Characterisation of the UK high energy proton research beamline for high and ultra-high dose rate (FLASH) irradiation.

Objective. This work sets out the capabilities of the high energy proton research beamline developed in the Christie proton therapy centre for Ultra-High Dose Rate (UHDR) irradiation and FLASH experiments. It also characterises the lower limits of UHDR operation for this Pencil Beam Scanning (PBS) proton hardware.Approach. Energy dependent nozzle transmission was measured using a Faraday Cup beam collector. Spot size was measured at the reference plane using a 2D scintillation detector. Integrated depth doses (IDDs) were measured. EBT3 Gafchromic film was used to compare UHDR and conventional dose rate spots. Our beam monitor calibration methodolgy for UHDR is described. A microDiamond detector was used to determine dose rates at zref. Instantaneous depth dose rates were calculated for 70-245 MeV. PBS dose rate distributions were calculated using Folkerts and Van der Water definitions.Main results. Transmission of 7.05 ± 0.1% is achieveable corresponding to a peak instantaneous dose rate of 112.7 Gy s-1. Beam parameters are comparable in conventional and UHDR mode with a spot size ofσx= 4.6 mm,σy= 6.6 mm. Dead time in the beam monitoring electonics warrants a beam current dependent MU correction in the present configuration. Fast beam scanning of 26.4 m s-1(X) and 12.1 m s-1(Y) allows PBS dose rates of the order tens of Grays per second.Significance. UHDR delivery is possible for small field sizes and high energies enabling research into the FLASH effect with PBS protons at our facility. To our knowledge this is also the first thorough characterisation of UHDR irradiation using the hardware of this clinical accelerator at energies less than 250 MeV. The data set out in this publication can be used for designing experiments at this UK research facility and inform the possible future clinical translation of UHDR PBS proton therapy.

Determining the parameter space for effective oxygen depletion for FLASH radiation therapy.

There has been a recent revival of interest in the FLASH effect, after experiments have shown normal tissue sparing capabilities of ultra-high-dose-rate radiation with no compromise on tumour growth restraint. A model has been developed to investigate the relative importance of a number of fundamental parameters considered to be involved in the oxygen depletion paradigm of induced radioresistance. An example eight-dimensional parameter space demonstrates the conditions under which radiation may induce sufficient depletion of oxygen for a diffusion-limited hypoxic cellular response. Initial results support experimental evidence that FLASH sparing is only achieved for dose rates on the order of tens of Gy s-1or higher, for a sufficiently high dose, and only for tissue that is slightly hypoxic at the time of radiation. We show that the FLASH effect is the result of a number of biological, radiochemical and delivery parameters. Also, the threshold dose for a FLASH effect occurring would be more prominent when the parameterisation was optimised to produce the maximum effect. The model provides a framework for further FLASH-related investigation and experimental design. An understanding of the mechanistic interactions producing an optimised FLASH effect is essential for its translation into clinical practice.

Evaluating very high energy electron RBE from nanodosimetric pBR322 plasmid DNA damage.

This paper presents the first plasmid DNA irradiations carried out with Very High Energy Electrons (VHEE) over 100-200 MeV at the CLEAR user facility at CERN to determine the Relative Biological Effectiveness (RBE) of VHEE. DNA damage yields were measured in dry and aqueous environments to determine that ~ 99% of total DNA breaks were caused by indirect effects, consistent with other published measurements for protons and photons. Double-Strand Break (DSB) yield was used as the biological endpoint for RBE calculation, with values found to be consistent with established radiotherapy modalities. Similarities in physical damage between VHEE and conventional modalities gives confidence that biological effects of VHEE will also be similar-key for clinical implementation. Damage yields were used as a baseline for track structure simulations of VHEE plasmid irradiation using GEANT4-DNA. Current models for DSB yield have shown reasonable agreement with experimental values. The growing interest in FLASH radiotherapy motivated a study into DSB yield variation with dose rate following VHEE irradiation. No significant variations were observed between conventional and FLASH dose rate irradiations, indicating that no FLASH effect is seen under these conditions.

In Silico Models of DNA Damage and Repair in Proton Treatment Planning: A Proof of Concept.

