Cancer Research Aziza Shad Telomeres
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Cancer Research Clinical Cancer Research
Cancer Epidemiology Biomarkers & Prevention Molecular Cancer Therapeutics
Molecular Cancer Research Cancer Prevention Research
Cancer Prevention Journals Portal Cancer Reviews Online
Annual Meeting Education Book Meeting Abstracts Online
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Biston, M.-C.
Right arrow Articles by Balosso, J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Biston, M.-C.
Right arrow Articles by Balosso, J.
Related Collections
Right arrow Therapeutics and Targets
Right arrow Therapeutics and Targets: Drug Mechanisms and Resistance
[Cancer Research 64, 2317-2323, April 1, 2004]
© 2004 American Association for Cancer Research

Reviews

Cure of Fisher Rats Bearing Radioresistant F98 Glioma Treated with cis-Platinum and Irradiated with Monochromatic Synchrotron X-Rays

Marie-Claude Biston1, Aurélie Joubert1, Jean-François Adam1, Hélène Elleaume1, Sylvain Bohic2, Anne-Marie Charvet1, François Estève1,3, Nicolas Foray1 and Jacques Balosso1,4

1 U647 Institut National de la Santé et de la Recherche Médicale "Rayonnement Synchrotron et Recherche Médicale," ID17 Medical Beamline and 2 ID22 Beamline, European Synchrotron Radiation Facility, Grenoble, France, and 3 Unité Imagerie par Resonance Magnétique and 4 Département de Cancérologie et d’Hématologie, Centre Hospitalier Universitaire A. Michallon, Grenoble, France


   ABSTRACT
Top
 ABSTRACT
Introduction
 Materials and Methods
 Results
 Discussion
 REFERENCES
 
High-grade gliomas are usually of poor prognosis, and conventional radiotherapy, even combined with chemotherapy, still fails to improve the survival of patients. Here, we propose an innovative therapeutic approach combining synchrotron radiation with cis-diamminedichloroplatinum (II) (CDDP). As suggested previously, monochromatic synchrotron irradiation of CDDP at 78.8 keV, just above the 78.4 keV platinum absorption K-edge, leads to an enhanced photoelectric effect and an increased local toxicity. To select a particular radiation energy that could provide supra-additive effect, we used pulsed-field gel electrophoresis to assess yields of DNA double-strand breaks induced in rat F98 glioma cells after CDDP treatment combined with synchrotron X-rays. Thereafter, intracerebral CDDP injection combined with synchrotron X-rays was applied to Fisher rats bearing F98 glioma. CDDP concentrations were mapped by synchrotron X-ray microfluorescence. An extra number of more slowly repaired double strand breaks were observed when irradiating CDDP-treated F98 cells at 78.8 keV. In vivo treatments were then performed with different radiation doses and CDDP concentrations. All cell inoculations in rat brain resulted in tumor development, and tumor presence was controlled by computed tomography. Among all of the conditions tested, the combination of 3 µg of CDDP with 15 Gy resulted in the largest median survival time (206 days). After 1 year, about 34% of treated rats were still alive. This preclinical finding, validated by molecular analysis, represents the most protracted survival reported with this radioresistant glioma model and demonstrates the interest in powerful monochromatic X-ray sources as new tools for cancer treatments.


   Introduction
Top
 ABSTRACT
 Introduction
Materials and Methods
 Results
 Discussion
 REFERENCES
 
The overall incidence rate of gliomas is about 5/100,000, and despite significant advances in cancer therapy, treatment of high-grade gliomas is still palliative. To date, the median survival for patients with glioblastoma is less than 1 year, and virtually no patients with glioblastoma multiforme survive 5 years after treatment (1 , 2) . Although radiotherapy significantly increases survival of patients, gliomas are some of the most radioresistant human tumors. Furthermore, adjuvant chemotherapy generally fails to improve patient outcome, because of inadequate drug delivery and/or concentration inside the tumor (1 , 2) . Hence, considerable efforts were provided to optimize chemo-radiotherapy treatments against brain tumors by increasing both chemotherapeutic drug concentration and radiation dose inside the tumor, while preserving healthy tissues (1) .

