Radiation Biological Toximetry Using Circulating Cell-Free DNA (cfDNA) for Rapid Radiation/Nuclear Triage.

Optimal triage biodosimetry would include risk stratification within minutes, and it would provide useful triage despite heterogeneous dosimetry, cytokine therapy, mixed radiation quality, race, and age. For regulatory approval, the U.S. Food and Drug Administration (FDA) Biodosimetry Guidance requires suitability for purpose and a validated species-independent mechanism. Circulating cell-free DNA (cfDNA) concentration assays may provide such triage information. To test this hypothesis, cfDNA concentrations were measured in unprocessed monkey plasma using a branched DNA (bDNA) technique with a laboratory developed test. The cfDNA levels, along with hematopoietic parameters, were measured over a 7-day period in Rhesus macaques receiving total body radiation doses ranging from 1 to 6.5 Gy. Low-dose irradiation (0-2 Gy) was easily distinguished from high-dose whole-body exposures (5.5 and 6.5 Gy). Fold changes in cfDNA in the monkey model were comparable to those measured in a bone marrow transplant patient receiving a supralethal radiation dose, suggesting that the lethal threshold of cfDNA concentrations may be similar across species. Average cfDNA levels were 50 ± 40 ng/mL [±1 standard deviation (SD)] pre-irradiation, 120 ± 13 ng/mL at 1 Gy; 242 ± 71 ng/mL at 2 Gy; 607 ± 54 at 5.5 Gy; and 1585 ± 351 at 6.5 Gy (±1 SD). There was an exponential increase in cfDNA concentration with radiation dose. Comparison of the monkey model with the mouse model and the Guskova model, developed using Chernobyl responder data, further demonstrated correlation across species, supporting a similar mechanism of action. The test is available commercially in a Clinical Laboratory Improvement Amendments (CLIA) ready form in the U.S. and the European Union. The remaining challenges include developing methods for further simplification of specimen processing and assay evaluation, as well as more accurate calibration of the triage category with cfDNA concentration cutoffs.

Implications of "flash" radiotherapy for biodosimetry.

Extremely high dose rate radiation delivery (FLASH) for cancer treatment has been shown to produce less damage to normal tissues while having the same radiotoxic effect on tumor tissue (referred to as the FLASH effect). Research on the FLASH effect has two very pertinent implications for the field of biodosimetry: (1) FLASH is a good model to simulate delivery of prompt radiation from the initial moments after detonating a nuclear weapon and (2) the FLASH effect elucidates how dose rate impacts the biological mechanisms that underlie most types of biological biodosimetry. The impact of dose rate will likely differ for different types of biodosimetry, depending on the specific underlying mechanisms. The greatest impact of FLASH effects is likely to occur for assays based on biological responses to radiation damage, but the consequences of differential effects of dose rates on the accuracy of dose estimates has not been taken into account.

Benefits and challenges of in vivo EPR nail biodosimetry in a second tier of medical triage in response to a large radiation event.

Following large-scale radiation events, an overwhelming number of people will potentially need mitigators or treatment for radiation-induced injuries. This necessitates having methods to triage people based on their dose and its likely distribution, so life-saving treatment is directed only to people who can benefit from such care. Using estimates of victims following an improvised nuclear device striking a major city, we illustrate a two-tier approach to triage. At the second tier, after first removing most who would not benefit from care, biodosimetry should provide accurate dose estimates and determine whether the dose was heterogeneous. We illustrate the value of using in vivo electron paramagnetic resonance nail biodosimetry to rapidly assess dose and determine its heterogeneity using independent measurements of nails from the hands and feet. Having previously established its feasibility, we review the benefits and challenges of potential improvements of this method that would make it particularly suitable for tier 2 triage. Improvements, guided by a user-centered approach to design and development, include expanding its capability to make simultaneous, independent measurements and improving its precision and universality.

FGF7 peptide (FGF7p) mimetic mitigates bladder urothelial injury from cyclophosphamide.