There is strong in vitro cell survival evidence that the relative biological effectiveness (RBE) of protons is variable, with dependence on factors such as linear energy transfer (LET) and dose. This is coupled with the growing in vivo evidence, from post-treatment image change analysis, of a variable RBE. Despite this, a constant RBE of 1.1 is still applied as a standard in proton therapy. However, there is a building clinical interest in incorporating a variable RBE. Recently, correlations summarising Monte Carlo-based mechanistic models of DNA damage and repair with absorbed dose and LET have been published as the Manchester mechanistic (MM) model. These correlations offer an alternative path to variable RBE compared to the more standard phenomenological models. In this proof of concept work, these correlations have been extended to acquire RBE-weighted dose distributions and calculated, along with other RBE models, on a treatment plan. The phenomenological and mechanistic models for RBE have been shown to produce comparable results with some differences in magnitude and relative distribution. The mechanistic model found a large RBE for misrepair, which phenomenological models are unable to do. The potential of the MM model to predict multiple endpoints presents a clear advantage over phenomenological models.

Mechanistic modelling supports entwined rather than exclusively competitive DNA double-strand break repair pathway.

Following radiation induced DNA damage, several repair pathways are activated to help preserve genome integrity. Double Strand Breaks (DSBs), which are highly toxic, have specified repair pathways to address them. The main repair pathways used to resolve DSBs are Non-Homologous End Joining (NHEJ) and Homologous Recombination (HR). Cell cycle phase determines the availability of HR, but the repair choice between pathways in the G2 phases where both HR and NHEJ can operate is not clearly understood. This study compares several in silico models of repair choice to experimental data published in the literature, each model representing a different possible scenario describing how repair choice takes place. Competitive only scenarios, where initial protein recruitment determines repair choice, are unable to fit the literature data. In contrast, the scenario which uses a more entwined relationship between NHEJ and HR, incorporating protein co-localisation and RNF138-dependent removal of the Ku/DNA-PK complex, is better able to predict levels of repair similar to the experimental data. Furthermore, this study concludes that co-localisation of the Mre11-Rad50-Nbs1 (MRN) complexes, with initial NHEJ proteins must be modeled to accurately depict repair choice.

Clinically relevant nanodosimetric simulation of DNA damage complexity from photons and protons.

Relative Biological Effectiveness (RBE), the ratio of doses between radiation modalities to produce the same biological endpoint, is a controversial and important topic in proton therapy. A number of phenomenological models incorporate variable RBE as a function of Linear Energy Transfer (LET), though a lack of mechanistic description limits their applicability. In this work we take a different approach, using a track structure model employing fundamental physics and chemistry to make predictions of proton and photon induced DNA damage, the first step in the mechanism of radiation-induced cell death. We apply this model to a proton therapy clinical case showing, for the first time, predictions of DNA damage on a patient treatment plan. Our model predictions are for an idealised cell and are applied to an ependymoma case, at this stage without any cell specific parameters. By comparing to similar predictions for photons, we present a voxel-wise RBE of DNA damage complexity. This RBE of damage complexity shows similar trends to the expected RBE for cell kill, implying that damage complexity is an important factor in DNA repair and therefore biological effect.

A New Standard DNA Damage (SDD) Data Format.

Our understanding of radiation-induced cellular damage has greatly improved over the past few decades. Despite this progress, there are still many obstacles to fully understand how radiation interacts with biologically relevant cellular components, such as DNA, to cause observable end points such as cell killing. Damage in DNA is identified as a major route of cell killing. One hurdle when modeling biological effects is the difficulty in directly comparing results generated by members of different research groups. Multiple Monte Carlo codes have been developed to simulate damage induction at the DNA scale, while at the same time various groups have developed models that describe DNA repair processes with varying levels of detail. These repair models are intrinsically linked to the damage model employed in their development, making it difficult to disentangle systematic effects in either part of the modeling chain. These modeling chains typically consist of track-structure Monte Carlo simulations of the physical interactions creating direct damages to DNA, followed by simulations of the production and initial reactions of chemical species causing so-called "indirect" damages. After the induction of DNA damage, DNA repair models combine the simulated damage patterns with biological models to determine the biological consequences of the damage. To date, the effect of the environment, such as molecular oxygen (normoxic vs. hypoxic), has been poorly considered. We propose a new standard DNA damage (SDD) data format to unify the interface between the simulation of damage induction in DNA and the biological modeling of DNA repair processes, and introduce the effect of the environment (molecular oxygen or other compounds) as a flexible parameter. Such a standard greatly facilitates inter-model comparisons, providing an ideal environment to tease out model assumptions and identify persistent, underlying mechanisms. Through inter-model comparisons, this unified standard has the potential to greatly advance our understanding of the underlying mechanisms of radiation-induced DNA damage and the resulting observable biological effects when radiation parameters and/or environmental conditions change.

