Having determined the feasibility of small-molecule profiling for radiation biodosimetry with total body, high dose-rate, external-beam γ-ray exposures and having established the field of radiation metabolomics using cultured cells and animals, our present efforts extend radiation metabolomics to studies of radiation exposures that will typically occur during a radiologic or nuclear event. Real-world, population-level exposure scenarios will include various qualities of radiation and differentially exposed tissues and organs. In particular, we are expanding our research into more real-world scenarios such as low dose-rate exposures, partial body exposures resulting from shielding of certain organs and tissues, exposures to radioisotopes following an IND or RDD, and mixed exposures to neutrons and low linear energy transfer (LET) radiation typical of an IND. We are also focusing attention on the development of prognostic biomarkers to predict individual outcomes from near-lethal exposures as well as the mechanisms involved in biomarker responses. Continuous development of bioinformatic approaches aims to answer complex questions about the connection of the biomarkers and the pathway involvement, in addition to developing new algorithms of biomarker identification.
In a situation of a radiological accident or dirty bomb dispersal, rapid triage of potentially exposed individuals will be of importance. For this, identification of radiation biomarkers through metabolomics was expanded to a human population. Patients undergoing total body irradiation prior to hematopoietic stem cell transplantation had urine collected before and after one dose of 125 cGy (6h post-irradiation). Metabolomic profiling, followed by validation of the markers through tandem mass spectrometry, and quantification revealed disturbances in two major pathways, as described below.
Developing a panel of biomarkers from human biofluids has the potential to evolve into a rapid method of identifying exposed individuals for effective triage in a situation where timely and precise diagnosis would be necessary.
In the event of a radiological event, radionuclides are feared as they will persist in the food chain and environment. Consumption of water and food will lead to internal exposure, leading to additional radiation exposure over time. We aim to understand the differences between internal and external exposure in terms of biomarker identification and metabolic pathways that are altered. The first study in collaboration with Lovelace Respiratory Research Institute focused on internal exposure to 137Cs. The study lasted for 30 days and mice received cumulative doses between 1.95 and 9.91 Gy (time-dependent exposure). As highlighted in the multidimensional scaling plot, the overall metabolic profiles lead to clustering of control versus treated mice in two distinct groups.
Although the health risks of exposure to high doses of radiation at high dose rates during a nuclear or a radiological disaster are well recognized, much research remains to be done to establish the role of dose rate, particularly at low dose rates. To date, we have employed metabolomic and bioinformatics approaches to study the unique effects of dose rate on the urinary excretion of metabolites in mice exposed to external beam irradiation at the low dose rate (LDR) of 0.00309 Gy/min compared to high dose rate (HDR). The mice were exposed to the total dose of 1.1 Gy and 4.45 Gy at both dose rates. We took advantage of the high resolving power of ultra-performance liquid chromatography (UPLC) and the high sensitivity of time-of-flight mass spectrometry (TOFMS) to determine the changes in the urinary metabolome of mice exposed to 4.45 Gy and 1.1 Gy at LDR compared to mice exposed to the same doses at HDR.
The results of this study revealed that exposure to external beam γ-irradiation perturbed energy metabolism, fatty acid oxidation, and amino acid metabolism independent of dose rate. Dose rate is speculated to modulate the magnitude of metabolic perturbations and the targets within metabolic pathways.
Future studies will probe intermediate, higher and lower dose rates with the aim of refining these signatures of effects over a wider range of dose rates.
While untargeted metabolomics focuses on global changes and profiling, targeted metabolomics assesses only specific metabolites or pathways. Additional benefits include, with enrichment of samples, to quantify molecules with extremely low concentrations in biological samples. A targeted metabolomics approach was undertaken to assess changes in the serum of irradiated mice.
Previous research on mutant mouse models focused on the effects of mutations in key DNA repair pathways to the metabolome, in urine and blood. Parp1-/- mice are deficient in base excision repair, ATM-/- in double-strand break repair and radiation signaling, and Prkdc-/- (Scid) deficient in non-homologous end joining. While Parp1-/- show delayed responses and a generally equal number of metabolites of high and low levels of metabolites between irradiated wild-type and mutant, Scid mice are characterized by an overall decreased level of metabolites, whereas Atm mice by high levels of excretion. New research will focus on pro- and anti-inflammatory mouse models to identify the contribution of immune responses and signaling in the metabolic signature.
Salivary metabolomics has been utilized before to discriminate individuals with oral cancer, periodontal disease, breast cancer, and pancreatic cancer. With the oral mucosa and salivary glands being exceptionally radiosensitive, we investigated the metabolic profiles of mouse saliva. Polar analysis showed a dose-response present in many metabolites. As this is the first study to show salivary responses with metabolomics, we will explore this biofluid further for its potential in radiation biodosimetry. The results of the pilot project are currently in press in Radiation Research.
In the possible scenario of a terrorist attack, a possible gun-type detonation of an improvised nuclear device with highly enriched uranium will produce a substantial neutron exposure risk. As fast neutrons have a much higher relative biological effectiveness (RBE) than gamma rays or x rays, it is expected that the same dose of either of the exposures will have a significantly different biological response. This, in turn, will translate to different radiation signatures in both urine and blood. In the new round, we aim to characterize the metabolic responses to mixed irradiation of 3 Gy x rays with increasing percentages of neutrons. All irradiations will be performed at RARAF.