Neurologic Changes Induced by Whole-Brain Synchrotron Microbeam Irradiation: 10-Month Behavioral and Veterinary Follow-Up.

PURPOSE: Novel radiation therapy approaches have increased the therapeutic efficacy for malignant brain tumors over the past decades, but the balance between therapeutic gain and radiotoxicity remains a medical hardship. Synchrotron microbeam radiation therapy, an innovative technique, deposes extremely high (peak) doses in micron-wide, parallel microbeam paths, whereas the diffusing interbeam (valley) doses lie in the range of conventional radiation therapy doses. In this study, we evaluated normal tissue toxicity of whole-brain microbeam irradiation (MBI) versus that of a conventional hospital broad beam (hBB). METHODS AND MATERIALS: Normal Fischer rats (n = 6-7/group) were irradiated with one of the two modalities, exposing the entire brain to MBI valley/peak doses of 0/0, 5/200, 10/400, 13/520, 17/680, or 25/1000 Gy or to hBB doses of 7, 10, 13, 17, or 25 Gy. Two additional groups of rats received an MBI valley dose of 10 Gy coupled with an hBB dose of 7 or 15 Gy (groups MBI17* and MBI25*). Behavioral parameters were evaluated for 10 months after irradiation combined with veterinary observations. RESULTS: MBI peak doses of ≥680 Gy caused acute toxicity and death. Animals exposed to hBB or MBI dose-dependently gained less weight than controls; rats in the hBB25 and MBI25* groups died within 6 months after irradiation. Increasing doses of MBI caused hyperactivity but no other detectable behavioral alterations in our tests. Importantly, no health concerns were seen up to an MBI valley dose of 17 Gy. CONCLUSIONS: While acute toxicity of microbeam exposures depends on very high peak doses, late toxicity mainly relates to delivery of high MBI valley doses. MBI seems to have a low impact on normal rat behavior, but further tests are warranted to fully explore this hypothesis. However, high peak and valley doses are well tolerated from a veterinary point of view. This normal tissue tolerance to whole-brain, high-dose MBI reveals a promising avenue for microbeam radiation therapy, that is, therapeutic applications of microbeams that are poised for translation to a clinical environment.

Monocrystalline diamond detector for online monitoring during synchrotron microbeam radiotherapy.

Microbeam radiation therapy (MRT) is a radiotherapy technique combining spatial fractionation of the dose distribution on a micrometric scale, X-rays in the 50-500 keV range and dose rates up to 16 × 103 Gy s-1. Nowadays, in vivo dosimetry remains a challenge due to the ultra-high radiation fluxes involved and the need for high-spatial-resolution detectors. The aim here was to develop a striped diamond portal detector enabling online microbeam monitoring during synchrotron MRT treatments. The detector, a 550 µm bulk monocrystalline diamond, is an eight-strip device, of height 3 mm, width 178 µm and with 60 µm spaced strips, surrounded by a guard ring. An eight-channel ASIC circuit for charge integration and digitization has been designed and tested. Characterization tests were performed at the ID17 biomedical beamline of the European Synchrotron Radiation Facility (ESRF). The detector measured direct and attenuated microbeams as well as interbeam fluxes with a precision level of 1%. Tests on phantoms (RW3 and anthropomorphic head phantoms) were performed and compared with simulations. Synchrotron radiation measurements were performed on an RW3 phantom for strips facing a microbeam and for strips facing an interbeam area. A 2% difference between experiments and simulations was found. In more complex geometries, a preliminary study showed that the absolute differences between simulated and recorded transmitted beams were within 2%. Obtained results showed the feasibility of performing MRT portal monitoring using a microstriped diamond detector. Online dosimetric measurements are currently ongoing during clinical veterinary trials at ESRF, and the next 153-strip detector prototype, covering the entire irradiation field, is being finalized at our institution.

MRT-boost as the last fraction may be the most efficient irradiation schedule for increased survival times in a rat glioma model.

Synchrotron microbeam radiation therapy (MRT) is based on the spatial fractionation of the incident synchrotron beam into arrays of parallel microbeams, typically a few tens of micrometres wide and depositing several hundred Gray. This high dose, high dose rate, spatially fractionated radiotherapy has a high therapeutic impact on tumors, especially in intracranial locations. MRT leads to better control of incurable high-grade glioma than from homogeneous radiotherapy. The schedule of MRT within a conventional irradiation protocol (three fractions of 11 Gy) of brain tumors was evaluated on the 9L glioma model in rats. MRT delivered as a first fraction increased the median survival time of the animals by four days compared with conventional radiotherapy, while the last MRT fraction improved the lifespan by 148% (+15.5 days compared with conventional radiotherapy, p < 0.0001). The most efficient radiation regimen was obtained when the MRT-boost was applied as the last fraction, following two conventional clinical exposures.