In Silico Non-Homologous End Joining Following Ion Induced DNA Double Strand Breaks Predicts That Repair Fidelity Depends on Break Density.

This work uses Monte Carlo simulations to investigate the dependence of residual and misrepaired double strand breaks (DSBs) at 24 hours on the initial damage pattern created during ion therapy. We present results from a nanometric DNA damage simulation coupled to a mechanistic model of Non-Homologous End Joining, capable of predicting the position, complexity, and repair of DSBs. The initial damage pattern is scored by calculating the average number of DSBs within 70 nm from every DSB. We show that this local DSB density, referred to as the cluster density, can linearly predict misrepair regardless of ion species. The models predict that the fraction of residual DSBs is constant, with 7.3% of DSBs left unrepaired following 24 hours of repair. Through simulation over a range of doses and linear energy transfer (LET) we derive simple correlations capable of predicting residual and misrepaired DSBs. These equations are applicable to ion therapy treatment planning where both dose and LET are scored. This is demonstrated by applying the correlations to an example of a clinical proton spread out Bragg peak. Here we see a considerable biological effect past the distal edge, dominated by residual DSBs.

Abstract ID: 171 A Monte Carlo study to reduce range uncertainty in proton beam therapy via prompt gamma-ray detection.

In proton beam therapy precise knowledge of the proton beam range is essential to guarantee the treatment's efficacy and to avoid unnecessary toxicities. Unlike photon beams, protons stop inside the patient's body, therefore a direct detection of the distal fall-off is impossible. One technique to determine the beam range is to detect the prompt gamma (PG) rays emitted from the nuclei de-exciting following proton bombardment [1]. PG emission is almost instantaneous and has a high-production rate. The aim of this project is to develop a new method, based on an optimized PG detector system, which can achieve 3D range determination with an uncertainty of no more than 2 mm. The presented method is based on the detection of discrete gamma-rays. As a first step, the position reconstruction capability of the PG detector system was examined by means of Geant4 simulations. The prototype system is comprised of 12 LaBr3(Ce) detectors. The information recorded by each individual detector is fed into a reconstruction algorithm to determine the gamma-ray emission point in 3 dimensions. The development of the algorithm, proof-of-principle and simulation validation, have all been conducted using a sealed 60Co source. Our simulations demonstrate that an ideal detector system with the current reconstruction algorithm is capable of determining the source position with sub-millimetre accuracy. Having obtained proof-of-principle for the reconstruction algorithm the next stage is to investigate how implementing a realistic detector system affects the reconstruction performance. In addition, the ability of the detector system to discriminate between multiple sources in different positions is under evaluation.

Nanodosimetric Simulation of Direct Ion-Induced DNA Damage Using Different Chromatin Geometry Models.

Monte Carlo based simulation has proven useful in investigating the effect of proton-induced DNA damage and the processes through which this damage occurs. Clustering of ionizations within a small volume can be related to DNA damage through the principles of nanodosimetry. For simulation, it is standard to construct a small volume of water and determine spatial clusters. More recently, realistic DNA geometries have been used, tracking energy depositions within DNA backbone volumes. Traditionally a chromatin fiber is built within the simulation and identically replicated throughout a cell nucleus, representing the cell in interphase. However, the in vivo geometry of the chromatin fiber is still unknown within the literature, with many proposed models. In this work, the Geant4-DNA toolkit was used to build three chromatin models: the solenoid, zig-zag and cross-linked geometries. All fibers were built to the same chromatin density of 4.2 nucleosomes/11 nm. The fibers were then irradiated with protons (LET 5-80 keV/µm) or alpha particles (LET 63-226 keV/µm). Nanodosimetric parameters were scored for each fiber after each LET and used as a comparator among the models. Statistically significant differences were observed in the double-strand break backbone size distributions among the models, although nonsignificant differences were noted among the nanodosimetric parameters. From the data presented in this article, we conclude that selection of the solenoid, zig-zag or cross-linked chromatin model does not significantly affect the calculated nanodosimetric parameters. This allows for a simulation-based cell model to make use of any of these chromatin models for the scoring of direct ion-induced DNA damage.

Automatic cell detection in bright-field microscopy for microbeam irradiation studies.