Although full-length fibroblast growth factor 7 (FGF7) blocks cyclophosphamide-induced urothelial apoptosis in mice, limitations include high production costs because of its large size. We previously identified a small peptide derived from FGF2 that mitigated acute radiation syndrome as well as full-length FGF2. Based on the sequence of the FGF2 peptide, we synthesized a corresponding 19 amino acid FGF7 peptide (FGF7p). Our objectives were to determine if systemic FGF7p triggered the downstream targets and protected against cyclophosphamide bladder injury similar to full-length FGF7. We administered FGF7p or vehicle subcutaneously (SQ) to mice subjected to no injury or intraperitoneal (IP) cyclophosphamide and harvested bladders 1 day after injury. We then performed hematoxylin and eosin, TUNEL and immunofluorescence (IF) staining. In uninjured mice, a 20 mg/kg threshold FGF7p dose induced expression of phosphorylated (activated) FRS2α (pFRS2α), and pAKT in urothelium (consistent with cytoprotective effects of FGF7). We then gave FGF7p (20 mg/kg) or vehicle at 72 and 48 h prior to cyclophosphamide. One day after injury, TUNEL staining revealed many more apoptotic urothelial cells with vehicle treatment versus FGF7p treatment. IF for pAKT and readouts of two anti-apoptotic AKT targets (BAD and mTORC1) revealed minimal staining with vehicle treatment, but strong urothelial expression for all markers with FGF7p treatment. In conclusion, FGF7p appears to block bladder urothelial apoptosis via AKT and its targets, similar to FGF7. FGF7p is much more inexpensive to make and has a longer shelf life and higher purity than FGF7.

Measuring Radiation Toxicity Using Circulating Cell-Free DNA in Prostate Cancer Patients.

BACKGROUND: After radiation therapy (RT), circulating plasma cell-free DNA (cfDNA) released in response to RT damage to tissue can be measured within hours. We examined for a correlation between cfDNA measured during the first week of therapy and early and late gastrointestinal (GI) and genitourinary (GU) toxicity. MATERIAL AND METHODS: Patients were eligible for enrollment if they planned to receive proton or photon RT for nonmetastatic prostate cancer in the setting of an intact prostate or after prostatectomy. Blood was collected before treatment and on sequential treatment days for the first full week of therapy. Toxicity assessments were performed at baseline, weekly during RT, and 6 months and 12 months after RT. Data were analyzed to examine correlations among patient-reported GI and GU toxicities. RESULTS: Fifty-four patients were evaluable for this study. Four (7%) and 3 (6%) patients experienced acute and late grade 2 GI toxicity, respectively. Twenty-two (41%) and 18 (35%) patients experienced acute and late grade 2 GU toxicity, respectively. No patients developed grade 3 or higher toxicity. Grade 2 acute GI toxicity, but not grade 2 acute GU toxicity, was significantly correlated with pre-RT cfDNA levels and on all days 1, 2, 3, 4, and 5 of RT (P < .005). Grade 2 late GI toxicity, but not GU toxicity, was significantly correlated with pre-RT cfDNA levels (P = .021). CONCLUSIONS: Based on this preliminary study, cfDNA levels can potentially predict the subset of patients destined to develop GI toxicity during prostate cancer treatment. Given that the toxicity profiles of the various fractionations and modalities are highly similar, the data support the expectation that cfDNA could provide a biological estimate to complement the dose-volume histogram. A test of this hypothesis is under evaluation in a National Cancer Institute-funded multi-institutional study.

Circulating Cell-Free DNA Correlates with Body Integral Dose and Radiation Modality in Prostate Cancer.

PURPOSE: The RadTox assay measures circulating cell-free DNA released in response to radiotherapy (RT)-induced tissue damage. The primary objectives for this clinical trial were to determine whether cell-free DNA numbers measured by the RadTox assay are (1) correlated with body integral dose, (2) lower with proton RT compared with photon RT, and (3) higher with larger prostate cancer RT fields. PATIENTS AND METHODS: Patients planned to receive proton or photon RT for nonmetastatic prostate cancer in the setting of an intact prostate or postprostatectomy were eligible for the trial. Plasma was collected pre-RT and at 5 additional daily collection points beginning 24 hours after the initiation of RT. Data from 54 evaluable patients were analyzed to examine any correlations among RadTox scores with body-integral dose, RT modality (photon versus proton), and RT field size (prostate or prostate bed versus whole pelvis). RESULTS: Body integral dose was significantly associated with the peak post-RT RadTox score (P = .04). Patients who received photon RT had a significant increase in peak post-RT RadTox score (P = .04), average post-RT RadTox score (P = .04), and day-2 RadTox score (all minus the pre-RT values for each patient) as compared with patients who received proton RT. Field size was not significantly associated with RadTox score. CONCLUSION: RadTox is correlated with body integral dose and correctly predicts which patients receive proton versus photon RT. Data collection remains ongoing for patient-reported RT toxicity outcomes to determine whether RadTox scores are correlated with toxicity.

