0.3 compared to 1.9 ± 0.1 (p = 0.0444) for the control irradiated cells. MMS350 was also an effective radiation protector and mitigator of human umbilical cord blood clonagenic multilineage progenitor cells in vitro. Results of in vivo survival curves demonstrated that mice injected with MMS350 at either 4 or 24 hr after irradiation had an increased survival compared to control irradiated mice (p = 0.0092 and 0.0097, respectively). Fifty percent of the control irradiated mice were dead at day 13 after irradiation while 50% of the MMS350 mice survived to day 40. These results demonstrate that MMS350 has a potential for both radiation protection and mitigation of normal tissues in humans.

Identification of Ionizing Irradiation Damage Mitigators by Evaluation of Clonogenic Survival of Human Umbilical Cord Blood Progenitor Cells

Evaluation of the effectiveness of small molecule radiation protectors and mitigators requires in vitro assays for clonogenic survival and analyses of drug effects on apoptosis and DNA damage repair. Application of novel mitigators for potential use in normal tissue protection during clinical radiation therapy and radiation counter terrorism mandates animal testing which, in the LD 50/30 assay for the effects of TBI on the hematopoietic syndrome, involves significant expense. We tested the hypothesis that early evaluation of candidate drugs in human cord blood clonogenic assay could potentially identify drugs which, while effective in the murine system, might be ineffective in humans.

Identification of Druggable Targets for Radiation Mitigation Using a Small Interfering RNA Screening Assay

Currently, there is a serious absence of pharmaceutically attractive small molecules that mitigate the lethal effects of an accidental or intentional public exposure to toxic doses of ionizing radiation. Moreover, cellular systems that emulate the radiobiologically relevant cell populations and that are suitable for high-throughput screening have not been established. Therefore, we examined two human pluripotent embryonal carcinoma cell lines for use in an unbiased phenotypic small interfering RNA (siRNA) assay to identify proteins with the potential of being drug targets for the protection of human cell populations against clinically relevant ionizing radiation doses that cause acute radiation syndrome. Of the two human cell lines tested, NCCIT cells had optimal growth characteristics in a 384 well format, exhibited radiation sensitivity (D(0) = 1.3 ± 0.1 Gy and ñ = 2.0 ± 0.6) comparable to the radiosensitivity of stem cell populations associated with human death within 30 days after total-body irradiation. Moreover, they internalized siRNA after 4 Gy irradiation enabling siRNA library screening. Therefore, we used the human NCCIT cell line for the radiation mitigation study with a siRNA library that silenced 5,520 genes known or hypothesized to be potential therapeutic targets. Exploiting computational methodologies, we identified 113 siRNAs with potential radiomitigative properties, which were further refined to 29 siRNAs with phosphoinositide-3-kinase regulatory subunit 1 (p85α) being among the highest confidence candidate gene products. Colony formation assays revealed radiation mitigation when the phosphoinositide-3-kinase inhibitor LY294002 was given after irradiation of 32D cl 3 cells (D(0) = 1.3 ± 0.1 Gy and ñ = 2.3 ± 0.3 for the vehicle control treated cells compared to D(0) = 1.2 ± 0.1 Gy and ñ = 6.0 ± 0.8 for the LY294002 treated cells, P = 0.0004). LY294002 and two other PI3K inhibitors, PI 828 and GSK 1059615, also mitigated radiation-induced apoptosis in NCCIT cells. Treatment of mice with a single intraperitoneal LY294002 dose of 30 mg/kg at 10 min, 4, or 24 h after LD(50/30) whole-body dose of irradiation (9.25 Gy) enhanced survival. This study documents that an unbiased siRNA assay can identify new genes, signaling pathways, and chemotypes as radiation mitigators and implicate the PI3K pathway in the human radiation response.

Normal tissue protection for improving radiotherapy: Where are the Gaps?

