Audrey Bouchet

Toward an image-guided microbeam radiation therapy using gadolinium-based nanoparticles.

Ultrasmall gadolinium-based nanoparticles (GBNs) induce both a positive contrast for magnetic resonance imaging and a radiosentizing effect. The exploitation of these characteristics leads to a greater increase in lifespan of rats bearing brain tumors since the radiosensitizing effect of GBNs can be activated by X-ray microbeams when the gadolinium content is, at the same time, sufficiently high in the tumor and low in the surrounding healthy tissue. GBNs exhibit therefore an interesting potential for image-guided radiotherapy.

Preferential Effect of Synchrotron Microbeam Radiation Therapy on Intracerebral 9L Gliosarcoma Vascular Networks

PURPOSE Synchrotron microbeam radiation therapy (MRT) relies on spatial fractionation of the incident photon beam into parallel micron-wide beams. Our aim was to analyze the effects of MRT on normal brain and 9L gliosarcoma tissues, particularly on blood vessels. METHODS AND MATERIALS Responses to MRT (two arrays, one lateral, one anteroposterior (2 × 400 Gy), intersecting orthogonally in the tumor region) were studied during 6 weeks using MRI, immunohistochemistry, and vascular endothelial growth factor Western blot. RESULTS MRT increased the median survival time of irradiated rats (×3.25), significantly increased blood vessel permeability, and inhibited tumor growth; a cytotoxic effect on 9L cells was detected 5 days after irradiation. Significant decreases in tumoral blood volume fraction and vessel diameter were measured from 8 days after irradiation, due to loss of endothelial cells in tumors as detected by immunochemistry. Edema was observed in the normal brain exposed to both crossfired arrays about 6 weeks after irradiation. This edema was associated with changes in blood vessel morphology and an overexpression of vascular endothelial growth factor. Conversely, vascular parameters and vessel morphology in brain regions exposed to one of the two arrays were not damaged, and there was no loss of vascular endothelia. CONCLUSIONS We show for the first time that preferential damage of MRT to tumor vessels versus preservation of radioresistant normal brain vessels contributes to the efficient palliation of 9L gliosarcomas in rats. Molecular pathways of repair mechanisms in normal and tumoral vascular networks after MRT may be essential for the improvement of such differential effects on the vasculature.

High-Precision Radiosurgical Dose Delivery by Interlaced Microbeam Arrays of High-Flux Low-Energy Synchrotron X-Rays

Microbeam Radiation Therapy (MRT) is a preclinical form of radiosurgery dedicated to brain tumor treatment. It uses micrometer-wide synchrotron-generated X-ray beams on the basis of spatial beam fractionation. Due to the radioresistance of normal brain vasculature to MRT, a continuous blood supply can be maintained which would in part explain the surprising tolerance of normal tissues to very high radiation doses (hundreds of Gy). Based on this well described normal tissue sparing effect of microplanar beams, we developed a new irradiation geometry which allows the delivery of a high uniform dose deposition at a given brain target whereas surrounding normal tissues are irradiated by well tolerated parallel microbeams only. Normal rat brains were exposed to 4 focally interlaced arrays of 10 microplanar beams (52 µm wide, spaced 200 µm on-center, 50 to 350 keV in energy range), targeted from 4 different ports, with a peak entrance dose of 200Gy each, to deliver an homogenous dose to a target volume of 7 mm3 in the caudate nucleus. Magnetic resonance imaging follow-up of rats showed a highly localized increase in blood vessel permeability, starting 1 week after irradiation. Contrast agent diffusion was confined to the target volume and was still observed 1 month after irradiation, along with histopathological changes, including damaged blood vessels. No changes in vessel permeability were detected in the normal brain tissue surrounding the target. The interlacing radiation-induced reduction of spontaneous seizures of epileptic rats illustrated the potential pre-clinical applications of this new irradiation geometry. Finally, Monte Carlo simulations performed on a human-sized head phantom suggested that synchrotron photons can be used for human radiosurgical applications. Our data show that interlaced microbeam irradiation allows a high homogeneous dose deposition in a brain target and leads to a confined tissue necrosis while sparing surrounding tissues. The use of synchrotron-generated X-rays enables delivery of high doses for destruction of small focal regions in human brains, with sharper dose fall-offs than those described in any other conventional radiation therapy.