Among recent therapeutic techniques, boron neutron capture therapy consists of accumulating a sufficient level of 10B in tumors before irradiating them with neutrons to increase local radiation absorption (3) . Another promising technique, computed tomography therapy, uses a modified X-ray scanner to irradiate brain tumors loaded with iodinated contrast agent. This technique was shown to result in a supra-additive response, perhaps because of an increased photoelectric effect (5, 6, 7, 8) . A monochromatic radiation beam would potentially enhance this physical effect. However, conventional X-ray medical irradiators are not manufactured with sufficient fluence to reach this requirement (9) .

Synchrotrons might overcome this problem: their high fluence (105 times higher than conventional X-ray sources) makes possible a sufficient residual monochromatic beam tuned over tens of keV to allow medical imaging and radiotherapy applications (10) . Over the last 5 years, several therapeutic techniques involving synchrotron radiation have been developed (10) . For example, microbeam radiation therapy, based on multiple parallel collimated synchrotron radiation beams, results in several hundreds of Gy delivered to the tumor bed and has been successfully used on rat brain tumors (11 , 12) . Recently, our group developed a variant computed tomography therapy technique involving synchrotron radiation at the European Synchrotron Radiation Facility (ESRF) (13) . Like microbeam radiation therapy, this technique also consists of delivering a radiation dose excess by targeting vascularization, but not necessarily the DNA of tumor cells.

Theoretically, the irradiation of high-Z elements at their K-edge absorption energy leads to emission of Auger electrons and photoelectrons, releasing a large amount of energy at their immediate vicinity. This phenomenon is called photoactivation (14, 15, 16, 17, 18, 19, 20, 21, 22) . Hence, to irreversibly damage the DNA of tumor cells, an ideal anticancer treatment would consist of a chemo-radiotherapy combining monochromatic synchrotron radiation with DNA-binding photoactivable high-Z compounds (19, 20, 21, 22) . Among the drugs of interest, molecules containing platinum atoms (Z = 78), such as cis-diamminedichloroplatinum (II) (CDDP), are used extensively (23 , 24) . CDDP molecules react with nucleophilic sites into DNA by forming DNA adducts (25) . A number of in vivo studies involving platinum compounds and medical X-rays have already been performed, on either mice or rats bearing various tumors. However, these experiments demonstrate a small increase in life span (26, 27, 28, 29, 30, 31, 32) .

By developing the synchrotron photoactivation of cis-platinum, we have recently proposed a molecular model for describing mechanisms implicated in the combination of CDDP treatment and irradiation with soft X-rays (33) . This treatment was shown to inhibit the major DNA double-strand break (DSB) repair pathway and to induce an extra number of more slowly repaired DSBs, suggesting that CDDP treatment combined with synchrotron X-rays above the K-edge of platinum produces supra-additive molecular effects (33) . However, these data were obtained from experiments on human cells, and our model has not yet been verified in rodents. Hence, in this study, synchrotron CDDP photoactivation was investigated in vitro and in vivo on the radioresistant and weakly immunogenic rat F98 glioma (34) . Our findings show that the combination of CDDP treatment with synchrotron irradiation at 78.0 and 78.8 keV (i.e., just below and just above the platinum K-edge) leads to both an excess of unrepaired DNA damage in treated cells and the most protracted survival obtained with this brain tumor model.


   Materials and Methods
Top
 ABSTRACT
 Introduction
 Materials and Methods
Results
 Discussion
 REFERENCES
 
Cell Culture.
The F98 glioma cell line used in this study was originally established by Drs. A. Koestner and W. Wechsler (Ohio State University; Ref. 34 ). Cells were routinely cultured as monolayers with DMEM (Gibco-Invitrogen-France, Cergy-Pontoise, France) supplemented with 10% FCS, penicillin, and streptomycin for both in vivo and in vitro experiments. All of the experiments were performed at the ESRF.

