Nanodosimetric effects of gold nanoparticles in megavoltage radiation therapy

Background and purpose: The addition of gold nanoparticles (GNPs) to tumours leads to an increase in dose due to their high density and energy absorption coefficient, making it a potential radiosensitiser. However, experiments have observed radiosensitisations significantly larger than the increase in dose alone, including at megavoltage energies where gold's relative energy absorption is lowest. This work investigates whether GNPs create dose inhomogeneities on a sub-cellular scale which combine with non-linear dose dependence of cell survival to be the source of radiosensitisation at megavoltage energies.

Cell survival responses after exposure to modulated radiation fields.

In the present study survival responses were determined in cells with differing radiosensitivity, specifically primary fibroblast (AG0-1522B), human breast cancer (MDA-MB-231), human prostate cancer (DU-145) and human glioma (T98G) cells, after exposure to modulated radiation fields delivered by shielding 50% of the tissue culture flask. A significant decrease (P < 0.05) in cell survival was observed in the shielded area, outside the primary treatment field (out-of-field), that was lower than predicted when compared to uniform exposures fitted to the linear-quadratic model. Cellular radiosensitivity was demonstrated to be an important factor in the level of response for both the in- and out-of-field regions. These responses were shown to be dependent on secretion-mediated intercellular communication, because inhibition of cellular secreted factors between the in- and out-of-field regions abrogated the response. Out-of-field cell survival was shown to increase after pretreatment of cells with agents known to inhibit factors involved in mediating radiation-induced bystander signaling (aminoguanidine, DMSO or cPTIO). These data illustrate a significant decrease in survival out-of-field, dependent upon intercellular communication, in several cell lines with varying radiosensitivity after exposure to a modulated radiation field. This study provides further evidence for the importance of intercellular signaling in modulated exposures, where dose gradients are present, and may inform the refinement of established radiobiological models to facilitate the optimization of advanced radiotherapy treatment plans.

Optical source model for the 23.2-23.6 nm radiation from the multielement germanium soft X-ray laser

Distributions of source intensity in two dimensions (designated the source model), averaged over a single laser pulse, based on experimental measurements of spatial coherence, are considered for radiation from the unresolved 23.2/23.6 nm spectral lines from the germanium collisional X-ray laser. The model derives from measurements of the visibility of Young slit interference fringes determined by a method based on the Wiener-Khinchin theorem. Output from amplifiers comprising three and four target elements have similar coherence properties in directions within the horizontal plane corresponding to strong plasma refraction effects and fitting the coherence data shows source dimensions (FWHM) are similar to 26 mu m (horizontal), significantly smaller than expected by direct imaging, and similar to 125 mu m (vertical: equivalent to the height of the driver excitation). (C) 1999 Elsevier Science B.V. All rights reserved.

A computational model of cellular response to modulated radiation fields

Purpose:
To develop a model to describe the response of cell populations to spatially modulated radiation exposures of relevance to advanced radiotherapies.

Materials and Methods:
A Monte Carlo model of cellular radiation response was developed. This model incorporated damage from both direct radiation and intercellular communication including bystander signaling. The predictions of this model were compared to previously measured survival curves for a normal human fibroblast line (AGO1522) and prostate tumor cells (DU145) exposed to spatially modulated fields.

Results:
The model was found to be able to accurately reproduce cell survival both in populations which were directly exposed to radiation and those which were outside the primary treatment field. The model predicts that the bystander effect makes a significant contribution to cell killing even in uniformly irradiated cells. The bystander effect contribution varies strongly with dose, falling from a high of 80% at low doses to 25% and 50% at 4 Gy for AGO1522 and DU145 cells, respectively. This was verified using the inducible nitric oxide synthase inhibitor aminoguanidine to inhibit the bystander effect in cells exposed to different doses, which showed significantly larger reductions in cell killing at lower doses.

Conclusions:
The model presented in this work accurately reproduces cell survival following modulated radiation exposures, both in and out of the primary treatment field, by incorporating a bystander component. In addition, the model suggests that the bystander effect is responsible for a significant portion of cell killing in uniformly irradiated cells, 50% and 70% at doses of 2 Gy in AGO1522 and DU145 cells, respectively. This description is a significant departure from accepted radiobiological models and may have a significant impact on optimization of treatment planning approaches if proven to be applicable in vivo.