Toward Neuro-Oncologic Clinical Trials of High-Dose-Rate Synchrotron Microbeam Radiation Therapy: First Treatment of a Spontaneous Canine Brain Tumor.

PURPOSE: The high potential of microbeam radiation therapy (MRT) in improving tumor control while reducing side effects has been shown by numerous preclinical studies. MRT offers a widened therapeutic window by using the periodical spatial fractionation of synchrotron generated x-rays into an array of intense parallel microbeams. MRT now enters a clinical transfer phase. As proof of principle and cornerstone for the safe clinical transfer of MRT, we conducted a "first in dog" trial under clinical conditions. In this report, we evaluated whether a 3-dimensional conformal MRT can be safely delivered as exclusive radiosurgical treatment in animal patients METHODS AND MATERIALS: We irradiated a 17.5-kg French bulldog for a spontaneous brain tumor (glioma suspected on magnetic resonance imaging) with conformal high-dose-rate microbeam arrays (50-µm-wide microbeams, replicated with a pitch of 400 µm) of synchrotron-generated x-rays. The dose prescription adjusted a minimal cumulated valley dose of 2.8 Gy to the plnning target volume (PTV) (cinical target volume (CTV)+ 1 mm). Thus, each beam delivered 20 to 25 Gy to the target as peak doses, and ∼1 Gy as valley doses RESULTS: The treatment was successfully delivered. Clinical follow-up over 3 months showed a significant improvement of the dog's quality of life: the symptoms disappeared. Magnetic resonance imaging, performed 3 months after irradiation, revealed reduction in tumor size (-87.4%) and mass effect with normalization of the left lateral ventricle. CONCLUSIONS: To our knowledge, this neuro-oncologic veterinary trial is the first 3-dimensional conformal synchrotron x-ray MRT treatment of a spontaneous intracranial tumor in a large animal. It is an essential last step toward the clinical transfer of MRT in the near future to demonstrate the feasibility and safety of treating deep-seated tumors using synchrotron-generated microbeams.

A high-resolution dose calculation engine for X-ray microbeams radiation therapy.

BACKGROUND: Microbeam radiation therapy (MRT) is a treatment modality based on spatial fractionation of synchrotron generated X-rays into parallel, high dose, microbeams of a few microns width. MRT is still an underdevelopment radiosurgery technique for which, promising preclinical results on brain tumors and epilepsy encourages its clinical transfer. PURPOSE: A safe clinical transfer of MRT needs a specific treatment planning system (TPS) that provides accurate dose calculations in human patients, taking into account the MRT beam's properties (high-dose gradients, spatial fractionation, polarization effects). So far, the most advanced MRT TPS, based on a hybrid dose calculation algorithm, is limited to a macroscopic rendering of the dose and does not account for the complex dose distribution inherent to MRT if delivered as conformal irradiations with multiple incidences. For overcoming these limitations, a multi-scale full Monte-Carlo calculation engine called penMRT has been developed and benchmarked against two general-purpose Monte Carlo (MC) codes: penmain based on PENELOPE and Gate based on Geant4. METHODS: PenMRT, is based on the PENELOPE (2018) MC code, modified to take into account the voxelized geometry of the patients (computed tomography [CT]-scans) and is offering an adaptive micrometric dose calculation grid independent of the CT size, location, and orientation. The implementation of the dynamic memory allocation in penMRT, makes the simulations feasible within a huge number of dose scoring bins. The possibility of using a source replication approach to simulate arrays of microbeams, and the parallelization using OpenMPI have been added to penMRT in order to increase the calculation speed for clinical usages. This engine can be implemented in a TPS as a dose calculation core. RESULTS: The performance tests highlight the reliability of penMRT to be used for complex irradiation conditions in MRT. The benchmarking against a standard PENELOPE code did not show any significant difference for calculations in centimetric beams, for a single microbeam and for a microbeam array. The comparisons between penMRT and Gate as an independent MC code did not show any difference in the beam paths, whereas, in valley regions, relative differences between the two codes rank from 1% to 7.5% which are probably due to the differences in physics lists that are used in these two codes. The reliability of the source replication approach has also been tested and validated with an underestimation of no more than 0.6% in low-dose areas. CONCLUSIONS: Good agreements (a relative difference between 0% and 8%) were found when comparing calculated peak to valley dose ratio values using penMRT, for irradiations with a full microbeam array, with calculated values in the literature. The high-resolution calculated dose maps obtained with penMRT are used to extract differential and cumulative dose-volume histograms (DVHs) and analyze treatment plans with much finer metrics regarding the irradiation complexity. To our knowledge, these are the first high-resolution dose maps and associated DVHs ever obtained for cross-fired microbeams irradiation, which is bringing a significant added value to the field of treatment planning in spatially fractionated radiation therapy.