Synchrotron microbeam radiation therapy for rat brain tumor palliation—influence of the microbeam width at constant valley dose

To analyze the effects of the microbeam width (25, 50 and 75 microm) on the survival of 9L gliosarcoma tumor-bearing rats and on toxicity in normal tissues in normal rats after microbeam radiation therapy (MRT), 9L gliosarcomas implanted in rat brains, as well as in normal rat brains, were irradiated in the MRT mode. Three configurations (MRT25, MRT50, MRT75), each using two orthogonally intersecting arrays of either 25, 50 or 75 microm wide microbeams, all spaced 211 microm on center, were tested. For each configuration, peak entrance doses of 860, 480 and 320 Gy, respectively, were calculated to produce an identical valley dose of 18 Gy per individual array at the center of the tumor. Two, 7 and 14 days after radiation treatment, 42 rats were killed to evaluate histopathologically the extent of tumor necrosis, and the presence of proliferating tumors cells and tumor vessels. The median survival times of the normal rats were 4.5, 68 and 48 days for MRT25, 50 and 75, respectively. The combination of the highest entrance doses (860 Gy per array) with 25 microm wide beams (MRT25) resulted in a cumulative valley dose of 36 Gy and was excessively toxic, as it led to early death of all normal rats and of approximately 50% of tumor-bearing rats. The short survival times, particularly of rats in the MRT25 group, restricted adequate observance of the therapeutic effect of the method on tumor-bearing rats. However, microbeams of 50 microm width led to the best median survival time after 9L gliosarcoma MRT treatment and appeared as the better compromise between tumor control and normal brain toxicity compared with 75 microm or 25 microm widths when used with a 211 microm on-center distance. Despite very high radiation doses, the tumors were not sterilized; viable proliferating tumor cells remained present at the tumor margin. This study shows that microbeam width and peak entrance doses strongly influence tumor responses and normal brain toxicity, even if valley doses are kept constant in all groups. The use of 50 microm wide microbeams combined with moderate peak doses resulted in a higher therapeutic ratio.

Brain tumor vessel response to synchrotron microbeam radiation therapy: a short-term in vivo study

The aim of this work focuses on the description of the short-term response of a 9L brain tumor model and its vasculature to microbeam radiation therapy (MRT) using magnetic resonance imaging (MRI). Rat 9L gliosarcomas implanted in nude mice brains were irradiated by MRT 13 days after tumor inoculation using two orthogonal arrays of equally spaced 28 planar microbeams (25 microm width, 211 microm spacing and dose 500 Gy). At 1, 7 and 14 days after MRT, apparent diffusion coefficient, blood volume and vessel size index were mapped by MRI. Mean survival time after tumor inoculation increased significantly between MRT-treated and untreated groups (23 and 28 days respectively, log-rank test, p < 0.0001). A significant increase of apparent diffusion coefficient was observed 24 h after MRT in irradiated tumors versus non-irradiated ones. In the untreated group, both tumor size and vessel size index increased significantly (from 7.6 +/- 2.2 to 19.2 +/- 4.0 mm(2) and +23%, respectively) between the 14th and the 21st day after tumor cell inoculation. During the same period, in the MRT-treated group, no difference in tumor size was observed. The vessel size index measured in the MRT-treated group increased significantly (+26%) between 14 and 28 days of tumor growth. We did not observe the significant difference in blood volume between the MRT-treated and untreated groups. MRT slows 9L tumor growth in a mouse brain but MRI results suggest that the increase in survival time after our MRT approach may be rather due to a cytoreduction than to early direct effects of ionizing radiation on tumor vessels. These results suggest that MRT parameters need to be optimized to further damage tumor vessels.

Synchrotron microbeam radiation therapy induces hypoxia in intracerebral gliosarcoma but not in the normal brain

PURPOSE Synchrotron microbeam radiation therapy (MRT) is an innovative irradiation modality based on spatial fractionation of a high-dose X-ray beam into lattices of microbeams. The increase in lifespan of brain tumor-bearing rats is associated with vascular damage but the physiological consequences of MRT on blood vessels have not been described. In this manuscript, we evaluate the oxygenation changes induced by MRT in an intracerebral 9L gliosarcoma model. METHODS Tissue responses to MRT (two orthogonal arrays (2 × 400Gy)) were studied using magnetic resonance-based measurements of local blood oxygen saturation (MR_SO2) and quantitative immunohistology of RECA-1, Type-IV collagen and GLUT-1, marker of hypoxia. RESULTS In tumors, MR_SO2 decreased by a factor of 2 in tumor between day 8 and day 45 after MRT. This correlated with tumor vascular remodeling, i.e. decrease in vessel density, increases in half-vessel distances (×5) and GLUT-1 immunoreactivity. Conversely, MRT did not change normal brain MR_SO2, although vessel inter-distances increased slightly. CONCLUSION We provide new evidence for the differential effect of MRT on tumor vasculature, an effect that leads to tumor hypoxia. As hypothesized formerly, the vasculature of the normal brain exposed to MRT remains sufficiently perfused to prevent any hypoxia.

