Track Structure-Based Simulations on DNA Damage Induced by Diverse Isotopes.

Diverse isotopes such as 2H, 3He, 10Be, 11C and 14C occur in nuclear reactions in ion beam radiotherapy, in cosmic ray shielding, or are intentionally accelerated in dating techniques. However, only a few studies have specifically addressed the biological effects of diverse isotopes and were limited to energies of several MeV/u. A database of simulations with the PARTRAC biophysical tool is presented for H, He, Li, Be, B and C isotopes at energies from 0.5 GeV/u down to stopping. The doses deposited to a cell nucleus and also the yields per unit dose of single- and double-strand breaks and their clusters induced in cellular DNA are predicted to vary among diverse isotopes of the same element at energies < 1 MeV/u, especially for isotopes of H and He. The results may affect the risk estimates for astronauts in deep space missions or the models of biological effectiveness of ion beams and indicate that radiation protection in 14C or 10Be dating techniques may be based on knowledge gathered with 12C or 9Be.

Internal microdosimetry of alpha-emitting radionuclides.

At the tissue level, energy deposition in cells is determined by the microdistribution of alpha-emitting radionuclides in relation to sensitive target cells. Furthermore, the highly localized energy deposition of alpha particle tracks and the limited range of alpha particles in tissue produce a highly inhomogeneous energy deposition in traversed cell nuclei. Thus, energy deposition in cell nuclei in a given tissue is characterized by the probability of alpha particle hits and, in the case of a hit, by the energy deposited there. In classical microdosimetry, the randomness of energy deposition in cellular sites is described by a stochastic quantity, the specific energy, which approximates the macroscopic dose for a sufficiently large number of energy deposition events. Typical examples of the alpha-emitting radionuclides in internal microdosimetry are radon progeny and plutonium in the lungs, plutonium and americium in bones, and radium in targeted radionuclide therapy. Several microdosimetric approaches have been proposed to relate specific energy distributions to radiobiological effects, such as hit-related concepts, LET and track length-based models, effect-specific interpretations of specific energy distributions, such as the dual radiation action theory or the hit-size effectiveness function, and finally track structure models. Since microdosimetry characterizes only the initial step of energy deposition, microdosimetric concepts are most successful in exposure situations where biological effects are dominated by energy deposition, but not by subsequently operating biological mechanisms. Indeed, the simulation of the combined action of physical and biological factors may eventually require the application of track structure models at the nanometer scale.

Track structure based modelling of chromosome aberrations after photon and alpha-particle irradiation.

A computational model of radiation-induced chromosome aberrations in human cells within the PARTRAC Monte Carlo simulation framework is presented. The model starts from radiation-induced DNA damage assessed by overlapping radiation track structures with multi-scale DNA and chromatin models, ranging from DNA double-helix in atomic resolution to chromatin fibre loops, heterochromatic and euchromatic regions, and chromosome territories. The repair of DNA double-strand breaks via non-homologous end-joining is followed. Initial spatial distribution and complexity, diffusive motion, enzymatic processing, synapsis and ligation of individual DNA ends from the breaks are simulated. To enable scoring of different chromosome aberration types resulting from improper joining of DNA fragments, the repair module has been complemented by tracking the chromosome origin of the ligated fragments and the positions of centromeres. The modelled motion of DNA ends has sub-diffusive characteristics and corresponds to measured chromatin mobility within time-scales of a few hours. The calculated formation of dicentrics after photon and α-particle irradiation in human fibroblasts is compared to experimental data (Cornforth et al., 2002, Radiat Res 158, 43). The predicted yields of dicentrics overestimate the measurements by factors of five for γ-rays and two for α-particle irradiation. Nevertheless, the observed relative dependence on radiation dose is correctly reproduced. Calculated yields and size distributions of other aberration types are discussed. The present work represents a first mechanistic approach to chromosome aberrations and their kinetics, combining full track structure simulations with detailed models of chromatin and accounting for the kinetics of DNA repair.