Automatic cell detection in bright-field illumination microscopy is challenging due to cells' inherent optical properties. Applications including individual cell microbeam irradiation demand minimisation of additional cell stressing factors, so contrast-enhancing fluorescence microscopy should be avoided. Additionally, the use of optically non-homogeneous substrates amplifies the problem. This research focuses on the design of a method for automatic cell detection on polypropylene substrate, suitable for microbeam irradiation. In order to fulfil the relative requirements, the Harris corner detector was employed to detect apparent cellular features. These features-corners were clustered based on a dual-clustering technique according to the density of their distribution across the image. Weighted centroids were extracted from the clusters of corners and constituted the targets for irradiation. The proposed method identified more than 88% of the 1,738 V79 Chinese hamster cells examined. Moreover, a processing time of 2.6 s per image fulfilled the requirements for a near real-time cell detection-irradiation system.

Investigation of gold nanoparticle radiosensitization mechanisms using a free radical scavenger and protons of different energies.

Gold nanoparticles (GNPs) have been shown to sensitize cancer cells to x-ray radiation, particularly at kV energies where photoelectric interactions dominate and the high atomic number of gold makes a large difference to x-ray absorption. Protons have a high cross-section for gold at a large range of relevant clinical energies, and so potentially could be used with GNPs for increased therapeutic effect.Here, we investigate the contribution of secondary electron emission to cancer cell radiosensitization and investigate how this parameter is affected by proton energy and a free radical scavenger. We simulate the emission from a realistic cell phantom containing GNPs after traversal by protons and x-rays with different energies. We find that with a range of proton energies (1-250 MeV) there is a small increase in secondaries compared to a much larger increase with x-rays. Secondary electrons are known to produce toxic free radicals. Using a cancer cell line in vitro we find that a free radical scavenger has no protective effect on cells containing GNPs irradiated with 3 MeV protons, while it does protect against cells irradiated with x-rays. We conclude that GNP generated free radicals are a major cause of radiosensitization and that there is likely to be much less dose enhancement effect with clinical proton beams compared to x-rays.

Influence of the nucleus area distribution on the survival fraction after charged particles broad beam irradiation.

It is well known that broad beam irradiation with heavy ions leads to variation in the number of hit(s) received by each cell as the distribution of particles follows the Poisson statistics. Although the nucleus area will determine the number of hit(s) received for a given dose, variation amongst its irradiated cell population is generally not considered. In this work, we investigate the effect of the nucleus area's distribution on the survival fraction. More specifically, this work aims to explain the deviation, or tail, which might be observed in the survival fraction at high irradiation doses. For this purpose, the nucleus area distribution was added to the beam Poisson statistics and the Linear-Quadratic model in order to fit the experimental data. As shown in this study, nucleus size variation, and the associated Poisson statistics, can lead to an upward survival trend after broad beam irradiation. The influence of the distribution parameters (mean area and standard deviation) was studied using a normal distribution, along with the Linear-Quadratic model parameters (α and ß). Finally, the model proposed here was successfully tested to the survival fraction of LN18 cells irradiated with a 85 keV µm(- 1) carbon ion broad beam for which the distribution in the area of the nucleus had been determined.

"Broadbeam" irradiation of mammalian cells using a vertical microbeam facility.

A "broadbeam" facility is demonstrated for the vertical microbeam at Surrey's Ion Beam Centre, validating the new technique used by Barazzuol et al. (Radiat Res 177:651-662, 2012). Here, droplets with a diameter of about 4 mm of 15,000 mammalian cells in suspension were pipetted onto defined locations on a 42-mm-diameter cell dish with each droplet individually irradiated in "broadbeam" mode with 2 MeV protons and 4 MeV alpha particles and assayed for clonogenicity. This method enables multiple experimental data points to be rapidly collected from the same cell dish. Initially, the Surrey vertical beamline was designed for the targeted irradiation of single cells with single counted ions. Here, the benefits of both targeted single-cell and broadbeam irradiations being available at the same facility are discussed: in particular, high-throughput cell irradiation experiments can be conducted on the same system as time-intensive focused-beam experiments with the added benefits of fluorescent microscopy, cell recognition and time-lapse capabilities. The limitations of the system based on a 2 MV tandem accelerator are also discussed, including the uncertainties associated with particle Poisson counting statistics, spread of linear energy transfer in the nucleus and a timed dose delivery. These uncertainties are calculated with Monte Carlo methods. An analysis of how this uncertainty affects relative biological effect measurements is made and discussed.

Dosimetry and spectral analysis of a radiobiological experiment using laser-driven proton beams.

