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Exposure to ionizing radiation induces cellular and molecular damage leading to a cascade of events resulting in tissue and organ injury. Our study strives to characterize and validate metabolomic changes in preterminal stage (immediately prior to death) samples collected from rhesus macaques lethally irradiated with one of two different doses of radiation. Peripheral blood samples were collected pre-exposure, post-exposure, and at the preterminal stage of nonhuman primates (NHPs that did not survive exposure with 7.2 Gy or 7.6 Gy total-body radiation (LD 60-80/60 )). We analyzed global metabolomic alterations using ultra-performance liquid chromatography (UPLC) quadrupole time-of-flight mass spectrometry (QTOF-MS) in serum samples collected at various timepoints in relation to radiation exposure. The goal of this study was to validate the metabolic shifts present in samples collected just prior to death, which were reported earlier in a preliminary study with a limited number of samples and a single dose of radiation. Here, we demonstrate that radiation exposure induced significant time-dependent metabolic alterations compared with pre-exposure samples. We observed significant metabolite dysregulation in animals exposed to 7.6 Gy compared to 7.2 Gy. Greater metabolic disruption was observed in the preterminal groups than all of the other post-irradiation timepoints in both cohorts. Metabolomic shifts in these preterminal groups also revealed consistent disturbances in sphingolipid metabolism, steroid hormone biosynthesis, and glycerophospholipid metabolism pathways. Overall, the sphingolipid metabolism pathway appears to be representative of the preterminal phenotype, confirming the results of our preliminary study. These results offer important and novel insights for identification and validation of biomarkers for lethality, and such observations would be valuable for triage during a radiological/nuclear mass casualty scenario., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

The identification and validation of radiation biomarkers is critical for assessing the radiation dose received in exposed individuals and for developing radiation medical countermeasures that can be used to treat acute radiation syndrome (ARS). Additionally, a fundamental understanding of the effects of radiation injury could further aid in the identification and development of therapeutic targets for mitigating radiation damage. In this study, blood samples were collected from fourteen male nonhuman primates (NHPs) that were exposed to 7.2 Gy ionizing radiation at various time points (seven days prior to irradiation; 1, 13, and 25 days post-irradiation; and immediately prior to the euthanasia of moribund (preterminal) animals). Plasma was isolated from these samples and was analyzed using a liquid chromatography tandem mass spectrometry approach in an effort to determine the effects of radiation on plasma proteomic profiles. The primary objective was to determine if the radiation-induced expression of specific proteins could serve as an early predictor for health decline leading to a preterminal phenotype. Our results suggest that radiation induced a complex temporal response in which some features exhibit upregulation while others trend downward. These statistically significantly altered features varied from pre-irradiation levels by as much as tenfold. Specifically, we found the expression of integrin alpha and thrombospondin correlated in peripheral blood with the preterminal stage. The differential expression of these proteins implicates dysregulation of biological processes such as hemostasis, inflammation, and immune response that could be leveraged for mitigating radiation-induced adverse effects., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Exposure to high, marginally lethal doses or higher of ionizing radiation, either intentional or accidental, results in injury to various organs. Currently, there is only a limited number of safe and effective radiation countermeasures approved by US Food and Drug Administration for such injuries. These approved agents are effective for only the hematopoietic component of the acute radiation syndrome and must be administered only after the exposure event: currently, there is no FDA-approved agent that can be used prophylactically. The nutraceutical, gamma-tocotrienol (GT3) has been found to be a promising radioprotector of such exposure-related injuries, especially those of a hematopoietic nature, when tested in either rodents or nonhuman primates. We investigated the nature of injuries and the possible protective effects of GT3 within select organ systems/tissues caused by both non-lethal level (4.0 Gy), as well as potentially lethal level (5.8 Gy) of ionizing radiation, delivered as total-body or partial-body exposure. Results indicated that the most severe, dose-dependent injuries occurred within those organ systems with strong self-renewing capacities (e.g., the lymphohematopoietic and gastrointestinal systems), while in other tissues (e.g., liver, kidney, lung) endowed with less self-renewal, the pathologies noted tended to be less pronounced and less dependent on the level of exposure dose or on the applied exposure regimen. The prophylactic use of the test nutraceutical, GT3, appeared to limit the extent of irradiation-associated pathology within blood forming tissues and, to some extent, within