Scientific and Logistical Considerations When Screening for Radiation Risks by Using Biodosimetry Based on Biological Effects of Radiation Rather than Dose: The Need for Prior Measurements of Homogeneity and Distribution of Dose.

An effective medical response to a large-scale radiation event requires prompt and effective initial triage so that appropriate care can be provided to individuals with significant risk for severe acute radiation injury. Arguably, it would be advantageous to use injury rather than radiation dose for the initial assessment; i.e., use bioassays of biological damage. Such assays would be based on changes in intrinsic biological response elements; e.g., up- or down-regulation of genes, proteins, metabolites, blood cell counts, chromosomal aberrations, micronuclei, micro-RNA, cytokines, or transcriptomes. Using a framework to evaluate the feasibility of biodosimetry for triaging up to a million people in less than a week following a major radiation event, Part 1 analyzes the logistical feasibility and clinical needs for ensuring that biomarkers of organ-specific injury could be effectively used in this context. We conclude that the decision to use biomarkers of organ-specific injury would greatly benefit by first having independent knowledge of whether the person's exposure was heterogeneous and, if so, what was the dose distribution (to determine which organs were exposed to high doses). In Part 2, we describe how these two essential needs for prior information (heterogeneity and dose distribution) could be obtained by using in vivo nail dosimetry. This novel physical biodosimetry method can also meet the needs for initial triage, providing non-invasive, point-of-care measurements made by non-experts with immediate dose estimates for four separate anatomical sites. Additionally, it uniquely provides immediate information as to whether the exposure was homogeneous and, if not, it can estimate the dose distribution. We conclude that combining the capability of methods such as in vivo EPR nail dosimetry with bioassays to predict organ-specific damage would allow effective use of medical resources to save lives.

Developments in Biodosimetry Methods for Triage With a Focus on X-band Electron Paramagnetic Resonance In Vivo Fingernail Dosimetry.

Instrumentation and application methodologies for rapidly and accurately estimating individual ionizing radiation dose are needed for on-site triage in a radiological/nuclear event. One such methodology is an in vivo X-band, electron paramagnetic resonance, physically based dosimetry method to directly measure the radiation-induced signal in fingernails. The primary components under development are key instrument features, such as resonators with unique geometries that allow for large sampling volumes but limit radiation-induced signal measurements to the nail plate, and methodological approaches for addressing interfering signals in the nail and for calibrating dose from radiation-induced signal measurements. One resonator development highlighted here is a surface resonator array designed to reduce signal detection losses due to the soft tissues underlying the nail plate. Several surface resonator array geometries, along with ergonomic features to stabilize fingernail placement, have been tested in tissue-equivalent nail models and in vivo nail measurements of healthy volunteers using simulated radiation-induced signals in their fingernails. These studies demonstrated radiation-induced signal detection sensitivities and quantitation limits approaching the clinically relevant range of ≤ 10 Gy. Studies of the capabilities of the current instrument suggest that a reduction in the variability in radiation-induced signal measurements can be obtained with refinements to the surface resonator array and ergonomic features of the human interface to the instrument. Additional studies are required before the quantitative limits of the assay can be determined for triage decisions in a field application of dosimetry. These include expanded in vivo nail studies and associated ex vivo nail studies to provide informed approaches to accommodate for a potential interfering native signal in the nails when calculating the radiation-induced signal from the nail plate spectral measurements and to provide a method for calibrating dose estimates from the radiation-induced signal measurements based on quantifying experiments in patients undergoing total-body irradiation or total-skin electron therapy.

POSSIBLE NATURE OF THE RADIATION-INDUCED SIGNAL IN NAILS: HIGH-FIELD EPR, CONFIRMING CHEMICAL SYNTHESIS, AND QUANTUM CHEMICAL CALCULATIONS.