CDDP Treatment and Irradiation of Cells.
CDDP (Cysplatyl; Rhône-Poulenc, Rorer, Montrouge, France) was kindly provided by Michallon Hospital (Grenoble, France). Cells were treated for 6 h with 30 µM CDDP in DMEM supplemented with 10% serum. They were irradiated in suspension in rotating plastic tubes. The beam size (0.85-mm high) and its homogenous part (8-cm wide) allowed two tubes to be irradiated simultaneously by vertical scanning. To avoid any artifacts due to DNA repair during irradiation, cells were irradiated at 4°C by using a cooling system based on liquid nitrogen vapor diffusion in a polystyrene box containing the rotating tubes. Temperature was monitored by a thermocouple throughout the irradiation. A cylindrical ion chamber coupled with a UNIDOS electrometer and a high-purity germanium detector (Eurisys Mesure, Lingolsheim, France) were used for radiation dose calibration. The dose rate delivered by synchrotron radiation is directly proportional to the storage ring current and was set at 0.1 Gy/scan at the center of the tube. The beam energy was precisely tuned, either 400 eV above (78.8 keV) or below (78.0 keV) the K-edge absorption of platinum (78.4 keV), with an energy bandwidth of 80 eV (35) . Other series of experiments have been performed at 30, 40 and 85 keV. A monochromator based on a pair of dispersive bent Laue Si (1,1,1) crystals mounted in fixed-exit geometry was used for all these settings (36) .

DSB Assays.
DSB induction and repair assays have been described previously in detail (37) . Briefly, cells were irradiated at 4°C and incubated at 37°C at the indicated repair times, when required. Agarose plugs containing 2 x 106 cells·ml–1 were prepared and incubated at 50°C for 38 h in lysis solution containing L-laurylsarcosine and proteinase K (Sigma-Aldrich, St. Louis, MO). Migration of DNA fragments was performed by using pulsed-field gel electrophoresis (CHEF DRII; Bio-Rad, Hercules, CA) with a 4-day migration program discriminating the megabase-sized fragments (37) . Under these particular conditions, only fragments of <15 Mb are able to migrate out of the well. To avoid any artifacts, we did not use radioactivity to label DNA. DSB data were therefore expressed as the fraction of DNA fragments migrating out of the well (FDM) after quantifying the light that each migration lane emitted in the ethidium bromide-stained gel by using a densitometer. Chromosomal S. pombe DNA size standards (Bio-Rad) were used in each experiment to evaluate DNA fragment size all along the migration lane.

Animals and Tumor Cell Inoculation.
Male 7–8-week-old Fisher 344 rats (Charles River Laboratories France, L’Asbresle, France) were used in all experiments. Their weights ranged between 220 and 240 g. All of the operative procedures and animal care were in conformity with the Guidelines of the French Government (Decree 87-848 of October 19, 1987, License 7593 and A38071). Each rat was anesthetized by isoflurane inhalation followed by i.p. infusion of chloral hydrate 4% (0.1 ml/100 g body weight). This procedure maintains animals under anesthesia for about 2 h.

The tumor inoculation technique itself has been described elsewhere (38) . Briefly, each rat was positioned on a stereotaxic frame. An incision was made in the skin 3.5 mm to the right of the bregma. A 0.5-mm-diameter hole for implanting tumor cells was drilled through the skull without breaking the dura. One thousand F98 cells diluted in 5 µl of DMEM were implanted stereotactically at a depth of 6 mm in the cerebral parenchyma. The hole was sealed with bone wax, and the scalp was sutured thereafter. Tumors were allowed to grow for 13 days after implantation. Under our conditions, this procedure resulted in 100% tumor uptake and a median survival time (MeST) of 26 days for the sham untreated controls.

CDDP Injection in Rat Brain.
CDDP inoculation was performed on day 13 after tumor implantation. As described above, the rats were anesthetized and set in a stereotactic frame. The drug was intracerebrally injected at the tumor site. The rats received 3 or 5 µg of CDDP in 5 µl of NaCl isotonic solution. The untreated and irradiated controls were injected with isotonic solution only.