Any tumor could be controlled by radiation therapy if sufficient dose were delivered to all tumor cells. Although technological advances in physical treatment delivery have been developed to allow more radiation dose conformity, normal tissues are invariably included in any radiation field within the tumor volume and also as part of the exit and entrance doses relevant for particle therapy. Mechanisms of normal tissue injury and related biomarkers are now being investigated, facilitating the discovery and development of a next generation of radiation protectors and mitigators. Bringing recent research advances stimulated by development of radiation countermeasures for mass casualties, to clinical cancer care requires understanding the impact of protectors and mitigators on tumor response. These may include treatments that modify cellular damage and death processes, inflammation, alteration of normal flora, wound healing, tissue regeneration and others, specifically to counter cancer site-specific adverse effects to improve outcome of radiation therapy. Such advances in knowledge of tissue and organ biology, mechanisms of injury, development of predictive biomarkers and mechanisms of radioprotection have re-energized the field of normal tissue protection and mitigation. Since various factors, including organ sensitivity to radiation, cellular turnover rate, and differences in mechanisms of injury manifestation and damage response vary among tissues, successful development of radioprotectors/mitigators/treatments may require multiple approaches to address cancer site specific needs. In this review, we discuss examples of important adverse effects of radiotherapy (acute and intermediate to late occurring, when it is delivered either alone or in conjunction with chemotherapy, and important limitations in the current approaches of using radioprotectors and/or mitigators for improving radiation therapy. Also, we are providing general concepts for drug development for improving radiation therapy.

A review of radiation countermeasure work ongoing at the Armed Forces Radiobiology Research Institute

Purpose: The hazard of exposure to ionizing radiation is a serious public and military health concern that has justified substantial efforts to develop medically effective radiation countermeasure approaches, including radiation protectors, mitigators, and therapeutics. Although such efforts were initiated more than half a century ago, no safe and effective radiation countermeasure has been approved by the United States Food and Drug Administration (FDA) for the acute radiation syndrome. This situation has prompted intensified research among government laboratories, academic institutions, and pharmaceutical companies to identify a new generation of countermeasures. In this communication we discuss selected promising radiation countermeasures at advanced stages of development.Conclusion: Other than granulocyte colony-stimulating factor, which has an Emergency Use Investigational New Drug (IND) status, four countermeasures have FDA IND status and other promising countermeasures are in development. Here we review primarily the in vivo efficacy of selected countermeasures in animal models and clinical studies.

Strategies for discovery of small molecule radiation protectors and radiation mitigators.

Mitochondrial targeted radiation damage protectors (delivered prior to irradiation) and mitigators (delivered after irradiation, but before the appearance of symptoms associated with radiation syndrome) have been a recent focus in drug discovery for (1) normal tissue radiation protection during fractionated radiotherapy, and (2) radiation terrorism counter measures. Several categories of such molecules have been discovered: nitroxide-linked hybrid molecules, including GS-nitroxide, GS-nitric oxide synthase inhibitors, p53/mdm2/mdm4 inhibitors, and pharmaceutical agents including inhibitors of the phosphoinositide-3-kinase pathway and the anti-seizure medicine, carbamazepine. Evaluation of potential new radiation dose modifying molecules to protect normal tissue includes: clonogenic radiation survival curves, assays for apoptosis and DNA repair, and irradiation-induced depletion of antioxidant stores. Studies of organ specific radioprotection and in total body irradiation-induced hematopoietic syndrome in the mouse model for protection/mitigation facilitate rational means by which to move candidate small molecule drugs along the drug discovery pipeline into clinical development.