Mechanistic modelling suggests that the size of preneoplastic lesions is limited by intercellular induction of apoptosis in oncogenically transformed cells.

Selective removal of oncogenically transformed cells by apoptosis induced via signalling by surrounding cells has been suggested to represent a natural anticarcinogenic process. To investigate its potential effect in detail, a mechanistic model of this process is proposed. The model is calibrated against in vitro data on apoptosis triggered in transformed cells by defined external inducers as well as through signalling by normal cells under coculture conditions. The model predicts that intercellular induction of apoptosis is capable of balancing the proliferation of oncogenically transformed cells and limiting the size of their populations over long times, even if their proliferation per se were unlimited. Experimental research is desired to verify whether the predicted stable population of transformed cells corresponds to a kind of dormancy during early-stage carcinogenesis (dormant preneoplastic lesions), and how this process relates to other anticarcinogenic mechanisms taking place under in vivo conditions.

Track structures, DNA targets and radiation effects in the biophysical Monte Carlo simulation code PARTRAC.

This review describes the PARTRAC suite of comprehensive Monte Carlo simulation tools for calculations of track structures of a variety of ionizing radiation qualities and their biological effects. A multi-scale target model characterizes essential structures of the whole genomic DNA within human fibroblasts and lymphocytes in atomic resolution. Calculation methods and essential results are recapitulated regarding the physical, physico-chemical and chemical stage of track structure development of radiation damage induction. Recent model extension towards DNA repair processes extends the time dimension by about 12 orders of magnitude and paves the way for superior predictions of radiation risks.

Analytical formulas representing track-structure simulations on DNA damage induced by protons and light ions at radiotherapy-relevant energies.

Track structure based simulations valuably complement experimental research on biological effects of ionizing radiation. They provide information at the highest level of detail on initial DNA damage induced by diverse types of radiation. Simulations with the biophysical Monte Carlo code PARTRAC have been used for testing working hypotheses on radiation action mechanisms, for benchmarking other damage codes and as input for modelling subsequent biological processes. To facilitate such applications and in particular to enable extending the simulations to mixed radiation field conditions, we present analytical formulas that capture PARTRAC simulation results on DNA single- and double-strand breaks and their clusters induced in cells irradiated by ions ranging from hydrogen to neon at energies from 0.5 GeV/u down to their stopping. These functions offer a means by which radiation transport codes at the macroscopic scale could easily be extended to predict biological effects, exploiting a large database of results from micro-/nanoscale simulations, without having to deal with the coupling of spatial scales and running full track-structure calculations.

Time- and space-resolved Monte Carlo study of water radiolysis for photon, electron and ion irradiation.

Time-dependent yields of the most important products of water radiolysis E(aq)(-), (*)OH, H(*), H(3)O(+), H(2), OH(-) and H(2)O(2) have been calculated for (60)Co-photons, electrons, protons, helium- and carbon-ions incident onto water. G values have been evaluated for the interval from 1 ps to 1 mus after initial energy deposition as a function of time, as well as after 1 ns and at the end of the chemical stage as a function of linear energy transfer (LET), covering an interval from approximately 0.2 up to 750 keV/microm by means of different particle types. In this work, the modules of the biophysical Monte Carlo track structure code PARTRAC dealing with the simulation of prechemical and chemical stages have been improved to extend interaction data sets for heavier ions. Among other newly selected parameter values, the thermalisation distance between the point of generation and hydration of subexcitation electrons has been adopted from recent data in the literature. As far as data from the literature are available, good agreement has been found with the calculated time- and LET-dependent yields in this work, supporting the selection of the revised parameter values.

Interaction of ion tracks in spatial and temporal proximity.