Platinum Content Mapping.
The platinum contents were evaluated by sacrificing some rats 24 h after CDDP inoculation and others 14 days after tumor implantation. The synchrotron induced X-ray fluorescence (SR-XRF) microprobe at ID22 ESRF beamline allows the assessment of elemental concentrations (with Z > 13) in biological samples. SR-XRF set-up was used for intracellular analysis as described elsewhere (39 , 40) . Briefly, a Si (111) monochromator and a Kirkpatrick-Baez X-ray focusing system were used to generate a 14-keV monochromatic X-ray beam. This incident radiation energy allows the K{alpha} fluorescence excitation of elements ranging between Al (Z = 13) and Br (Z = 35) and the L{alpha} fluorescence excitation of Pt (Z = 78). Unstained sections of the entire rat brain (15-µm thick) were deposited on 4-µm thin film target (Ultralene; Spex Certiprep, Metuchen, NJ). This type of backing optimized for SR-XRF analysis is free of contaminants. A spot size of 3 x 10 µm2 (vertical x horizontal) was used. The sample was rastered through the focused beam with a 4-s dwell time, and the entire X-ray fluorescence spectrum was recorded for each map pixel using an energy-dispersive Si(Li) detector (150 eV energy resolution at 5.9 keV). A noncontiguous pixel step-size (50 x 100 µm2) was deliberately chosen because of the large scanned area required (7.6 x 6.4 mm2). This was the best compromise between spatial resolution, measurement stability, and reasonable mapping time (10–12 h). Consequently, some information was lost, which slightly degrades the resolution of the SR-XRF elemental images. The AXIL (analysis of X-ray spectra by iterative least squares) code was used for spectral evaluation to correct the background fluorescence contribution and the overlaps in the fluorescence peaks of neighboring elements (39) . Elemental concentrations (µg·cm–2) were then calculated from areas under peak from thin-film standards [SRM 1832 and 1833; National Institute of Standards and Technology (39 , 40) ].

Irradiation of Rats.
Irradiation was performed stereotactically with monochromatic X-rays from the ESRF synchrotron source. The radiation beam energy was tuned at either 78.8 or 78.0 keV. As described elsewhere, rats were anesthetized at day 14 after implantation, set in a stereotactic frame, and irradiated while rotating. The rotation axis was positioned at the injection point (Fig. 1ACitation ; Ref. 13 ). The radiation beam was fixed at values of 10-mm wide and 0.85-mm high, centered on the injection point. The total scanned height was 12.75 mm, corresponding to fifteen 0.85-mm joined slices. The dose rate was 0.3 Gy·s–1 at the skin entrance. Each tumor was assumed to be surrounded by 1.5 cm of equivalent tissue. As an example, the attenuation coefficient (µatt) at 78.8 keV is 0.1818 cm–1, corresponding to a 24% dose reduction (ICRU4). Hence, the device rotation speed was adjusted to deliver 15 Gy to the tumor, corresponding to 19.7 Gy at the skin entrance.



View larger version (31K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 1. A, schematic illustration of the vertical stereotactic position of rats during radiation treatment. Ear bars and nose sticker maintained rats on the rotating frame. Tumor position is represented by the closed circle in the right hemisphere. The dotted line indicates the rotation axis during radiation treatment. B, after each irradiation, rats received injection with iodinated contrast agent as indicated in "Materials and Methods." Thereafter, tumor computed tomography imaging was performed by taking 12 images in the positions indicated in A. Slice thickness was 1 mm. The tumor is visible on slices 2–5. Ears bars are clearly visible in slice 12.

 
Postirradiation Imaging.
After each irradiation, an image was taken with Iomeron (Bracco, Milan, Italy), an iodinated contrast agent, to verify whether rats bore tumors. It is noteworthy that imaging was performed after radiotherapy to avoid any potential synergy between contrast agent and radiation. Two ml of 350 mg·ml–1 Iomeron solution were injected into the tail vein. Images were taken at 35 keV to get maximal contrast. The radiation dose delivered during imaging is 0.2 Gy (about 0.1 Gy in the tumor). Fig. 1Citation shows a representative example of tumor imaging on a rat surviving 1 year after treatment.