Monitoring Blood-Brain Barrier Status in a Rat Model of Glioma Receiving Therapy: Dual Injection of Low-Molecular-Weight and Macromolecular MR Contrast Media

PURPOSE To evaluate the sequential injection of a low-molecular-weight (gadoterate meglumine [Gd-DOTA], 0.5 kDa) and a macromolecular (P846, 3.5 kDa) contrast media in monitoring the effect of antitumor therapies (antiangiogenic therapy and/or microbeam radiation therapy [MRT]) on healthy brain tissue and implanted tumors. MATERIALS AND METHODS Animal use was compliant with official French guidelines and was assessed by the local Internal Evaluation Committee for Animal Welfare and Rights. Eighty male rats bearing 9L gliosarcoma were randomized into four groups: untreated, antiangiogenic (sorafenib) therapy, MRT, and both treatments. Magnetic resonance (MR) imaging was performed 1 day before and 1, 5, and 8 days after the start of the treatment. At all time points, vascular integrity to a macromolecular contrast medium (P846) and, 11 minutes 30 seconds later, to low-molecular-weight contrast medium (Gd-DOTA) was evaluated by using a dynamic contrast material-enhanced MR imaging approach. To quantify vessel wall integrity, areas under the signal intensity curves were computed for each contrast medium. Unpaired t tests and one-way analysis of variance were used for statistical analyses. RESULTS Tumor vessels receiving antiangiogenic therapy became less permeable to the macromolecular contrast medium, but their permeability to the low-molecular-weight contrast medium remained unchanged. Healthy double-irradiated vessels became permeable to the low-molecular-weight contrast medium but not to the macromolecular contrast medium. CONCLUSION Antiangiogenic therapy and MRT generate different effects on the extravasation of contrast medium in tumoral and healthy tissues. This study indicates that the use of a low-molecular-weight contrast medium and a macromolecular contrast medium provides complementary information and suggests that the use of two contrast media within the same MR imaging session is feasible.

First trial of spatial and temporal fractionations of the delivered dose using synchrotron microbeam radiation therapy

The technical feasibility of temporal and spatial fractionations of the radiation dose has been evaluated using synchrotron microbeam radiation therapy for brain tumors in rats. A significant increase in lifespan (216%, p < 0.0001) resulted when three fractions of microbeam irradiation were applied to the tumor through three different ports, orthogonal to each other, at 24 h intervals. However, there were no long-term survivors, and immunohistological studies revealed that 9 L tumors were not entirely ablated.

Molecular X-ray computed tomography of myelin in a rat brain

In this work we demonstrate the feasibility of applying small-angle X-ray scattering computed tomography (SAXS-CT) for non-invasive molecular imaging of myelin sheaths in a rat brain. Our results show that the approach yields information on several quantities, including the relative myelin concentration, its periodicity, the total thickness of the myelin sheaths, and the relative concentration of cytoskeletal neurofilaments. For example the periodicity of the myelin sheaths varied in the range from 17.0 to 18.2 nm around an average of 17.6 (±0.3) nm. We believe that imaging, i.e., spatially resolved measuring these quantities could provide general means for understanding the relation to a number of neurodegenerative diseases.

Increase of lifespan for glioma-bearing rats by using minibeam radiation therapy

This feasibility work assesses the therapeutic effectiveness of minibeam radiation therapy, a new synchrotron radiotherapy technique. In this new approach the irradiation is performed on 9L gliosarcoma-bearing rats with arrays of parallel beams of width 500-700 µm. Two irradiation configurations were compared: a lateral unidirectional irradiation and two orthogonal arrays interlacing at the target. A dose escalation study was performed. A factor of three gain in the mean survival time obtained for some animals paves the way for further exploration of the different possibilities of this technique and its further optimization.