Exposure of finger- and toe-nails to ionizing radiation generates an Electron Paramagnetic Resonance (EPR) signal whose intensity is dose dependent and stable at room temperature for several days. The dependency of the radiation-induced signal (RIS) on the received dose may be used as the basis for retrospective dosimetry of an individual's fortuitous exposure to ionizing radiation. Two radiation-induced signals, a quasi-stable (RIS2) and stable signal (RIS5), have been identified in nails irradiated up to a dose of 50 Gy. Using X-band EPR, both RIS signals exhibit a singlet line shape with a line width around 1.0 mT and an apparent g-value of 2.0044. In this work, we seek information on the exact chemical nature of the radiation-induced free radicals underlying the signal. This knowledge may provide insights into the reason for the discrepancy in the stabilities of the two RIS signals and help develop strategies for stabilizing the radicals in nails or devising methods for restoring the radicals after decay. In this work an analysis of high field (94 GHz and 240 GHz) EPR spectra of the RIS using quantum chemical calculations, the oxidation-reduction properties and the pH dependence of the signal intensities are used to show that spectroscopic and chemical properties of the RIS are consistent with a semiquinone-type radical underlying the RIS. It has been suggested that semiquinone radicals formed on trace amounts of melanin in nails are the basis for the RIS signals. However, based on the quantum chemical calculations and chemical properties of the RIS, it is likely that the radicals underlying this signal are generated from the radiolysis of L-3,4-dihydroxyphenylalanine (DOPA) amino acids in the keratin proteins. These DOPA amino acids are likely formed from the exogenous oxidation of tyrosine in keratin by the oxygen from the air prior to irradiation. We show that these DOPA amino acids can work as radical traps, capturing the highly reactive and unstable sulfur-based radicals and/or alkyl radicals generated during the radiation event and are converted to the more stable o-semiquinone anion-radicals. From this understanding of the oxidation-reduction properties of the RIS, it may be possible to regenerate the unstable RIS2 following its decay through treatment of nail clippings. However, the treatment used to recover the RIS2 also has the ability to recover an interfering, mechanically-induced signal (MIS) formed when the nail is clipped. Therefore, to use the recovered (regenerated) RIS2 to increase the detection limits and precision of the RIS measurements and, therefore, the dose estimates calculated from the RIS signal amplitudes, will require the application of methods to differentiate the RIS2 from the recovered MIS signal.

Dielectric-Backed Aperture Resonators for X-Band in vivo EPR Nail Dosimetry.

A new resonator for X-band in vivo EPR nail dosimetry, the dielectric-backed aperture resonator (DAR), is developed based on rectangular TE102 geometry. This novel geometry for surface spectroscopy improves at least a factor of 20 compared to a traditional non-backed aperture resonator. Such an increase in EPR sensitivity is achieved by using a non-resonant dielectric slab, placed on the aperture inside the cavity. The dielectric slab provides an increased magnetic field at the aperture and sample, while minimizing sensitive aperture resonance conditions. This work also introduces a DAR semi-spherical (SS)-TE011 geometry. The SS-TE011 geometry is attractive due to having twice the incident magnetic field at the aperture for a fixed input power. It has been shown that DAR provides sufficient sensitivity to make biologically relevant measurements both in vitro and in vivo Although in vivo tests have shown some effects of physiological motions that suggest the necessity of a more robust finger holder, equivalent dosimetry sensitivity of approximately 1.4 Gy has been demonstrated.

Evaluating the Special Needs of The Military for Radiation Biodosimetry for Tactical Warfare Against Deployed Troops: Comparing Military to Civilian Needs for Biodosimetry Methods.