Laser-driven proton and ion acceleration is an area of increasing research interest given the recent development of short pulse-high intensity lasers. Several groups have reported experiments to understand whether a laser-driven beam can be applied for radiobiological purposes and in each of these, the method to obtain dose and spectral analysis was slightly different. The difficulty with these studies is that the very large instantaneous dose rate is a challenge for commonly used dosimetry techniques, so that other more sophisticated procedures need to be explored. This paper aims to explain a method for obtaining the energetic spectrum and the dose of a laser-driven proton beam irradiating a cell dish used for radiobiology studies. The procedure includes the use of a magnet to have charge and energy separation of the laser-driven beam, Gafchromic films to have information on dose and partially on energy, and a Monte Carlo code to expand the measured data in order to obtain specific details of the proton spectrum on the cells. Two specific correction factors have to be calculated: one to take into account the variation of the dose response of the films as a function of the proton energy and the other to obtain the dose to the cell layer starting from the dose measured on the films. This method, particularly suited to irradiation delivered in a single laser shot, can be applied in any other radiobiological experiment performed with laser-driven proton beams, with the only condition that the initial proton spectrum has to be at least roughly known. The method was tested in an experiment conducted at Queen's University of Belfast using the TARANIS laser, where the mean energy of the protons crossing the cells was between 0.9 and 5 MeV, the instantaneous dose rate was estimated to be close to 109 Gy s−1 and doses between 0.8 and 5 Gy were delivered to the cells in a single laser shot. The combination of the applied corrections modified the initial estimate of dose by up to 40%.

Family group cognitive-behavioral preventive intervention for families of depressed parents: 18- and 24-month outcomes.

OBJECTIVE: In a long-term follow-up of a randomized controlled trial (Compas et al., 2009) to examine the effects at 18- and 24-month follow-ups of a family group cognitive-behavioral (FGCB) preventive intervention for mental health outcomes for children and parents from families (N = 111) of parents with a history of major depressive disorder (MDD). METHOD: Parents with a history of MDD and their 9- to 15-year-old children were randomly assigned to a FGCB intervention or a written information comparison condition. Children's internalizing, externalizing, anxiety/depression, and depressive symptoms; episodes of MDD and other psychiatric diagnoses; and parents' depressive symptoms and episodes of MDD were assessed at 18 and 24 months after randomization. RESULTS: Children in the FGCB condition were significantly lower in self-reports of anxiety/depression and internalizing symptoms at 18 months and were significantly lower in self-reports of externalizing symptoms at 18 and 24 months. Rates of MDD were significantly lower for children in the FGCB intervention over the 24-month follow-up (odds ratio = 2.91). Marginal effects were found for parents' symptoms of depression at 18 and 24 months but not for episodes of MDD. CONCLUSIONS: Support was found for a FGCB preventive intervention for children of parents with a history of MDD significantly reducing children's episodes of MDD over a period of 2 years. Significant effects for the FGCB intervention were also found on internalizing and externalizing symptoms, with stronger effects at 18- than at 24-month follow-up.

Evidence of specialized leukocyte-vascular homing interactions at the maternal/fetal interface.

In normal pregnancy, the maternal immune system fails to reject the fetus or the placenta as an allogeneic graft. We hypothesize that specialized mechanisms of leukocyte recruitment might limit access of circulating maternal immune cells to the maternal/fetal interface. During the critical period of initial trophoblast invasion there is an elegantly orchestrated progression of leukocyte homing events in the decidua basalis, associated with highly regulated expression of vascular addressins and segregation of specialized leukocyte subsets into well-defined decidual microdomains. Neutrophils are limited to the region of necrosis associated with enzymatic digestion at the leading edge of the invading trophoblast, where an almost linear array of maternal blood vessels displays the neutrophil ligand E-selectin. Cells with the phenotype of monocytes but expressing alpha4beta7 integrin are localized in the blood vessels of the specialized "vascular zone", which display the unusual combination of P-selectin (partially associated with platelets) and the alpha4beta7 ligand mucosal vascular addressin-1 (MAdCAM-1). Granulated metrial gland cells (alpha4+beta7-, probably alpha4beta1+) constitute a well-defined cluster positioned in the central decidua basalis around venules prominently expressing the alpha4beta1 ligand VCAM-1 (but not MAdCAM-1). T and B lymphocytes are rare. Our results suggest that selective mechanisms for regulating leukocyte access, associated with microdomain specialization within the decidua basalis, may play a fundamental role in immune regulation during the invasive period of placental development.