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Development of the FAST-DOSE assay system for high-throughput biodosimetry and radiation triage.

ABSTRACT: Following a large-scale radiological incident, there is a need for FDA-approved biodosimetry devices and biomarkers with the ability to rapidly determine past radiation exposure with sufficient accuracy for early population triage and medical management. Towards this goal, we have developed FAST-DOSE (Fluorescent Automated Screening Tool for Dosimetry), an immunofluorescent, biomarker-based system designed to reconstruct absorbed radiation dose in peripheral blood samples collected from potentially exposed individuals. The objective of this study was to examine the performance of the FAST-DOSE assay system to quantify intracellular protein changes in blood leukocytes for early biodosimetry triage from humanized NOD-scid-gamma (Hu-NSG) mice and non-human primates (NHPs) exposed to ionizing radiation up to 8 days after radiation exposure. In the Hu-NSG mice studies, the FAST-DOSE biomarker panel was able to generate delivered dose estimates at days 1, 2 and 3 post exposure, whereas in the NHP studies, the biomarker panel was able to successfully classify samples by dose categories below or above 2 Gy up to 8 days after total body exposure. These results suggest that the FAST-DOSE bioassay has large potential as a useful diagnostic tool for rapid and reliable screening of potentially exposed individuals to aid early triage decisions within the first week post-exposure.

SUBMITTER: Wang Q

PROVIDER: S-EPMC7392759 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Development of the FAST-DOSE assay system for high-throughput biodosimetry and radiation triage.

Wang Qi Q Lee Younghyun Y Shuryak Igor I Pujol Canadell Monica M Taveras Maria M Perrier Jay R JR Bacon Bezalel A BA Rodrigues Matthew A MA Kowalski Richard R Capaccio Christopher C Brenner David J DJ Turner Helen C HC

Scientific reports 20200729 1

Following a large-scale radiological incident, there is a need for FDA-approved biodosimetry devices and biomarkers with the ability to rapidly determine past radiation exposure with sufficient accuracy for early population triage and medical management. Towards this goal, we have developed FAST-DOSE (Fluorescent Automated Screening Tool for Dosimetry), an immunofluorescent, biomarker-based system designed to reconstruct absorbed radiation dose in peripheral blood samples collected from potentia ...[more]

PMID: 32728041

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Project description:BackgroundThe effects of dose-rate and its implications on radiation biodosimetry methods are not well studied in the context of large-scale radiological scenarios. There are significant health risks to individuals exposed to an acute dose, but a realistic scenario would include exposure to both high and low dose-rates, from both external and internal radioactivity. It is important therefore, to understand the biological response to prolonged exposure; and further, discover biomarkers that can be used to estimate damage from low-dose rate exposures and propose appropriate clinical treatment.MethodsWe irradiated human whole blood ex vivo to three doses, 0.56 Gy, 2.23 Gy and 4.45 Gy, using two dose rates: acute, 1.03 Gy/min and a low dose-rate, 3.1 mGy/min. After 24 h, we isolated RNA from blood cells and these were hybridized to Agilent Whole Human genome microarrays. We validated the microarray results using qRT-PCR.ResultsMicroarray results showed that there were 454 significantly differentially expressed genes after prolonged exposure to all doses. After acute exposure, 598 genes were differentially expressed in response to all doses. Gene ontology terms enriched in both sets of genes were related to immune processes and B-cell mediated immunity. Genes responding to acute exposure were also enriched in functions related to natural killer cell activation and cell-to-cell signaling. As expected, the p53 pathway was found to be significantly enriched at all doses and by both dose-rates of radiation. A support vectors machine classifier was able to distinguish between dose-rates with 100 % accuracy using leave-one-out cross-validation.ConclusionsIn this study we found that low dose-rate exposure can result in distinctive gene expression patterns compared with acute exposures. We were able to successfully distinguish low dose-rate exposed samples from acute dose exposed samples at 24 h, using a gene expression-based classifier. These genes are candidates for further testing as markers to classify exposure based on dose-rate.