the small intestine of the gastrointestinal tract. No distinct, global pattern of bodily protection was noted with the agent's use, although a hint of a possible radioprotective benefit was suggested not only by a lessening of apparent injury within select organ systems, but also by way of noting the lack of early onset of moribundity within select GT3-treated animals., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Despite remarkable scientific progress over the past six decades within the medical arts and in radiobiology in general, limited radiation medical countermeasures (MCMs) have been approved by the United States Food and Drug Administration for the acute radiation syndrome (ARS). Additional effort is needed to develop large animal models for improving the prediction of clinical safety and effectiveness of MCMs for acute and delayed effects of radiation in humans. Nonhuman primates (NHPs) are considered the animal models that reproduce the most appropriate representation of human disease and are considered the gold standard for drug development and regulatory approval. The clinical and histopathological effects of supralethal, total- or partial-body irradiations (12 Gy) of NHPs were assessed, along with possible protective actions of a promising radiation MCM, gamma-tocotrienol (GT3). Results show that these supralethal radiation exposures induce severe injuries that manifest both clinically as well as pathologically, as evidenced by the noted functionally crippling lesions within various major organ systems of experimental NHPs. The MCM, GT3, has limited radioprotective efficacy against such supralethal radiation doses., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Exposure to ionizing radiation (IR) presents a formidable clinical challenge. Total-body or significant partial-body exposure at a high dose and dose rate leads to acute radiation syndrome (ARS), the complex pathologic effects that arise following IR exposure over a short period of time. Early and accurate diagnosis of ARS is critical for assessing the exposure dose and determining the proper treatment. Serum microRNAs (miRNAs) may effectively predict the impact of irradiation and assess cell viability/senescence changes and inflammation. We used a nonhuman primate (NHP) model-rhesus macaques (Macaca mulatta)-to identify the serum miRNA landscape 96 h prior to and following 7.2 Gy total-body irradiation (TBI) at four timepoints: 24, 36, 48, and 96 h. To assess whether the miRNA profile reflects the therapeutic effect of a small molecule ON01210, commonly known as Ex-Rad, that has demonstrated radioprotective efficacy in a rodent model, we administered Ex-Rad at two different schedules of NHPs; either 36 and 48 h post-irradiation or 48 and 60 h post-irradiation. Results of this study corroborated our previous findings obtained using a qPCR array for several miRNAs and their modulation in response to irradiation: some miRNAs demonstrated a temporary increased serum concentration within the first 24-36 h (miR-375, miR-185-5p), whereas others displayed either a prolonged decline (miR-423-5p) or a long-term increase (miR-30a-5p, miR-27b-3p). In agreement with these time-dependent changes, hierarchical clustering of differentially expressed miRNAs showed that the profiles of the top six miRNA that most strongly correlated with radiation exposure were inconsistent between the 24 and 96 h timepoints following exposure, suggesting that different biodosimetry miRNA markers might be required depending on the time that has elapsed. Finally, Ex-Rad treatment restored the level of several miRNAs whose expression was significantly changed after radiation exposure, including miR-16-2, an miRNA previously associated with radiation survival. Taken together, our findings support the use of miRNA expression as an indicator of radiation exposure and the use of Ex-Rad as a potential radioprotectant., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Radiological incidents or terrorist attacks would likely expose civilians and military personnel to high doses of ionizing radiation, leading to the development of acute radiation syndrome. We examined the effectiveness of prophylactic administration of a developmental radiation countermeasure, γ-tocotrienol (GT3), in a total-body irradiation (TBI) mouse model. CD2F1 mice received GT3 24 h prior to 11 Gy cobalt-60 gamma-irradiation. This dose of radiation induces severe hematopoietic acute radiation syndrome and moderate gastrointestinal injury. GT3 provided 100% protection, while the vehicle control group had 100% mortality. Two-dimensional differential in-gel electrophoresis was followed by mass spectrometry and Ingenuity Pathway Analysis (IPA). Analysis revealed a change in expression of 18 proteins in response to TBI, and these changes were reversed with prophylactic treatment of GT3. IPA revealed a network of associated proteins involved in cellular movement, immune cell trafficking, and inflammatory response. Of particular interest, significant expression changes in beta-2-glycoprotein 1, alpha-1-acid glycoprotein 1, alpha-2-macroglobulin, complement C3, mannose-binding protein C, and major urinary protein 6 were noted after TBI and reversed with GT3 treatment. This study reports the untargeted approach, the network, and specific serum proteins which could be translated as biomarkers of both radiation injury and protection by countermeasures., (© 2022. The Author(s).)