Posttreatment Observations and Histological Studies.
Feeding ability, external appearance, locomotion, and weights of rats were routinely controlled. Rats exhibiting unusual weakness, bleeding eyes, tremors, and loss of weight were euthanized by i.p. Doletal injection (150 mg·kg–1; Vetoquinol, Lure, France). The surviving rats were euthanized 1 year after treatment. The brain of each treated rat was removed and frozen in N-methylbutane. Tissues were then stored at –80°C. Ten-µm brain slices were prepared and stained with hematoxylin and erythrosin.

Statistical Methods.
Each group of rats consisted of 7–18 animals bearing tumors. Survival times were determined from the day of tumor implantation to the day of death. For the euthanized rats, 1 day was added to the life span. All of the animals that died immediately after intracerebral drug injection because of either anesthesia or injection itself were excluded from the statistical study. The mean survival time and MeST were calculated for each group, and Kaplan-Meier survival data were plotted against times after tumor inoculation. The groups were compared using the log-rank test. The percentage increase in life span (%ILS) relative to MeST was defined by the following equation:

where t and u represent treated and untreated animals, respectively. The same formula can be applied to mean survival time to calculate percentage increase in life span relative to mean survival time.


   Results
Top
 ABSTRACT
 Introduction
 Materials and Methods
 Results
Discussion
 REFERENCES
 
DNA DSB Induction and Repair.
DSB induction was examined in rat F98 glioma cells treated with CDDP and irradiated at 30, 40, 78.0, 78.8, and 85 keV. Treatment of F98 cells with CDDP alone did not elicit any detectable DSBs (data not shown). Conversely, synchrotron radiation alone lead to a FDM of about 10%, whatever the radiation energies tested (Fig. 2A)Citation . When combined with 30 µM CDDP, synchrotron radiation resulted in a larger FDM than seen with the two above-mentioned treatments alone. CDDP-treated cells irradiated at 30, 40, and 85 keV showed two and three times lower FDM than at 78.0 and 78.8 keV, respectively, suggesting that DSB induction in CDDP treated cells is strongly enhanced when approaching the platinum K-edge (Fig. 2A)Citation . DSB repair rate was examined thereafter; when CDDP treatment was applied, DSBs induced at 78.8 keV were found to be 1.3, 1.5, and 2 times more slowly repaired than those induced at 78.0, 85, and 30 or 40 keV, respectively (Fig. 2B)Citation . Even if differences in yields of induced and repaired DSB at 78.0 and 78.8 keV are smaller than those obtained previously in SQ20B cells, our findings are consistent with an excess of severe DSBs at 78.8 keV (33) . Photoelectric effect and potential subsequent Auger cascades, more probable at this energy, should supposedly increase energy deposition at close vicinity of CDDP molecules. Consequently, this photoactivation phenomenon should consist of the production of numerous short DNA fragments close to DNA-bound CDDP molecules. To verify this assumption, the 10–100-kb fragment region of the gel was analyzed; an excess of FDM was observed in CDDP-treated cells at 78.0 and 78.8 keV compared with the other energies tested (Fig. 2C)Citation . In addition, the FDM was 1.5 times larger at 78.8 keV than at 78.0 keV. Taken together, these findings strongly suggest that to irradiate CDDP-treated cells at an energy above the platinum K-edge results in a supra-additive molecular effect. The biological relevance of these molecular findings was then examined in rats bearing brain tumors.



View larger version (15K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 2. A, yields of double-strand break (DSB) induced, assessed immediately after treatment of F98 cells irradiated at 4°C (30 Gy) at different synchrotron radiation energies in presence of 30 µM cis-diamminedichloroplatinum (II) (CDDP). DSB data are expressed in fraction of DNA fragments migrating out of the gel (FDM). Data represent the mean ± SE of two replicate experiments. B, DSB repair data from 30 µM CDDP-treated F98 cells irradiated (30 Gy) at different synchrotron radiation energies, expressed as percentage of FDM and plotted as a function of postirradiation time at 37°C. The dotted line links FDM data obtained from cells treated with only 78.8 keV radiation. These last FDM data are not significantly different from those obtained with irradiation alone, whatever the energies tested (data not shown). Data represent the mean ± SE of two replicate experiments. C, representative example of an estimation of FDM concerning only the 10–100-kb fragment region in an electrophoresis gel analysis (whole data are shown in A).