Aberrant neuronal connectivity in the cortex drives generation of seizures in rat absence epilepsy.

Absence epilepsy belongs to genetic epilepsies and is characterized by recurrent generalized seizures that are concomitant with alterations of consciousness and associated with cognitive comorbidities. Little is known about the mechanisms leading to occurrence of epileptic seizures (i.e. epileptogenesis) and, in particular, it remains an open question as to whether neuronal hypersynchronization, a key feature in seizure initiation, could result from aberrant structural connectivity within neuronal networks endowing them with epileptic properties. In the present study, we addressed this question using a genetic model of absence epilepsy in the rat where seizures initiate in the whisker primary somatosensory cortex (wS1). We hypothesized that alterations in structural connectivity of neuronal networks within wS1 contribute to pathological neuronal synchronization responsible for seizures. First, we used rabies virus-mediated retrograde synaptic tracing and showed that cortical neurons located in both upper- and deep-layers of wS1 displayed aberrant and significantly increased connectivity in the genetic model of absence epilepsy, as highlighted by a higher number of presynaptic partners. Next, we showed at the functional level that disrupting these aberrant wS1 neuronal networks with synchrotron X-ray-mediated cortical microtransections drastically decreased both the synchronization and seizure power of wS1 neurons, as revealed by in vivo local field potential recordings with multichannel probes. Taken together, our data provide for the first time strong evidence that increased structural connectivity patterns of cortical neurons represent critical pathological substrates for increased neuronal synchronization and generation of absence seizures.

Tolerance of Normal Rabbit Facial Bones and Teeth to Synchrotron X-Ray Microbeam Irradiation.

Microbeam radiation therapy, an alternative radiosurgical treatment under preclinical investigation, aims to safely treat muzzle tumors in pet animals. This will require data on the largely unknown radiation toxicity of microbeam arrays for bones and teeth. To this end, the muzzle of six young adult New Zealand rabbits was irradiated by a lateral array of microplanar beamlets with peak entrance doses of 200, 330 or 500 Gy. The muzzles were examined 431 days postirradiation by computed microtomographic imaging (micro-CT) ex vivo, and extensive histopathology. The boundaries of the radiation field were identified histologically by microbeam tracks in cartilage and other tissues. There was no radionecrosis of facial bones in any rabbit. Conversely, normal incisor teeth exposed to peak entrance doses of 330 Gy or 500 Gy developed marked caries-like damage, whereas the incisors of the two rabbits exposed to 200 Gy remained unscathed. A single, unidirectional array of microbeams with a peak entrance dose ≤200 Gy (valley dose14 Gy) did not damage normal bone, teeth and soft tissues of the muzzle of normal rabbits longer than one year after irradiation. Because of that, Microbeam radiation therapy of muzzle tumors in pet animals is unlikely to cause sizeable damage to normal teeth, bone and soft tissues, if a single array as used here delivers a limited entrance dose of 200 Gy and a valley dose of ≤14 Gy.

Unexpected Benefits of Multiport Synchrotron Microbeam Radiation Therapy for Brain Tumors.