Project description:Dosimetry is an important tool for triage and treatment planning following any radiation exposure accident, and biological dosimetry, which estimates exposure dose using a biological parameter, is a practical means of determining the specific dose an individual receives. The cytokinesis-blocked micronucleus assay (CBMN) is an established biodosimetric tool to measure chromosomal damage in mitogen-stimulated human lymphocytes. The CBMN method is especially valuable for biodosimetry in triage situations thanks to simplicity in scoring and adaptability to high-throughput automated sample processing systems. While this technique produces dose-response data which fit very well to a linear-quadratic model for exposures to low linear energy transfer (LET) radiation and for doses up for 5 Gy, limitations to the accuracy of this method arise at larger doses. Accuracy at higher doses is limited by the number of cells reaching mitosis. Whereas it would be expected that the yield of micronuclei increases with the dose, in many experiments it has been shown to actually decrease when normalized over the total number of cells. This variation from a monotonically increasing dose response poses a limitation for retrospective dose reconstruction. In this study we modified the standard CBMN assay to increase its accuracy following exposures to higher doses of photons or a mixed neutron-photon beam. The assay is modified either through inhibitions of the G2/M and spindle checkpoints with the addition of caffeine and/or ZM447439 (an Aurora kinase inhibitor), respectively to the blood cultures at select times during the assay. Our results showed that caffeine addition improved assay performance for photon up to 10 Gy. This was achieved by extending the assay time from the typical 70 h to just 74 h. Compared to micronuclei yields without inhibitors, addition of caffeine and ZM447439 resulted in improved accuracy in the detection of micronuclei yields up to 10 Gy from photons and 4 Gy of mixed neutrons-photons. When the dose-effect curves were fitted to take into account the turnover phenomenon observed at higher doses, best fitting was achieved when the combination of both inhibitors was used. These techniques permit reliable dose reconstruction after high doses of radiation with a method that can be adapted to high-throughput automated sample processing systems.

Project description:There is a need for research to rapidly determine an individual's absorbed dose and its potential health effects after a potential radiological or nuclear event that could expose large portions of a population to ionizing radiation (IR). Studies on biomarker identification after radiation exposure could aid in biodosimetry, identifying individual dose absorbed, as well as biologic response, and administering immediate and proper medical care. Metabolomics on easily accessible biofluids is an emerging field with potential for high-throughput biodosimetry. While tremendous effort has been put into obtaining discovery based global radiation signatures from a number of biofluids and model organisms, quantitative targeted analysis on a subset of known radiation biomarkers is required to develop an optimized panel of biomarkers for future clinical applications. The current study analyzes levels of several known broad chemical groups (acylcarnitines, amino acids, phosphatidylcholines, and biogenic amines) affected by IR in serum from nonhuman primates (NHP) 7 days after exposure through multiple reaction monitoring (MRM) analysis with a triple quadrupole mass spectrometry (MS) platform. We identified several novel metabolites affected by IR exposure through univariate and unsupervised multivariate analyses. Levels of acylcarnitines, amino acids, and phospholipids were perturbed indicating altered protein metabolism, fatty acid β-oxidation, and inflammation. Fold changes in carnitine and short-chain acylcarnitines (acetylcarnitine, propionylcarnitine, butyrylcarnitine, and valerylcarnitine) complement previous global radiation signatures on NHP; notably, the levels of change were lower than previously observed in urine. Decreased levels of glutamate, citrulline, and arginine after IR are biomarkers indicating gastrointestinal syndrome and perturbations to the urea cycle. Sex differences were also assessed and were more prevalent in circulating acylcarnitines and phospholipids after IR exposure. These biomarkers may be combined with previously described compounds from DNA damage to develop a defined metabolomic biodosimetry panel to be analyzed by MS platforms, which are increasingly available in clinical laboratories.

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Development of the FAST-DOSE assay system for high-throughput biodosimetry and radiation triage.

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Development of the FAST-DOSE assay system for high-throughput biodosimetry and radiation triage.

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