To date, the United States Food and Drug Administration (FDA) has approved four drugs to mitigate hematopoietic acute radiation syndrome and all four are repurposed radiomitigators. There are several additional drug candidates currently under evaluation that may also be helpful for use during a widespread emergency. One possible candidate is Ex-Rad, also known as ON01210, a chlorobenzyl sulfone derivative (organosulfur compound), which is a novel, small-molecule kinase inhibitor with demonstrated efficacy in the murine model. In this study, we have evaluated the metabolomic and lipidomic profiles in serum samples of nonhuman primates (NHPs) treated with Ex-Rad after exposure to ionizing radiation. Two different dose administration schedules (Ex-Rad I administered 24 and 36 h post-irradiation, and Ex-Rad II administered 48 and 60 h post-irradiation), were used and evaluated using a global molecular profiling approach. We observed alterations in biochemical pathways relating to inflammation and oxidative stress after radiation exposure that were alleviated in animals that received Ex-Rad I or Ex-Rad II. The results from this study lend credence to the possible radiomitigative effects of this drug possibly via a dampening of metabolism-based tissue injury, thus aiding in recovery of vital, radiation-injured organ systems.

Hematopoietic acute radiation syndrome (H-ARS) and delayed effects of acute radiation exposure (DEARE) are detrimental health effects that occur after exposure to high doses of ionizing radiation. BIO 300, a synthetic genistein nanosuspension, was previously proven safe and effective against H-ARS when administered (via the oral (po) or intramuscular (im) route) prior to exposure to lethal doses of total-body radiation. In this study, we evaluated the proteomic changes in serum of nonhuman primates (NHP) after administering BIO 300 by different routes (po and im). We utilized nanoflow-ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (NanoUPLC-MS/MS) methods for comprehensive global profiling and quantification of serum proteins. The results corroborate previous findings that suggest a very similar metabolic profile following both routes of drug administration. Furthermore, we observed minor alterations in protein levels, 2 hours after drug administration, which relates to the C max of BIO 300 for both routes of administration. Taken together, this assessment may provide an insight into the mechanism of radioprotection of BIO 300 and a reasonable illustration of the pharmacodynamics of this radiation countermeasure.

Although multiple radioprotectors are currently being investigated preclinically for efficacy and safety, few studies have investigated concomitant metabolic changes. This study examines the effects of amifostine on the metabolic profiles in tissues of mice exposed to cobalt-60 total-body gamma-radiation. Global metabolomic and lipidomic changes were analyzed using ultra-performance liquid chromatography (UPLC) quadrupole time-of-flight mass spectrometry (QTOF-MS) in bone marrow, jejunum, and lung samples of amifostine-treated and saline-treated control mice. Results demonstrate that radiation exposure leads to tissue specific metabolic responses that were corrected in part by treatment with amifostine in a drug-dose dependent manner. Bone marrow exhibited robust responses to radiation and was also highly responsive to protective effects of amifostine, while jejunum and lung showed only modest changes. Treatment with amifostine at 200 mg/kg prior to irradiation seemed to impart maximum survival benefit, while the lower dose of 50 mg/kg offered only limited survival benefit. These findings show that the administration of amifostine causes metabolic shifts that would provide an overall benefit to radiation injury and underscore the utility of metabolomics and lipidomics to determine the underlying physiological mechanisms involved in the radioprotective efficacy of amifostine. This approach may be helpful in identifying biomarkers for radioprotective efficacy of amifostine and other countermeasures under development.