In the present work, a systematic analysis of the impact of spatial and temporal proximity of ion tracks on the yield of higher-order radiolytic species as well as of DNA damage patterns is presented. This potential impact may be of concern when laser-driven particle accelerators are used for ion radiation therapy. The biophysical Monte Carlo track structure code PARTRAC was used and, to this end, extended in two aspects: first, the temporal information about track evolution has been included in the track structure module and, second, the simulation code has been modified to enable parallel multiple track processing during simulation of subsequent modelling stages. Depending on the spatial and temporal separation between ion-track pairs, the yield of chemical species has been calculated for incident protons with start energies of 20 MeV, for He(2+) ions with start energies of 1 and 20 MeV, and for 60 MeV C(6+) ions. Provided the overlap of the considered ion tracks is sufficient in all four dimensions (space and time), the yield of hydroxyl radicals was found to be reduced compared to that of single tracks, for all considered ion types. The biological endpoints investigated were base damages, single-strand breaks, double-strand breaks, and clustered lesions for incident pairs of protons and He(2+) ions, each with start energies of 20 MeV. The yield of clustered lesions produced by 20 MeV protons turned out to be influenced by the spatial separation of the proton pair; in contrast, no influence was found for different start times of the protons. The yield of single-strand breaks and base hits was found neither to depend on the spatial separation nor on the temporal separation between the incident protons. For incident 20 MeV He(2+) ions, however, a dependence on the spatial and temporal separation of the ion pair was found for all considered biological endpoints. Nevertheless, spatial proximity conditions where such intertrack effects were obtained are not met in the case of tumour radiation therapy; thus, no impact on radiation effects due to short pulse duration of laser-driven accelerators can be expected from alterations during the chemical stage.

MECHANISTIC MODELING PREDICTS ANTI-CARCINOGENIC RADIATION EFFECTS ON INTERCELLULAR SIGNALING IN VITRO TURN PRO-CARCINOGENIC IN VIVO.

Oncogenic transformed cells represent an in vitro system mimicking early-stage carcinogenesis. These precancerous cells are subject to a selective removal via apoptosis induced by neighbor cells. By modulating the underpinning intercellular signaling mediated by cytokines and reactive oxygen/nitrogen species, ionizing radiation enhances this removal of precancerous cells in vitro, at doses from a few mGy to a few Gy. However, epidemiological data demonstrate that radiation exposure induces cancer, at least above 100 mGy. Mechanistic modeling of the given anti-carcinogenic process explains this discrepancy: The model reproduces in vitro data on apoptosis and its enhancement by radiation. For in vivo-like conditions with signal lifetimes shorter and cell densities higher than in vitro, radiation is predicted to reduce this anti-carcinogenic mechanism. Early-stage lesions that would be turned dormant or completely removed may grow large and escape this control mechanism upon irradiation.

BIOPHYSICAL SIMULATION TOOL PARTRAC: MODELLING PROTON BEAMS AT THERAPY-RELEVANT ENERGIES.

The biophysical simulation tool PARTRAC has been primarily developed to model radiation physics, chemistry and biology on nanometre to micrometre scales. However, the tool can be applied in simulating radiation effects in an event-by-event manner over macroscopic volumes as well. Benchmark simulations are reported showing that PARTRAC does reproduce the macroscopic Bragg peaks of proton beams, although the penetration depths are underestimated by a few per cent for high-energy beams. PARTRAC also quantifies the increase in DNA damage and its complexity along the beam penetration depth. Enhanced biological effectiveness is predicted in particular within distal Bragg peak parts of therapeutic proton beams.

Track-structure simulations of energy deposition patterns to mitochondria and damage to their DNA.