The aim of this paper is to delineate characteristics of biodosimetry most suitable for assessing individuals who have potentially been exposed to significant radiation from a nuclear device explosion when the primary population targeted by the explosion and needing rapid assessment for triage is civilians vs. deployed military personnel. The authors first carry out a systematic analysis of the requirements for biodosimetry to meet the military's needs to assess deployed troops in a warfare situation, which include accomplishing the military mission. Then the military's special capabilities to respond and carry out biodosimetry for deployed troops in warfare are compared and contrasted systematically, in contrast to those available to respond and conduct biodosimetry for civilians who have been targeted by terrorists, for example. Then the effectiveness of different biodosimetry methods to address military vs. civilian needs and capabilities in these scenarios was compared and, using five representative types of biodosimetry with sufficient published data to be useful for the simulations, the number of individuals are estimated who could be assessed by military vs. civilian responders within the timeframe needed for triage decisions. Analyses based on these scenarios indicate that, in comparison to responses for a civilian population, a wartime military response for deployed troops has both more complex requirements for and greater capabilities to use different types of biodosimetry to evaluate radiation exposure in a very short timeframe after the exposure occurs. Greater complexity for the deployed military is based on factors such as a greater likelihood of partial or whole body exposure, conditions that include exposure to neutrons, and a greater likelihood of combined injury. These simulations showed, for both the military and civilian response, that a very fast rate of initiating the processing (24,000 d) is needed to have at least some methods capable of completing the assessment of 50,000 people within a 2- or 6-d timeframe following exposure. This in turn suggests a very high capacity (i.e., laboratories, devices, supplies and expertise) would be necessary to achieve these rates. These simulations also demonstrated the practical importance of the military's superior capacity to minimize time to transport samples to offsite facilities and use the results to carry out triage quickly. Assuming sufficient resources and the fastest daily rate to initiate processing victims, the military scenario revealed that two biodosimetry methods could achieve the necessary throughput to triage 50,000 victims in 2 d (i.e., the timeframe needed for injured victims), and all five achieved the targeted throughput within 6 d. In contrast, simulations based on the civilian scenario revealed that no method could process 50,000 people in 2 d and only two could succeed within 6 d.

A microwave resonator for limiting depth sensitivity for electron paramagnetic resonance spectroscopy of surfaces.

A microwave Surface Resonator Array (SRA) structure is described for use in Electron Paramagnetic Resonance (EPR) spectroscopy. The SRA has a series of anti-parallel transmission line modes that provides a region of sensitivity equal to the cross-sectional area times its depth sensitivity, which is approximately half the distance between the transmission line centers. It is shown that the quarter-wave twin-lead transmission line can be a useful element for design of microwave resonators at frequencies as high as 10 GHz. The SRA geometry is presented as a novel resonator for use in surface spectroscopy where the region of interest is either surrounded by lossy material, or the spectroscopist wishes to minimize signal from surrounding materials. One such application is in vivo spectroscopy of human finger-nails at X-band (9.5 GHz) to measure ionizing radiation dosages. In order to reduce losses associated with tissues beneath the nail that yield no EPR signal, the SRA structure is designed to limit depth sensitivity to the thickness of the fingernail. Another application, due to the resonator geometry and limited depth penetration, is surface spectroscopy in coating or material science. To test this application, a spectrum of 1.44 µM of Mg(2+) doped polystyrene 1.1 mm thick on an aluminum surface is obtained. Modeling, design, and simulations were performed using Wolfram Mathematica (Champaign, IL; v. 9.0) and Ansys High Frequency Structure Simulator (HFSS; Canonsburg, PA; v. 15.0). A micro-strip coupling circuit is designed to suppress unwanted modes and provide a balanced impedance transformation to a 50 Ω coaxial input. Agreement between simulated and experimental results is shown.

Development and validation of an ex vivo electron paramagnetic resonance fingernail biodosimetric method.