 
Preparation of Preclinical Trials and Platinum Content in Treated Tumors.
From preliminary data with rats bearing brain tumors, the efficiency of the anticancer treatment appeared to be strongly influenced by the CDDP microdistribution within the tumor. Hence, by using SR-XRF, platinum content was assessed after CDDP injection in rat brain. To our knowledge, this sensitive technique has never been applied to brain histological samples at this scale. Thirteen days after brain tumor inoculation, rats received intracerebral injection with 3 µg of CDDP, and brain sections were prepared the next day. Fig. 3Citation shows the potassium, zinc, iron, and platinum distributions in the whole treated rat brain hemisphere. Potassium and zinc, which were more highly concentrated in the cortex part, reflected the structure of the brain tissue (Fig. 3, A and B)Citation . The needle route was revealed by iron contained in the large amount of red blood cells due to the injection wound (Fig. 3C)Citation . Platinum mapping showed that CDDP molecules were highly concentrated at the end of the needle pathway (average, 0.18 µg·cm–2; some hot spots at 0.62 µg·cm–2; Fig. 3DCitation ). An average platinum concentration of 0.08 µg·cm–2 was assessed around the high concentration zone (50–100 times higher than that after i.p. injections), and 0.03 µg·cm–2 was found in the edges of the inoculated hemisphere (Refs. 41 and 42 ; Fig. 3DCitation ). No traces of platinum were detected in the contralateral hemisphere. It is noteworthy that, by using argon-coupled plasma mass spectrometry, a constant level of platinum remains in the inoculated hemisphere 48 h after injection (data not shown). Taken together, these findings show that CDDP molecules are very abundant in the region of interest and diffuse very slowly after injection.



View larger version (108K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 3. Potassium (A), zinc (B), iron (C), and platinum (D) concentration mapping carried out in a 15-µm histological cut of a rat inoculated with 3 µg of cis-diamminedichloroplatinum (II) (CDDP) on day 13 by using synchrotron X-ray fluorescence. The needle route (arrows) is revealed by iron contained in the large amount of RBCs due to the injection. The platinum mapping reflects high CDDP concentrations at the end of the needle pathway, where the drug has been released.

 
Survival of Rats Bearing F98 Glioma.
CDDP treatment combined with synchrotron irradiation at energies surrounding the platinum K-edge was applied to Fisher rats bearing F98 glioma. All of the F98 cell inoculations (103 cells) resulted in tumor development, clearly distinguishable by computed tomography on day 14 (Fig. 1)Citation . At this time, the average tumor volume was estimated to be 20 ± 10 mm3, corresponding to about 15% of an average rat brain volume (1500 mm3). To optimize rat survival, several CDDP and radiation doses (3–5 µg and 5–15 Gy, respectively) were tested. Fig. 4Citation shows MeSTs obtained from different combined treatments such as 5 µg of CDDP + 5 Gy, 5 µg of CDDP + 10 Gy, and 3 µg of CDDP + 15 Gy. The MeST of sham untreated rats was found to be 26 days (Table 1)Citation . Compared with controls, CDDP intracerebral injections of 3 and 5 µg significantly improved survival, with a MeST of 37 and 39.5 days, respectively (P < 0.0018; Fig. 4Citation ). When synchrotron radiation alone was applied (5, 10, and 15 Gy), the MeST was found to be 31, 36, and 48 days, respectively. When combined with CDDP treatment, a dose of 5 and 10 Gy provided a MeST of about 53 days. Increasing the dose up to 15 Gy at 78.0 and 78.8 keV drastically enhanced the MeST, which reached 214 and 194 days, respectively (Fig. 4)Citation . CDDP treatment (3 µg) combined with 15 Gy of synchrotron irradiation at 78.0 and 78.8 keV appeared to be the most efficient treatment tested and was thereafter systematically applied to rats bearing F98 gliomas. Fig. 5ACitation shows the in vivo survival curves from the first series of experiment; an obvious gain of survival was obtained with the combined treatment by comparison with the 15-Gy irradiation alone (P < 0.0001). One year after the 3 µg of CDDP + 15 Gy treatment, 33% (6 of 18) of treated rats were still alive. The same experiment was performed again 4 months later. Although a higher early death rate was observed in the second experiment, survivals were not significantly different from those of the first experiment, whatever the radiation energies tested (78.0 and 78.8 keV; P > 0.6; Fig. 5BCitation ; Table 1Citation ). Similar to the first series, 36% (4 of 11) of treated rats were still alive 1 year after treatment.