Delivery of high-radiation doses to brain tumors via multiple arrays of synchrotron X-ray microbeams permits huge therapeutic advantages. Brain tumor (9LGS)-bearing and normal rats were irradiated using a conventional, homogeneous Broad Beam (BB), or Microbeam Radiation Therapy (MRT), then studied by behavioral tests, MRI, and histopathology. A valley dose of 10 Gy deposited between microbeams, delivered by a single port, improved tumor control and median survival time of tumor-bearing rats better than a BB isodose. An increased number of ports and an accumulated valley dose maintained at 10 Gy delayed tumor growth and improved survival. Histopathologically, cell death, vascular damage, and inflammatory response increased in tumors. At identical valley isodose, each additional MRT port extended survival, resulting in an exponential correlation between port numbers and animal lifespan (r2 = 0.9928). A 10 Gy valley dose, in MRT mode, delivered through 5 ports, achieved the same survival as a 25 Gy BB irradiation because of tumor dose hot spots created by intersecting microbeams. Conversely, normal tissue damage remained minimal in all the single converging extratumoral arrays. Multiport MRT reached exceptional ~2.5-fold biological equivalent tumor doses. The unique normal tissue sparing and therapeutic index are eminent prerequisites for clinical translation.

Synchrotron X-Ray Boost Delivered by Microbeam Radiation Therapy After Conventional X-Ray Therapy Fractionated in Time Improves F98 Glioma Control.

PURPOSE: Synchrotron microbeam radiation therapy (MRT) is based on the spatial fractionation of the incident, highly collimated synchrotron beam into arrays of parallel microbeams depositing several hundred grays. It appears relevant to combine MRT with a conventional treatment course, preparing a treatment scheme for future patients in clinical trials. The efficiency of MRT delivered after several broad-beam (BB) fractions to palliate F98 brain tumors in rats in comparison with BB fractions alone was evaluated in this study. METHODS AND MATERIALS: Rats bearing 106 F98 cells implanted in the caudate nucleus were irradiated by 5 fractions in BB mode (3 × 6 Gy + 2 × 8 Gy BB) or by 2 boost fractions in MRT mode to a total of 5 fractions (3 × 6 Gy BB + MRT 2 × 8 Gy valley dose; peak dose 181 Gy [50/200 µm]). Tumor growth was evaluated in vivo by magnetic resonance imaging follow-up at T-1, T7, T12, T15, T20, and T25 days after radiation therapy and by histology and flow cytometry. RESULTS: MRT-boosted tumors displayed lower cell density and cell proliferation compared with BB-irradiated tumors. The MRT boost completely stopped tumor growth during ∼4 weeks and led to a significant increase in median survival time, whereas tumors treated with BB alone recurred within a few days after the last radiation fraction. CONCLUSIONS: The first evidence is presented that MRT, delivered as a boost of conventionally fractionated irradiation by orthovoltage broad x-ray beams, is feasible and more efficient than conventional radiation therapy alone.

Locomotion and eating behavior changes in Yucatan minipigs after unilateral radio-induced ablation of the caudate nucleus.

The functional roles of the Caudate nucleus (Cd) are well known. Selective Cd lesions can be found in neurological disorders. However, little is known about the dynamics of the behavioral changes during progressive Cd ablation. Current stereotactic radiosurgery technologies allow the progressive ablation of a brain region with limited adverse effects in surrounding normal tissues. This could be of high interest for the study of the modified behavioral functions in relation with the degree of impairment of the brain structures. Using hypofractionated stereotactic radiotherapy combined with synchrotron microbeam radiation, we investigated, during one year after irradiation, the effects of unilateral radio-ablation of the right Cd on the behavior of Yucatan minipigs. The right Cd was irradiated to a minimal dose of 35.5 Gy delivered in three fractions. MRI-based morphological brain integrity and behavioral functions, i.e. locomotion, motivation/hedonism were assessed. We detected a progressive radio-necrosis leading to a quasi-total ablation one year after irradiation, with an additional alteration of surrounding areas. Transitory changes in the motivation/hedonism were firstly detected, then on locomotion, suggesting the influence of different compensatory mechanisms depending on the functions related to Cd and possibly some surrounding areas. We concluded that early behavioral changes related to eating functions are relevant markers for the early detection of ongoing lesions occurring in Cd-related neurological disorders.

Film dosimetry studies for patient specific quality assurance in microbeam radiation therapy.