Ionizing Irradiation Protection and Mitigation by Carbamazepine Is p53 and Autophagy Independent,

Abstract 3400 Introduction:Carbamazepine (CBZ), an established sodium channel blocker, used in treatment of epilepsy and trigeminal neuralgia, induces autophagy. CBZ has recently been demonstrated to be an ionizing radiation mitigator and protector (Kim,H.et. al IJRB-in press). CBZ protected C57BL/6NHsd female mice from irradiation if given 24 hr before total body irradiation (TBI) and mitigated if given 12 hours after irradiation.Materials and Methods: To elucidate the mechanism of CBZ action, autophagy incompetent (ATG5−/−) and competent (ATG5+/+) mouse embryo fibroblasts (MEF), p53−/− and p53+/+ bone marrow stromal cells and 32D cl 3 murine IL-3 dependent hematopoietic progenitor cells were tested for CBZ mediated radiation protection and mitigation in clonagenic irradiation survival curves. We also measured CBZ effect on irradiation-induced apoptosis, and depletion of antioxidant stores in vitro and after (TBI) in vivo in control mice and in hind limb irradiated mice with orthotopic tumors. Results:CBZ was a significant radiation protector and mitigator for both ATG5−/− and ATG5 +/+ cell lines by an increased ñ (a measurement of the shoulder on the clonogenic survival curve). CBZ treated ATG5 +/+ cells has an increased ñ of 11.1 +/− 0.2 or 8.8 +/− 0.2 for CBZ added before or after irradiation respectively compared to 5.4 +/− 0.9 for irradiation control cells (p = 0.0287 or 0.0119,respectively). ATG −/− cells were also protected and mitigated by CBZ (ñ of 16.1 +/− 2.6 as a radioprotector or 9.8 +/− 1.5 as a mitigator compared to 4.6 +/− 0.7 for irradiated control cells (p = 0.0002 or 0.0037, respectively). Thus, CBZ functions independent of autophagy. Incubation of p53 +/+ and p53−/− cell lines in 10 μM CBZ for one hour before irradiation protected (ñ of 5.6 +/− 0.9 compared to 1.9 +/− 0.5 for control irradiated p53 +/+ cells (p = 0.0126) and 3.7 +/− 1.9 compared to 1.8 +/− 0.4 for irradiated control p53 −/− cells (p = 0.0018). Thus, CBZ functions in a p53 independent manner. Other pro-autophagy drugs, Valproic Acid and Lithium Carbonate, were ineffective radiation protectors or mitigators. CBZ treatment of 32D cl 3 cells for one hour before or immediately following irradiation had no effect on mitochondrial membrane depolarization, apoptosis or viability but was protective and mitigative in clonagenic survival curve assays. Irradiation initially decreased antioxidant stores in both ATG5+/+ and ATG5−/− cells 10 to 30 min following irradiation. However, cells treated with CBZ had a faster recovery of antioxidant stores to pre-irradiation levels by 90 min compared to 120 min for control irradiated cells. To determine if CBZ protected both normal tissues and tumors in vivo mice with 3LL Lewis Lung carcinoma hind limb xenografts were treated with 10 mg/kg CBZ before or after 20 Gy limb irradiation. There was no significant difference in tumor growth inhibition by 20 Gy in mice treated with CBZ before or after irradiation (p = 0.2431 or 0.5439, respectively). Conclusion:Thus, CBZ is an effective radiation protector and mitigator of normal but not tumor tissues by a novel mechanism independent of autophagy, p53 and apoptosis. Acknowledgments:This project was supported by U191A168021–06. Disclosures:No relevant conflicts of interest to declare.