PURPOSE: Mitochondria have been implicated in initiating and/or amplifying the biological effects of ionizing radiation not mediated via damage to nuclear DNA. To help elucidate the underlying mechanisms, energy deposition patterns to mitochondria and radiation damage to their DNA have been modelled. METHODS: Track-structure simulations have been performed with PARTRAC biophysical tool for 60Co γ-rays and 5 MeV α-particles. Energy deposition to the cell's mitochondria has been analyzed. A model of mitochondrial DNA reflecting experimental information on its structure has been developed and used to assess its radiation-induced damage. RESULTS: Energy deposition to mitochondria is highly inhomogeneous, especially at low doses. Although a dose-dependent fraction of mitochondria sees no energy deposition at all, the hit ones receive rather high amounts of energy. Nevertheless, only little damage to mitochondrial DNA occurs, even at large doses. CONCLUSION: Mitochondrial DNA does not represent a critical target for radiation effects. Likely, the key role of mitochondria in radiation-induced biological effects arises from the communication between mitochondria and/or with the nucleus. Through this signaling, initial modifications in a few heavily hit mitochondria seem to be amplified to a massive long-term effect manifested in the whole cell or even tissue.

Predicting DNA damage foci and their experimental readout with 2D microscopy: a unified approach applied to photon and neutron exposures.

The consideration of how a given technique affects results of experimental measurements is a must to achieve correct data interpretation. This might be challenging when it comes to measurements on biological systems, where it is unrealistic to have full control (e.g. through a software replica) of all steps in the measurement chain. In this work we address how the effectiveness of different radiation qualities in inducing biological damage can be assessed measuring DNA damage foci yields, only provided that artefacts related to the scoring technique are adequately considered. To this aim, we developed a unified stochastic modelling approach that, starting from radiation tracks, predicts both the induction, spatial distribution and complexity of DNA damage, and the experimental readout of foci when immunocytochemistry coupled to 2D fluorescence microscopy is used. The approach is used to interpret γ-H2AX data for photon and neutron exposures. When foci are reconstructed in the whole cell nucleus, we obtain information on damage characteristics "behind" experimental observations, as the average damage content of a focus. We reproduce how the detection technique affects experimental findings, e.g. contributing to the saturation of foci yields scored at 30 minutes after exposure with increasing dose and to the lack of dose dependence for yields at 24 hours.

MODELLING γ-H2AX FOCI INDUCTION TO MIMIC LIMITATIONS IN THE SCORING TECHNIQUE.

An approach based on track-structure calculations has been developed to take account of artefacts occurring during γ-H2AX foci detection in 2D images of samples analyzed through immunocytochemistry. The need of this works stems from the observed saturation in foci yields measured after X-ray doses higher than few grays, hindering an unambiguous quantification of DNA damage and of radiation effectiveness. The proposed modelling approach allows to simulate the observer's point of view for foci scoring, mimicking the selection of a slice Δz of the cell nucleus due to the microscope depth of field, and applying a clustering algorithm to group together damages within a resolution parameter r. Calculation results were benchmarked with experimental measurements at an early time-point for mouse breast cancer cells, irradiated with X-ray doses in the range 0-5 Gy. The model is able to reproduce the saturation in experimental data.

The COOLER Code: A Novel Analytical Approach to Calculate Subcellular Energy Deposition by Internal Electron Emitters.