Microbeam radiation therapy (MRT) uses synchrotron arrays of X-ray microbeams to take advantage of the spatial fractionation effect for normal tissue sparing. In this study, radiochromic film dosimetry was performed for a treatment where MRT is introduced as a dose boost in a hypofractionated stereotactic radiotherapy (SRT) scheme. The isocenter dose was measured using an ionization chamber and two dimensional dose distributions were determined using radiochromic films. To compare the measured dose distribution to the MRT treatment plan, peak and valley were displayed in separate dosemaps. The measured and computed isocenter doses were compared and a two-dimensional 2%/2 mm normalized γ-index analysis with a 90% passing rate criterion was computed. For SRT, a difference of 2.6% was observed in the dose at the isocenter from the treatment plan and film measurement, with a passing rate of 96% for the γ-index analysis. For MRT, peak and valley doses differences of 25.6% and 8.2% were observed, respectively but passing rates of 96% and 90% respectively were obtained from the normalized γ-index maps. The differences in isocenter doses measured in MRT should be further investigated. We present the methodology of patient specific quality assurance (QA) for studying MRT dose distributions and discuss ideas to improve absolute dosimetry. This patient specific QA will be used for large animal trials quality assurance where MRT will be administered as a dose boost in conventional SRT. The observed remaining discrepancies should be studied against approximations in the TPS phantom materials, beams characteristics or film read-out procedures.

Ultra high dose rate Synchrotron Microbeam Radiation Therapy. Preclinical evidence in view of a clinical transfer.

This paper reviews the current state of the art of an emerging form of radiosurgery dedicated to brain tumour treatment and which operates at very high dose rate (kGy·s-1). Microbeam Radiation Therapy uses synchrotron-generated X-rays which triggered normal tissue sparing partially mediated by FLASH effect.

X-rays can trigger the FLASH effect: Ultra-high dose-rate synchrotron light source prevents normal brain injury after whole brain irradiation in mice.

This study is the first proof of concept that the FLASH effect can be triggered by X-rays. Our results show that a 10 Gy whole-brain irradiation delivered at ultra-high dose-rate with synchrotron generated X-rays does not induce memory deficit; it reduces hippocampal cell-division impairment and induces less reactive astrogliosis.

Cluster versus ROI analysis to assess combined antiangiogenic therapy and radiotherapy in the F98 rat-glioma model.

For glioblastoma (GBM), current therapeutic approaches focus on the combination of several therapies, each of them individually approved for GBM or other tumor types. Many efforts are made to decipher the best sequence of treatments that would ultimately promote the most efficient tumor response. There is therefore a strong interest in developing new clinical in vivo imaging procedures that can rapidly detect treatment efficacy and allow individual modulation of the treatment. In this preclinical study, we propose to evaluate tumor tissue changes under combined therapies, tumor vascular normalization under antiangiogenic treatment followed by radiotherapy, using a voxel-based clustering approach. This approach was applied to a rat model of glioma (F98). Six MRI parameters were mapped: apparent diffusion coefficient, vessel wall permeability, cerebral blood volume fraction, cerebral blood flow, tissue oxygen saturation and vessel size index. We compared the classical region of interest (ROI)-based analysis with a cluster-based analysis. Five clusters, defined by their MRI features, were sufficient to characterize tumor progression and tumor changes during treatments. These results suggest that the cluster-based analysis was as efficient as the ROI-based analysis to assess tumor physiological changes during treatment, but also gave additional information regarding the voxels impacted by treatments and their localization within the tumor. Overall, cluster-based analysis appears to be a powerful tool for subtle monitoring of tumor changes during combined therapies.

Permeability of Brain Tumor Vessels Induced by Uniform or Spatially Microfractionated Synchrotron Radiation Therapies.

PURPOSE: To compare the blood-brain barrier permeability changes induced by synchrotron microbeam radiation therapy (MRT, which relies on spatial fractionation of the incident x-ray beam into parallel micron-wide beams) with changes induced by a spatially uniform synchrotron x-ray radiation therapy. METHODS AND MATERIALS: Male rats bearing malignant intracranial F98 gliomas were randomized into 3 groups: untreated, exposed to MRT (peak and valley dose: 241 and 10.5 Gy, respectively), or exposed to broad beam irradiation (BB) delivered at comparable doses (ie, equivalent to MRT valley dose); both applied by 2 arrays, intersecting orthogonally the tumor region. Vessel permeability was monitored in vivo by magnetic resonance imaging 1 day before (T-1) and 1, 2, 7, and 14 days after treatment start. To determine whether physiologic parameters influence vascular permeability, we evaluated vessel integrity in the tumor area with different values for cerebral blood flow, blood volume, edema, and tissue oxygenation. RESULTS: Microbeam radiation therapy does not modify the vascular permeability of normal brain tissue. Microbeam radiation therapy-induced increase of tumor vascular permeability was detectable from T2 with a maximum at T7 after exposure, whereas BB enhanced vessel permeability only at T7. At this stage MRT was more efficient at increasing tumor vessel permeability (BB vs untreated: +19.1%; P=.0467; MRT vs untreated: +44.8%; P<.0001), and its effects lasted until T14 (MRT vs BB, +22.6%; P=.0199). We also showed that MRT was more efficient at targeting highly oxygenated (high blood volume and flow) and more proliferative parts of the tumor than BB. CONCLUSIONS: Microbeam radiation therapy-induced increased tumor vascular permeability is: (1) significantly greater; (2) earlier and more prolonged than that induced by BB irradiation, especially in highly proliferative tumor areas; and (3) targets all tumor areas discriminated by physiologic characteristics, including those not damaged by homogeneous irradiation.