There is an imperative need to develop methods that can rapidly and accurately determine individual exposure to radiation for screening (triage) populations and guiding medical treatment in an emergency response to a large-scale radiological/nuclear event. To this end, a number of methods that rely on dose-dependent chemical and/or physical alterations in biomaterials or biological responses are in various stages of development. One such method, ex vivo electron paramagnetic resonance (EPR) nail dosimetry using human nail clippings, is a physical biodosimetry technique that takes advantage of a stable radiation-induced signal (RIS) in the keratin matrix of fingernails and toenails. This dosimetry method has the advantages of ubiquitous availability of the dosimetric material, easy and non-invasive sampling, and the potential for immediate and rapid dose assessment. The major challenge for ex vivo EPR nail dosimetry is the overlap of mechanically induced signals and the RIS. The difficulties of analysing the mixed EPR spectra of a clipped irradiated nail were addressed in the work described here. The following key factors lead to successful spectral analysis and dose assessment in ex vivo EPR nail dosimetry: (1) obtaining a thorough understanding of the chemical nature, the decay behaviour, and the microwave power dependence of the EPR signals, as well as the influence of variation in temperature, humidity, water content, and O2 level; (2) control of the variability among individual samples to achieve consistent shape and kinetics of the EPR spectra; (3) use of correlations between the multiple spectral components; and (4) use of optimised modelling and fitting of the EPR spectra to improve the accuracy and precision of the dose estimates derived from the nail spectra. In the work described here, two large clipped nail datasets were used to test the procedures and the spectral fitting model of the results obtained with it. A 15-donor nail set with 90 nail samples from 15 donors was used to validate the sample handling and spectral analysis methods that have been developed but without the interference of a native background signal. Good consistency has been obtained between the actual RIS and the estimated RIS computed from spectral analysis. In addition to the success in RIS estimation, a linear dose response has also been achieved for all individuals in this study, where the radiation dose ranges from 0 to 6 Gy. A second 16-donor nail set with 96 nail samples was used to test the spectral fitting model where the background signal was included during the fitting of the clipped nail spectra data. Although the dose response for the estimated and actual RIS calculated in both donor nail sets was similar, there was an increased variability in the RIS values that was likely due to the variability in the background signal between donors. Although the current methods of sample handling and spectral analysis show good potential for estimating the RIS in the EPR spectra of nail clippings, there is a remaining degree of variability in the RIS estimate that needs to be addressed; this should be achieved by identifying and accounting for demographic sources of variability in the background nail signal and the composition of the nail matrix.

Clinical EPR: unique opportunities and some challenges.

Electron paramagnetic resonance (EPR) spectroscopy has been well established as a viable technique for measurement of free radicals and oxygen in biological systems, from in vitro cellular systems to in vivo small animal models of disease. However, the use of EPR in human subjects in the clinical setting, although attractive for a variety of important applications such as oxygen measurement, is challenged with several factors including the need for instrumentation customized for human subjects, probe, and regulatory constraints. This article describes the rationale and development of the first clinical EPR systems for two important clinical applications, namely, measurement of tissue oxygen (oximetry) and radiation dose (dosimetry) in humans. The clinical spectrometers operate at 1.2 GHz frequency and use surface-loop resonators capable of providing topical measurements up to 1 cm depth in tissues. Tissue pO2 measurements can be carried out noninvasively and repeatedly after placement of an oxygen-sensitive paramagnetic material (currently India ink) at the site of interest. Our EPR dosimetry system is capable of measuring radiation-induced free radicals in the tooth of irradiated human subjects to determine the exposure dose. These developments offer potential opportunities for clinical dosimetry and oximetry, which include guiding therapy for individual patients with tumors or vascular disease by monitoring of tissue oxygenation. Further work is in progress to translate this unique technology to routine clinical practice.

Electron paramagnetic resonance dosimetry for a large-scale radiation incident.

With possibilities for radiation terrorism and intensified concerns about nuclear accidents since the recent Fukushima Daiichi event, the potential exposure of large numbers of individuals to radiation that could lead to acute clinical effects has become a major concern. For the medical community to cope with such an event and avoid overwhelming the medical care system, it is essential to identify not only individuals who have received clinically significant exposures and need medical intervention but also those who do not need treatment. The ability of electron paramagnetic resonance to measure radiation-induced paramagnetic species, which persist in certain tissues (e.g., teeth, fingernails, toenails, bone, and hair), has led to this technique becoming a prominent method for screening significantly exposed individuals. Although the technical requirements needed to develop this method for effective application in a radiation event are daunting, remarkable progress has been made. In collaboration with General Electric and through funding committed by the Biomedical Advanced Research and Development Authority, electron paramagnetic resonance tooth dosimetry of the upper incisors is being developed to become a Food and Drug Administration-approved and manufacturable device designed to carry out triage for a threshold dose of 2 Gy. Significant progress has also been made in the development of electron paramagnetic resonance nail dosimetry based on measurements of nails in situ under point-of-care conditions, and in the near future this may become a second field-ready technique. Based on recent progress in measurements of nail clippings, it is anticipated that this technique may be implementable at remotely located laboratories to provide additional information when the measurements of dose on-site need to be supplemented. The authors conclude that electron paramagnetic resonance dosimetry is likely to be a useful part of triage for a large-scale radiation incident.

Advances towards using finger/toenail dosimetry to triage a large population after potential exposure to ionizing radiation.