The Autophagy Inducing Drug Carbamazepine is a Radiation Protector and Mitigator

AbstractPurpose: To determine whether Carbamazepine (CBZ) is a radiation protector and/or mitigator.Materials and methods: Murine hematopoietic progenitor 32D cl 3 cells were incubated in 1, 10, or 100 μM CBZ 1 h before or immediately after 0–8 Gy irradiation and assayed for clonogenic survival. Autophagy was assayed by immunoblot for microtubule-associated protein light chain 3 (LC3). In vivo radioprotection and mitigation were determined with C57BL/6NTac mice.Results: CBZ treatment at 1, 10 or 100 μM for 1 h prior to irradiation increased radioresistance (the dose for 37% survival or D0) from control 1.5 ± 0.1 Gy to 2.1 ± 0.2 Gy (P = 0.012), 2.3 ± 0.1 Gy (P = 0.010), and 3.6 ± 0.7 Gy (P = 0.003), respectively; after irradiation increased the extrapolation number (n) from 1.5 ± 0.3 to 10.1 ± 4.2 (P = 0.011), 5.5 ± 1.7 (P = 0.019), and 3.6 ± 0.8 (P = 0.014), respectively, and increased autophagy. CBZ treated mice 10 min or 24 h before or 10 min or 12 h after 9.25 Gy total body irradiation (TBI) showed inc...

A Manganese–Porphyrin Complex Decomposes H2O2, Inhibits Apoptosis, and Acts as a Radiation Mitigator in Vivo

Ionizing radiation triggers mitochondrial overproduction of H(2)O(2) with concomitant induction of intrinsic apoptosis, whereby clearance of H(2)O(2) upon overexpression of mitochondrial catalase increases radioresistance in vitro and in vivo. As an alternative to gene therapy, we tested the potential of Mn((III))-porphyrin complexes to clear mitochondrial H(2)O(2). We report that triphenyl-[(2E)-2-[4-[(1Z,4Z,9Z,15Z)-10,15,20-tris(4-aminophenyl)-21,23-dihydroporphyrin-5-yl]phenyl]iminoethyl]phosphonium-Mn((III)) compartmentalizes preferentially into mitochondria of mouse embryonic cells, reacts with H(2)O(2), impedes γ-ray-induced mitochondrial apoptosis, and increases the survival of mice exposed to whole body irradiation with γ-rays.

Bone Marrow Stromal Cell Transplantation Mitigates Radiation-Induced Gastrointestinal Syndrome in Mice

BackgroundNuclear accidents and terrorism presents a serious threat for mass casualty. While bone-marrow transplantation might mitigate hematopoietic syndrome, currently there are no approved medical countermeasures to alleviate radiation-induced gastrointestinal syndrome (RIGS), resulting from direct cytocidal effects on intestinal stem cells (ISC) and crypt stromal cells. We examined whether bone marrow-derived adherent stromal cell transplantation (BMSCT) could restitute irradiated intestinal stem cells niche and mitigate radiation-induced gastrointestinal syndrome.Methodology/Principal FindingsAutologous bone marrow was cultured in mesenchymal basal medium and adherent cells were harvested for transplantation to C57Bl6 mice, 24 and 72 hours after lethal whole body irradiation (10.4 Gy) or abdominal irradiation (16–20 Gy) in a single fraction. Mesenchymal, endothelial and myeloid population were characterized by flow cytometry. Intestinal crypt regeneration and absorptive function was assessed by histopathology and xylose absorption assay, respectively. In contrast to 100% mortality in irradiated controls, BMSCT mitigated RIGS and rescued mice from radiation lethality after 18 Gy of abdominal irradiation or 10.4 Gy whole body irradiation with 100% survival (p<0.0007 and p<0.0009 respectively) beyond 25 days. Transplantation of enriched myeloid and non-myeloid fractions failed to improve survival. BMASCT induced ISC regeneration, restitution of the ISC niche and xylose absorption. Serum levels of intestinal radioprotective factors, such as, R-Spondin1, KGF, PDGF and FGF2, and anti-inflammatory cytokines were elevated, while inflammatory cytokines were down regulated.Conclusion/SignificanceMitigation of lethal intestinal injury, following high doses of irradiation, can be achieved by intravenous transplantation of marrow-derived stromal cells, including mesenchymal, endothelial and macrophage cell population. BMASCT increases blood levels of intestinal growth factors and induces regeneration of the irradiated host ISC niche, thus providing a platform to discover potential radiation mitigators and protectors for acute radiation syndromes and chemo-radiation therapy of abdominal malignancies.