COmputation Of Local Electron Release (COOLER), a software program has been designed for dosimetry assessment at the cellular/subcellular scale, with a given distribution of administered low-energy electron-emitting radionuclides in cellular compartments, which remains a critical step in risk/benefit analysis for advancements in internal radiotherapy. The software is intended to overcome the main limitations of the medical internal radiation dose (MIRD) formalism for calculations of cellular S-values (i.e., dose to a target region in the cell per decay in a given source region), namely, the use of the continuous slowing down approximation (CSDA) and the assumption of a spherical cell geometry. To this aim, we developed an analytical approach, entrusted to a MATLAB-based program, using as input simulated data for electron spatial energy deposition directly derived from full Monte Carlo track structure calculations with PARTRAC. Results from PARTRAC calculations on electron range, stopping power and residual energy versus traveled distance curves are presented and, when useful for implementation in COOLER, analytical fit functions are given. Example configurations for cells in different culture conditions (V79 cells in suspension or adherent culture) with realistic geometrical parameters are implemented for use in the tool. Finally, cellular S-value predictions by the newly developed code are presented for different cellular geometries and activity distributions (uniform activity in the nucleus, in the entire cell or on the cell surface), validated against full Monte Carlo calculations with PARTRAC, and compared to MIRD standards, as well as results based on different track structure calculations (Geant4-DNA). The largest discrepancies between COOLER and MIRD predictions were generally found for electrons between 25 and 30 keV, where the magnitude of disagreement in S-values can vary from 50 to 100%, depending on the activity distribution. In calculations for activity distribution on the cell surface, MIRD predictions appeared to fail the most. The proposed method is suitable for Auger-cascade electrons, but can be extended to any energy of interest and to beta spectra; as an example, the 3H case is also discussed. COOLER is intended to be accessible to everyone (preclinical and clinical researchers included), and may provide important information for the selection of radionuclides, the interpretation of radiobiological or preclinical results, and the general establishment of doses in any scenario, e.g., with cultured cells in the laboratory or with therapeutic or diagnostic applications. The software will be made available for download from the DTU-Nutech website: http://www.nutech.dtu.dk/ .

Simulation of light ion induced DNA damage patterns.

The biophysical simulation code PARTRAC was extended by a module to handle ions heavier than alpha particles. Cross sections for ion-electron interactions were taken from He(++) ions of the same velocity and scaled by Z(eff(2))/4. Calculated linear energy transfer values, radial dose distributions and secondary electron spectra were found in agreement with experimental results. DNA damage due to irradiation of human fibroblast cells by several light ions from H to S was calculated for various energies complemented by 220 kV(p) X rays as reference radiation. With increasing linear energy transfer, the calculated total yield of double-strand breaks per dose showed saturation behaviour at about twice the value for reference radiation. When data analysis methods for experimental double-strand break yield determination were applied to the simulated DNA damage patterns, the two data sets were found in accord. The calculated patterns of DNA damage clusters were analysed on local and regional scale finding regional clusters in closer correlation to experimental cell inactivation data.

Enhanced release of primary signals may render intercellular signalling ineffective due to spatial aspects.

Detailed mechanistic modelling has been performed of the intercellular signalling cascade between precancerous cells and their normal neighbours that leads to a selective removal of the precancerous cells by apoptosis. Two interconnected signalling pathways that were identified experimentally have been modelled, explicitly accounting for temporal and spatial effects. The model predicts highly non-linear behaviour of the signalling. Importantly, under certain conditions, enhanced release of primary signals by precancerous cells renders the signalling ineffective. This counter-intuitive behaviour arises due to spatial aspects of the underlying signalling scheme: Increased primary signalling by precancerous cells does, upon reaction with factors derived from normal cells, produce higher yields of apoptosis-triggering molecules. However, the apoptosis-triggering signals are formed farther from the precancerous cells, so that these are attacked less efficiently. Spatial effects thus may represent a novel analogue of negative feedback mechanisms.

Role of DNA organisation and environmental scavenging capacity in the evolution of radiobiological damage: models and simulations.

BACKGROUND AND PURPOSE: Theoretical models and Monte Carlo simulations were developed, aimed to investigate the role played by the organisation of interphase DNA and the environmental scavenging capacity conditions in the induction of radiobiological damage. METHODS: The induction of single- and double-strand breaks by gamma rays impinging on different DNA structures (e.g. linear DNA, SV40 minichromosome and cellular DNA) was simulated as a function of the environment scavenging capacity. Furthermore, yields of chromosome aberrations (CA) induced by gamma rays and light ions were simulated with a purposely developed MC code that explicitly takes into account the DNA higher-order organisation as chromosome territories. RESULTS AND CONCLUSIONS: Simulations performed with the PARTRAC code allowed quantification of the dependence of dsb and ssb both on the target structure, and on the scavenging capacity. The results relative to CA showed the importance of DNA damage complexity (nanometre scale) and interphase chromosome domains (micrometre scale) in the process of aberration formation. Very good agreement was found between the model predictions on ssb, dsb and CA and available experimental data.