Synchrotron microbeam irradiation induces neutrophil infiltration, thrombocyte attachment and selective vascular damage in vivo.

Our goal was the visualizing the vascular damage and acute inflammatory response to micro- and minibeam irradiation in vivo. Microbeam (MRT) and minibeam radiation therapies (MBRT) are tumor treatment approaches of potential clinical relevance, both consisting of parallel X-ray beams and allowing the delivery of thousands of Grays within tumors. We compared the effects of microbeams (25-100 µm wide) and minibeams (200-800 µm wide) on vasculature, inflammation and surrounding tissue changes during zebrafish caudal fin regeneration in vivo. Microbeam irradiation triggered an acute inflammatory response restricted to the regenerating tissue. Six hours post irradiation (6 hpi), it was infiltrated by neutrophils and fli1a(+) thrombocytes adhered to the cell wall locally in the beam path. The mature tissue was not affected by microbeam irradiation. In contrast, minibeam irradiation efficiently damaged the immature tissue at 6 hpi and damaged both the mature and immature tissue at 48 hpi. We demonstrate that vascular damage, inflammatory processes and cellular toxicity depend on the beam width and the stage of tissue maturation. Minibeam irradiation did not differentiate between mature and immature tissue. In contrast, all irradiation-induced effects of the microbeams were restricted to the rapidly growing immature tissue, indicating that microbeam irradiation could be a promising tumor treatment tool.

In vivo pink-beam imaging and fast alignment procedure for rat brain tumor radiation therapy.

A fast positioning method for brain tumor microbeam irradiations for preclinical studies at third-generation X-ray sources is described. The three-dimensional alignment of the animals relative to the X-ray beam was based on the X-ray tomography multi-slices after iodine infusion. This method used pink-beam imaging produced by the ID17 wiggler. A graphical user interface has been developed in order to define the irradiation parameters: field width, height, number of angles and X-ray dose. This study is the first reporting an image guided method for soft tissue synchrotron radiotherapy. It allowed microbeam radiation therapy irradiation fields to be reduced by a factor of ∼20 compared with previous studies. It permitted more targeted, more efficient brain tumor microbeam treatments and reduces normal brain toxicity of the radiation treatment.

Multiparametric MRI as an early biomarker of individual therapy effects during concomitant treatment of brain tumours.

The aim of this study was to determine the ability of multiparametric MRI to identify the early effects of individual treatment, during combined chemo-radiotherapy on brain tumours. Eighty male rats bearing 9L gliosarcomas were randomized into four groups: untreated, anti-angiogenic therapy (SORA group), microbeam radiation therapy (MRT group) and both treatments (MRT+SORA group). Multiparametric MRI (tumour volume, diffusion-weighted MR imaging (ADC), blood volume fraction (BVf), microvessel index (VSI), vessel wall integrity (AUC(P846)) and tissue oxygen saturation (StO2)) was performed 1 day before and 2, 5 and 8 days after treatment initiation. Unpaired t-tests and one-way ANOVA were used for statistical analyses. Each MR parameter measured in our protocol was revealed to be sensitive to tumour changes induced by any of the therapies used (individually or combined). When compared with untreated tumours, SORA induced a decrease in BVf, VSI, StO2 and AUC(P846), MRT generated an increase in ADC and AUC(P846) and combined therapies yielded mixed effects: an increase in ADC and AUC(P846) and a decrease in BVf, StO2 and AUC(P846). MRT and MRT+SORA significantly slowed tumour growth. Despite these two groups presenting with similar tumour sizes, the information yielded from MR multiparameter assessment indicated that, when used concomitantly, each therapy induced distinguishable and appreciable physiological changes in the tumour. Our results suggest that multiparametric MRI can monitor the effects of individual treatments, used concomitantly, on brain tumours. Such monitoring would be useful for the detection of tumour resistance to drug/radiotherapy in patients undergoing concomitant therapies.