Rapid and accurate retrospective dosimetry is of critical importance and strategic value for the emergency medical response to a large-scale radiological/nuclear event. One technique that has the potential for rapid and accurate dosimetry measurements is electron paramagnetic resonance (EPR) spectroscopy of relatively stable radiation-induced signals (RIS) in fingernails and toenails. Two approaches are being developed for EPR nail dosimetry. In the approach using ex vivo measurements on nail clippings, accurate estimation of the dose-dependent amplitude of the RIS is complicated by the presence of mechanically-induced signals (MIS) that are generated during the nail clipping. Recent developments in ex vivo nail dosimetry, including a thorough characterization of the MIS and an appreciation of the role of hydration and the development of effective analytic techniques, have led to improvements in the accuracy and precision of this approach. An in vivo nail dosimetry approach is also very promising, as it eliminates the problems of MIS from the clipping and it has the potential to be an effective and efficient approach for field deployment. Two types of EPR resonators are being developed for in vivo measurements of fingernails and toenails.

Mechanisms of direct radiation damage to DNA: the effect of base sequence on base end products.

It has been generally assumed that product formation in DNA damaged by ionizing radiation is relatively independent of base sequence, i.e., that the yield of a given product depends primarily on the chemical properties of each DNA constituent and not on its base sequence context. We examined this assumption by comparing direct-type end products produced in films of d(CTCTCGAGAG)(2) with those produced in films of d(GCACGCGTGC)(2). Here we report the product yields in d(CTCTCGAGAG)(2) hydrated to Γ = 2.5 and 15, where Γ is the hydration level given in moles of H(2)O/mole of nucleotide. Of the 17 products monitored by GC/MS, seven exhibited statistically significant yields: 8-oxoGua, 8-oxoAde, 5-OHMeUra, 5,6-diHUra, 5,6-diHThy, 5-OHCyt, and 5-OHUra. These yields at Γ = 2.5 are compared with the yields from our previously reported study of d(GCACGCGTGC)(2) (after projecting the yields to a CG/AT ratio of 1). The ratio of projected yields, d(CTCTCGAGAG)(2) divided by d(GCACGCGTGC)(2), are 1.3 ± 0.9, 1.8 ± 0.3, 1.6 ± 0.6, 11.4 ± 4.7, 0.2 ± 0.1, >28, and 0.8 ± 1.1, respectively. Considering just d(CTCTCGAGAG)(2), the ratios of yields at Γ = 2.5 divided by yields at Γ = 15 are 0.7 ± 0.2, 0.5 ± 0.1, 2.3 ± 4.0, 3.4 ± 1.2, 3.5 ± 3.3, 1.2 ± 0.2, and 0.4 ± 0.2, respectively. The effects of sequence and hydration on base product yields are explained by a working model emphasizing the difference between two distinctly different types of reaction: (i) radical reactions that progress to nonradical intermediates and product prior to dissolution and (ii) reactions that stem from radicals trapped in the solid state at room temperature that go on to yield nonradical product after sample dissolution. Based on these findings, insights into rates of hole and excess electron-transfer relative to rates of proton transfer are discussed.

Antioxidant properties of select radiation mitigators based on semicarbazone and pyrazole derivatives of curcumin.

Fifty-eight semicarbazone and pyrazole derivatives of curcumin have been developed as potential mitigation agents to treat acute radiation syndrome (ARS). Pyridyl (D12, D13), furyl (D56), and phenyl (D68) derivatives of curcumin semi-carbazones were found to provide the highest dose modifying factors (DMF) with respect to survival in sub-TBI (bone marrow sparing) exposures in mouse models. To investigate the basis for the mitigating effects of these agents on ARS, we examined their oxidation potentials and radical scavenging properties in comparison to other semicarbazone and pyrazole curcumin derivatives with less effective DMFs. Comparisons between D12, D13, D56, and D68 and other semicarbazone and pyrazole derivatives of curcumin did not show a sufficient difference in reducing properties and hydrogen atom donating properties for these properties to be the basis of the dose modifying activities of these compounds. Therefore, their DMFs likely reflect structure-activity relationship(s),wherein interaction with key receptors or alteration of enzyme expression result in modifications of cellular or tissue responses to radiation, rather than on the derivatives' ability to modify radiation-induced flux of free radicals through direct interaction with these radicals.