DNA damage induction in dry and hydrated DNA by synchrotron radiation

Radiation biophysics has sought to understand at a molecular level, the mechanisms through which ionizing radiations damage DNA, and other molecules within living cells. The complexity of lesions produced in the DNA by ionizing radiations is thought to depend on the amount of energy deposited at the site of each lesion. To study the relationship between the energy deposited and the damage produced, we have developed novel techniques for irradiating dry prasmid DNA, partially re-hydrated DNA and DNA in solution using monochromatic vacuum-UV synchrotron radiation. We have used photons in the energy range 7-150 eV, corresponding to the range of energies typically involved in the efficient production of DNA single-strand (SSB), and double-strand breaks (DSB) by ionizing radiation. The data show that both types of breaks are produced at all energies investigated (with, or without water present). Also, the energy dependence for DSB induction follows a similar trend to SSB induction but at a 20-30-fold reduced incidence, suggesting a common precursor for both types of damage. Preliminary studies where DNA has been irradiated in solution indicate a change in the shape of the dose-effect curve (from linear, to linear-quadratic for double-strand break induction) and a large increase in sensitivity due to the presence of water.

DNA double-strand break distributions in X-ray and alpha-particle irradiated V79 cells: Evidence for non-random breakage

Many studies have shown that with increasing LET of ionizing radiation the RBE (relative biological effectiveness) for dsb (double strand breaks) induction remains around 1.0 despite the increase in the RBE for cell killing. This has been attributed to an increase in the complexity of lesions, classified as dsb with current techniques, at multiply damaged sites. This study determines the molecular weight distributions of DNA from Chinese hamster V79 cells irradiated with X-rays or 110 keV/mu m alpha-particles. Two running conditions for pulsed-field gel-electrophoresis were chosen to give optimal separation of fragments either in the 225 kbp-5.7 Mbp range or the 0.3 kbp to 225 kbp range. Taking the total fraction of DNA migrating into the gel as a measure of fragmentation, the RBE for dsb induction was less than 1.0 for both molecular weight regions studied. The total yields of dsb were 8.2 x 10(-9) dsb/Gy/bp for X-rays and 7.8 x 10(-9) dsb/Gy/bp for a-particles, measured using a random breakage model. Analysis of the RBE of alpha-particles versus molecular weight gave a different response. In the 0.4 Mbp-57 Mbp region the RBE was less than 1.0; however, below 0.4 Mbp the RBE increased above 1.0. The frequency distributions of fragment sizes were found to differ from those predicted by a model assuming random breakage along the length of the DNA and the differences were greater for alpha-particles than for X-rays. An excess of fragments induced by a single-hit mechanism was found in the 8-300 kbp region and for X-rays and alpha-particles these corresponded to an extra 0.8 x 10(-9) and 3.4 x 10(-9) dsb/bp/Gy, respectively. Thus for every alpha-particle track that induces a dsb there is a 44% probability of inducing a second break within 300 kbp and for electron tracks the probability is 10%. This study shows that the distribution of damage from a high LET alpha-particle track is significantly different from that observed with low LET X-rays. In particular, it suggests that the fragmentation patterns of irradiated DNA may be related to the higher-order chromatin repealing structures found in intact cells.

A study of DNA fragmentation patterns in cells irradiated with charged particles: evidence for non-random distributions

Many studies have shown that the effectiveness of radiations of varying LET is similar when yields of dsb have been measured, despite large differences in biological response. Recent evidence has suggested however, that current techniques underestimate the yields of dsb. By monitoring the fragmentation of DNA over a wide range of fragment sizes ( 6 Mbp) by pulsed field electrophoresis, RBE values greater than 1.0 for radiations of around 100 keV/mm have been determined. The data provide evidence for the production of correlated breaks produced within cells as particle tracks traverse the nucleus. The highly ordered structure of DNA within mammalian cells may lead to clustering of breaks over distances related to the repeating unit structures of the chromatin. As well as these regionally damaged sites, a major contributor to radiation effectiveness will be the localised clustering of damage in the 1 - 20 bp region. A major effort is required to elucidate the relative importance of these levels of clustering and their importance in biological response.