Effects of microbeam radiation therapy on normal and tumoral blood vessels.

Microbeam radiation therapy (MRT) is a new form of preclinical radiotherapy using quasi-parallel arrays of synchrotron X-ray microbeams. While the deposition of several hundred Grays in the microbeam paths, the normal brain tissues presents a high tolerance which is accompanied by the permanence of apparently normal vessels. Conversely, the efficiency of MRT on tumor growth control is thought to be related to a preferential damaging of tumor blood vessels. The high resistance of the healthy vascular network was demonstrated in different animal models by in vivo biphoton microscopy, magnetic resonance imaging, and histological studies. While a transient increase in permeability was shown, the structure of the vessels remained intact. The use of a chick chorioallantoic membrane at different stages of development showed that the damages induced by microbeams depend on vessel maturation. In vivo and ultrastructural observations showed negligible effects of microbeams on the mature vasculature at late stages of development; nevertheless a complete destruction of the immature capillary plexus was found in the microbeam paths. The use of MRT in rodent models revealed a preferential effect on tumor vessels. Although no major modification was observed in the vasculature of normal brain tissue, tumors showed a denudation of capillaries accompanied by transient increased permeability followed by reduced tumor perfusion and finally, a decrease in number of tumor vessels. Thus, MRT is a very promising treatment strategy with pronounced tumor control effects most likely based on the anti-vascular effects of MRT.

Characterization of the 9L gliosarcoma implanted in the Fischer rat: an orthotopic model for a grade IV brain tumor.

Among rodent models for brain tumors, the 9L gliosarcoma is one of the most widely used. Our 9L-European Synchrotron Radiation Facility (ESRF) model was developed from cells acquired at the Brookhaven National Laboratory (NY, USA) in 1997 and implanted in the right caudate nucleus of syngeneic Fisher rats. It has been largely used by the user community of the ESRF during the last decade, for imaging, radiotherapy, and chemotherapy, including innovative treatments based on particular irradiation techniques and/or use of new drugs. This work presents a detailed study of its characteristics, assessed by magnetic resonance imaging (MRI), histology, immunohistochemistry, and cytogenetic analysis. The data used for this work were from rats sampled in six experiments carried out over a 3-year period in our lab (total number of rats = 142). The 9L-ESRF tumors were induced by a stereotactic inoculation of 10(4) 9L cells in the right caudate nucleus of the brain. The assessment of vascular parameters was performed by MRI (blood volume fraction and vascular size index) and by immunostaining of vessels (rat endothelial cell antigen-1 and type IV collagen). Immunohistochemistry and regular histology were used to describe features such as tumor cell infiltration, necrosis area, nuclear pleomorphism, cellularity, mitotic characteristics, leukocytic infiltration, proliferation, and inflammation. Moreover, for each of the six experiments, the survival of the animals was assessed and related to the tumor growth observed by MRI or histology. Additionally, the cytogenetic status of the 9L cells used at ESRF lab was investigated by comparative genomics hybridization analysis. Finally, the response of the 9L-ESRF tumor to radiotherapy was estimated by plotting the survival curves after irradiation. The median survival time of 9L-ESRF tumor-bearing rats was highly reproducible (19-20 days). The 9L-ESRF tumors presented a quasi-exponential growth, were highly vascularized with a high cellular density and a high proliferative index, accompanied by signs of inflammatory responses. We also report an infiltrative pattern which is poorly observed on conventional 9 L tumor. The 9L-ESRF cells presented some cytogenetic specificities such as altered regions including CDK4, CDKN2A, CDKN2B, and MDM2 genes. Finally, the lifespan of 9L-ESRF tumor-bearing rats was enhanced up to 28, 35, and 45 days for single doses of 10, 20, and 2 × 20 Gy, respectively. First, this report describes an animal model that is used worldwide. Second, we describe few features typical of our model if compared to other 9L models worldwide. Altogether, the 9L-ESRF tumor model presents characteristics close to the human high-grade gliomas such as high proliferative capability, high vascularization and a high infiltrative pattern. Its response to radiotherapy demonstrates its potential as a tool for innovative radiotherapy protocols.