Simulation of DNA damage after proton irradiation.

The biophysical radiation track simulation model PARTRAC was improved by implementing new interaction cross sections for protons in water. Computer-simulated tracks of energy deposition events from protons and their secondary electrons were superimposed on a higher-order DNA target model describing the spatial coordinates of the whole genome inside a human cell. Induction of DNA double-strand breaks was simulated for proton irradiation with LET values between 1.6 and 70 keV/microm and various reference radiation qualities. The yield of DSBs after proton irradiation was found to rise continuously with increasing LET up to about 20 DSBs per Gbp and Gy, corresponding to an RBE up to 2.2. About half of this increase resulted from a higher yield of DSB clusters associated with small fragments below 10 kbp. Exclusion of experimentally unresolved multiple DSBs reduced the maximum DSB yield by 30% and shifted it to an LET of about 40 keV/microm. Simulated fragment size distributions deviated significantly from random breakage distributions over the whole size range after irradiation with protons with an LET above 10 keV/microm. Determination of DSB yields using equations derived for random breakage resulted in an underestimation by up to 20%. The inclusion of background fragments had only a minor influence on the distribution of the DNA fragments induced by radiation. Despite limited numerical agreement, the simulations reproduced the trends in proton-induced DNA DSBs and fragment induction found in recent experiments.

Track structure calculations on intracellular targets responsible for signal release in bystander experiments with transfer of irradiated cell-conditioned medium.

PURPOSE: To investigate alternative scenarios for the dose-dependent emission of bystander signals by irradiated cells in medium transfer experiments. METHODS: Energy deposition patterns to hypothetical intracellular targets whose hit by radiation initiates the emission of bystander signals have been simulated by Monte Carlo code PARTRAC, evaluating the effects of target size, multiplicity and threshold energy for activation. Scenarios in which individual irradiated cells release signals independently as well as those with signal amplification by neighbour cells have been analyzed. The non-linear response of unirradiated cells to signals in the transferred medium has been considered. RESULTS: The experimentally observed dose dependence of bystander effects is consistent with cell-autonomous signal release with a wide distribution of characteristic doses, covering the range of 3 mGy to 3 Gy. Alternatively, the data can be explained by assuming that only cells receiving a high specific energy (3 Gy to 0.5 µm targets) release primary signals, which are then amplified by secondary signalling by neighbour cells within about a millimetre distance. CONCLUSION: Alternative signal emission scenarios are consistent with the observed dose dependence of bystander effects in medium transfer experiments. Thus, further experimental research is needed to identify the actual mechanism of bystander signal emission.

Non-linear response of cells to signals leads to revised characteristics of bystander effects inferred from their modelling.

PURPOSE: To analyze the response of naïve cells to bystander signals, thus complementing previous studies on signal emission by irradiated cells and improving quantitative understanding of bystander effects. MATERIALS AND METHODS: Published data on reduced clonogenic survival and mutation induction in bystander experiments with undiluted and diluted irradiated cell-conditioned medium were analyzed using linear and non-linear response functions. RESULTS: The data indicated a highly non-linear response of cells to bystander signals. It can be described with sigmoid response functions, involving only a single additional parameter compared to the linear response assumed in existing models. Accounting for this non-linearity significantly modifies bystander characteristics inferred from the modelling, such as signal lifetime or dose dependence of signal release. Some signal release models are even ruled out. CONCLUSIONS: The sigmoid response to signals reflects complex intracellular pathways triggered and, together with the non-linear release of signals, supports the involvement of cytokines and/or reactive oxygen species in bystander effects. Further research combining experimental and modelling approaches is needed to elucidate the mechanisms of intercellular communication and their modifications by radiation, in particular to determine the nature of bystander signals, dynamics of their release after irradiation, and cellular responses to these signals.