View larger version (14K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 4. Median survival time (MeST) expressed in days after tumor inoculation and indicated treatments of rats bearing F98 glioma. Because no significant differences have been observed between MeST from irradiation treatments alone at 78.0 and 78.8 keV, only the MeST from 78.8 keV irradiation is represented. Each group of treated rats contained at least six animals. The untreated control group contained at least 10 rats.

 

View this table:
[in this window]
[in a new window]

  		Table 1 Survival data from rats bearing F98 glioma and treated with the indicated protocols

 


View larger version (11K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 5. Kaplan-Meier survival curves of rats bearing F98 glioma and subjected to the indicated treatments. The survivals are plotted against time (days) after tumor inoculation. A: {blacktriangleup}, untreated controls; {triangledown}, 3 µg of cis-diamminedichloroplatinum (II) (CDDP) alone; {circ}, 15 Gy alone; {blacktriangledown}, 3 µg of CDDP combined with irradiation at 78.0 keV; {bullet}, 3 µg of CDDP combined with irradiation at 78.8 keV. B, intercomparisons between survival fractions obtained from two series of independent experiments with 3 µg of CDDP treatment combined with irradiation (15 Gy) at 78.0 ({blacktriangledown}) or 78.8 keV ({bullet}). Data from experiment 1 and 2 are represented by solid and dotted lines, respectively. The gray confidence zone under the curve combines the survival fraction of untreated controls, rats treated with 3 µg of CDDP and rats treated with 15 Gy.

 
Histology.
Among the surviving rats, we investigated the histopathological changes in the brains of treated rats. To this end, the brains of some rats subjected to the 3 µg of CDDP + 15 Gy treatment were stained with hematoxylin and erythrosin. The brain from a rat that died at day 177 showed evidence of tumor growth with necrotic areas (Fig. 6, A and B)Citation . Conversely, the brain of a rat sacrificed 1 year after treatment exhibited an acellular pseudo cyst cavity surrounded by a fibrosis margin but no trace of residual tumor (Fig. 6, C and D)Citation , suggesting its irreversible regression.



View larger version (128K):
[in this window]
[in a new window]
[Download PPT slide]
  		Fig. 6. Histopathological changes in the brain from a rat that died at day 177 (A and B) and the brain from a cured rat sacrificed 1 year after treatment (C and D). A shows evidence of a tumor extension whose necrotic regions (1) and F98 tumor cells (2) are magnified in B. C shows absence of residual tumor with a cellular pseudo cyst cavity (3) surrounded by a fibrosis margin (4) magnified in D.

 

   Discussion
Top
 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
REFERENCES
 
In this study, we attempted to optimize a chemo-radiotherapy protocol against aggressive glioma consisting of CDDP treatment combined with monochromatic synchrotron radiation. Our in vitro findings suggest that CDDP treatment combined with irradiation just above the platinum K-edge results in a supra-additive molecular effect. In vivo experiments show that the combination of 3 µg of CDDP and 15 Gy monochromatic radiation resulted in the most protracted survival reported with the radioresistant F98 glioma.