A review of studies of ionizing radiation-induced double-strand break clustering

Underpinning current models of the mechanisms of the action of radiation is a central role for DNA damage and in particular double-strand breaks (DSBs). For radiations of different LET, there is a need to know the exact yields and distributions of DSBs in human cells. Most measurements of DSB yields within cells now rely on pulsed-field gel electrophoresis as the technique of choice. Previous measurements of DSB yields have suggested that the yields are remarkably similar for different types of radiation with RBE values less than or equal to1.0. More recent studies in mammalian cells, however, have suggested that both the yield and the spatial distribution of DSBs are influenced by radiation quality. RBE values for DSBs induced by high-LET radiations are greater than 1.0, and the distributions are nonrandom. Underlying this is the interaction of particle tracks with the higher-order chromosomal structures within cell nuclei. Further studies are needed to relate nonrandom distributions of DSBs to their rejoining kinetics. At the molecular level, we need to determine the involvement of clustering of damaged bases with strand breakage, and the relationship between higher-order clustering over sizes of kilobase pairs and above to localized clustering at the DNA level. Overall, these studies will allow us to elucidate whether the nonrandom distributions of breaks produced by high-LET particle tracks have any consequences for their repair and biological effectiveness. (C) 2001 by Radiation Research Society.

Low-dose hypersensitivity in Chinese hamster V79 cells targeted with counted protons using a charged-particle microbeam

The Gray Laboratory charged-particle microbeam has been used to assess the clonogenic ability of Chinese hamster V79 cells after irradiation of their nuclei with a precisely defined number of protons with energies of 1.0 and 3.2 MeV. The microbeam uses a 1-mum. silica capillary collimator to deliver protons to subcellular targets with high accuracy. The detection system is based on a miniature photomultiplier tube positioned above the cell dish, which detects the photons generated by the passage of the charged particles through an 18-mum-thick scintillator placed below the cells. With this system, a detection efficiency of greater than 99% is achieved. The cells are plated on specially designed dishes (3-mum-thick Mylar base), and the nuclei are identified by fluorescence microscopy. After an incubation period of 3 days, the cells are revisited individually to assess the formation of colonies from the surviving cells. For each energy investigated, the survival curve obtained for the microbeam shows a significant deviation below I Gy from a response extrapolated using the LQ model for the survival data above 1 Gy. The data are well fitted by a model that supports the hypothesis that radioresistance is induced by low-dose hypersensitivity. These studies demonstrate the potential of the microbeam for performing studies of the effects of single charged particles on cells in vitro. The hypersensitive responses observed are comparable with those reported by others using different radiations and techniques. (C) 2001 by Radiation Research Society.

The potential of electron beam radiation for simultaneous surface modification and bioresorption control of PLLA

Bioresorbable polymers have been widely investigated as materials exhibiting significant potential for successful application in the fields of tissue engineering and drug delivery. Further to the ability to control degradation, surface engineering of polymers has been highlighted as a key method central to their development. Previous work has demonstrated the ability of electron beam (e-beam) technology to control the degradation profiles and bioresorption of a number of commercially relevant bioresorbable polymers (poly-l-lactic acid (PLLA), Llactide/DL-lactide co-polymer (PLDL) and poly(lactic-co-glycolic acid (PLGA)). This work investigates the further potential of ebeam technology to impart added biofunctionality through the manipulation of polymer (PLLA) surface properties. PLLA samples were subjected to e-beam treatments in air, with varying beam energies and doses. Surface characterization was then performed using contact angle analysis, X-ray photoelectron spectroscopy (XPS), Raman spectroscopy, and atomic force microscopy. Results demonstrated a significant increase in surface wettability post e-beam treatment. In correlation with this, XPS data showed the introduction of oxygen-containing functional groups to the surface of PLLA. Raman spectroscopy indicated chain scission in the near surface region of PLLA (as predicted). However, e-beam effects on surface properties were not shown to be dependent on beam energy or dose. E-beam irradiation did not seem to affect the surface roughness of PLLA as a direct consequence of the treatment.

In-vitro investigation of out-of-field cell survival following the delivery of conformal, intensity-modulated radiation therapy (IMRT) and volumetric modulated arc therapy (VMAT) plans

The aim of this work is to determine the out-of-field survival of cells irradiated with either the primary field or scattered radiation in the presence and absence of intercellular communication following delivery of conformal, IMRT and VMAT treatment plans. Single beam, conformal, IMRT and VMAT plans were created to deliver 3 Gy to half the area of a T80 flask containing either DU-145 or AGO-1522 cells allowing intercellular communication between the in-and out-of-field cell populations. The same plans were delivered to a similar custom made phantom used to hold two T25 culture flasks, one flask in-field and one out-of-field to allow comparison of cell survival responses when intercellular communication is physically inhibited. Plans were created for the delivery of 8 Gy to the more radio-resistant DU-145 cells only in the presence and absence of intercellular communication. Cell survival was determined by clonogenic assay. In both cell lines, the out-of-field survival was not statistically different between delivery techniques for either cell line or dose. There was however, a statistically significant difference between survival out-of-field when intercellular communication was intact (single T80 culture flask) or inhibited (multiple T25 culture flasks) to in-field for all plans. No statistically significant difference was observed in-field with or without cellular communication to out-of-field for all plans. These data demonstrate out-of-field effects as important determinants of cell survival following exposure to modulated irradiation fields when cellular communication between differentially irradiated cell populations is present. This data is further evidence that refinement of existing radiobiological models to include indirect cell killing effects is required.

DNA damage responses following exposure to modulated radiation fields

During the delivery of advanced radiotherapy treatment techniques modulated beams are utilised to increase dose conformity across the target volume. Recent investigations have highlighted differential cellular responses to modulated radiation fields particularly in areas outside the primary treatment field that cannot be accounted for by scattered dose alone. In the present study, we determined the DNA damage response within the normal human fibroblast AG0-1522B and the prostate cancer cell line DU-145 utilising the DNA damage assay. Cells plated in slide flasks were exposed to 1 Gy uniform or modulated radiation fields. Modulated fields were delivered by shielding 25%, 50% or 75% of the flask during irradiation. The average number of 53BP1 or ?H2AX foci was measured in 2 mm intervals across the slide area. Following 30 minutes after modulated radiation field exposure an increase in the average number of foci out-of-field was observed when compared to non-irradiated controls. In-field, a non-uniform response was observed with a significant decrease in the average number of foci compared to uniformly irradiated cells. Following 24 hrs after exposure there is evidence for two populations of responding cells to bystander signals in-and out-of-field. There was no significant difference in DNA damage response between 25%, 50% or 75% modulated fields. The response was dependent on cellular secreted intercellular signalling as physical inhibition of intercellular communication abrogated the observed response. Elevated residual DNA damage observed within out-of-field regions decreased following addition of an inducible nitric oxide synthase inhibitor (Aminoguanidine). These data show, for the first time, differential DNA damage responses in-and out-of-field following modulated radiation field delivery. This study provides further evidence for a role of intercellular communication in mediating cellular radiobiological response to modulated radiation fields and may inform the refinement of existing radiobiological models for the optimization of advanced radiotherapy treatment plans. © 2012 Trainor et al.

Optimization of image quality and dose for Varian aS500 electronic portal imaging devices (EPIDs)

This study was carried out to investigate whether the electronic portal imaging (EPI) acquisition process could be optimized, and as a result tolerance and action levels be set for the PIPSPro QC-3V phantom image quality assessment. The aim of the optimization process was to reduce the dose delivered to the patient while maintaining a clinically acceptable image quality. This is of interest when images are acquired in addition to the planned patient treatment, rather than images being acquired using the treatment field during a patient's treatment. A series of phantoms were used to assess image quality for different acquisition settings relative to the baseline values obtained following acceptance testing. Eight Varian aS500 EPID systems on four matched Varian 600C/D linacs and four matched Varian 2100C/D linacs were compared for consistency of performance and images were acquired at the four main orthogonal gantry angles. Images were acquired using a 6 MV beam operating at 100 MU min(-1) and the low-dose acquisition mode. Doses used in the comparison were measured using a Farmer ionization chamber placed at d(max) in solid water. The results demonstrated that the number of reset frames did not have any influence on the image contrast, but the number of frame averages did. The expected increase in noise with corresponding decrease in contrast was also observed when reducing the number of frame averages. The optimal settings for the low-dose acquisition mode with respect to image quality and dose were found to be one reset frame and three frame averages. All patients at the Northern Ireland Cancer Centre are now imaged using one reset frame and three frame averages in the 6 MV 100 MU min(-1) low-dose acquisition mode. Routine EPID QC contrast tolerance (+/-10) and action (+/-20) levels using the PIPSPro phantom based around expected values of 190 (Varian 600C/D) and 225 (Varian 2100C/D) have been introduced. The dose at dmax from electronic portal imaging has been reduced by approximately 28%, and while the image quality has been reduced, the images produced are still clinically acceptable.