The Choice of the High-Z Photoactivable Compound.
Previous evidence has already suggested that chemo-radiotherapy treatment is more efficient than radiotherapy or chemotherapy alone. CDDP was deliberately chosen as a chemotherapeutic agent because of its DNA binding properties, the high-Z number of platinum, and its frequent use against ovarian, testicular, bladder, and squamous cell carcinomas and small cell lung carcinomas, especially in combination with conventional radiotherapy (43) . However, the efficiency of CDDP treatment remains to be confirmed against primary and metastatic brain tumors (44, 45, 46, 47) .

Molecular mechanisms at the origin of the supra-additive effect observed with the combined treatment with CDDP and X-rays are not yet fully understood. CDDP induces DNA adducts, which are generally repaired by a recombination-like pathway. In parallel, ionizing radiation produces DSBs, which are more likely recognized by the Ku protein and rejoined by nonhomologous end joining, suggesting that CDDP treatment combined with radiation leads to the interplay of two major DNA repair pathways (25 , 33 , 48 , 49) . Moreover, concentrated CDDP was shown to prevent the translocation of Ku all along the DNA, affecting the nonhomologous end joining pathway and contributing to cell lethality (50) . Hence, particular efforts were provided to optimize the CDDP injection protocol resulting in the highest platinum concentrations in rat brains. These requirements were reached by intracerebral CDDP injection. The assessment of platinum concentrations with the SR-XRF technique under the conditions of in vivo treatment confirms that the highest CDDP levels are reached around the injection point and are clearly higher than those seen when CDDP is injected i.p. (Fig. 3)Citation .

Molecular Findings.
In presence of CDDP, yields of radiation-induced DSBs were found to be 2–3 times larger at 78.0 and 78.8 keV than at any other energy tested (30, 40, and 85 keV). Our DSB data are therefore consistent with an excess of the delivered dose in the immediate vicinity of the CDDP molecules bound to DNA, the production of smaller DNA fragments at 78.0 and 78.8 keV than at the other energies, the generation of more severe damage with more DSBs surrounding DNA adducts, and the difficulty for tumor cells to repair these DSBs in conditions of CDDP photoactivation. Hence, it is shown here that the biological efficiency of synchrotron photoactivation of platinum is strongly dependent on monochromatic energy of radiation treatment.

When compared with the other energies tested, the efficiency of the combined treatment is maximal at 78.0 and 78.8 keV in CDDP-treated cells. However, significant differences also exist between these two energies: induced DSB yields were found to be higher at 78.8 keV than at 78.0 keV. Such an effect was reported previously (33) , but with a 1:3 ratio, in human SQ20B carcinoma cells subjected to the same treatment. Unrepaired DSB yields were found to be 1.2 and 2 times higher at 78.8 keV than at 78.0 keV in F98 and SQ20B cells, respectively (33) . These differences between SQ20B and F98 cells do not affect our general conclusion and may be attributed to species-dependent features (more condensed chromatin and faster DSB repair rate in rodents than in humans; Ref. 51 ). The differences observed in vitro between 78.0 and 78.8 keV are not detectable in survival curves. The magnitude of the extra number of DNA damage between 78.8 and 78.0 keV may be insufficient to induce differences in vivo; one typical example is the case of ATM- and LIG4-mutated cells that, despite similar cellular radiosensitivity, exhibit a 1:3 ratio with regard to unrepaired DSB yields (52) . Our DSB assays only take into account nonrepaired but not misrepaired DSBs, which may also contribute to cellular lethality. Further investigations are required to examine whether larger differences observed in vitro may lead to any detectable effect in vivo.

The Choice of Animal Tumor Model and Rat Survival Intercomparisons.
The F98 tumor model, which is known to be refractory to a number of therapeutic modalities, has been deliberately chosen because of its extreme radioresistance (surviving fraction at 2 Gy of about 75%) and its proliferation capacities (34) . Furthermore, F98 glioma is a very aggressive model, perhaps due to its weak immunogenicity. Note that some unexpected immune responses have been reported with 9L gliosarcoma, one of the most widely used rat brain tumor models (34) .

An impressive MeST (206 days) was found in rats treated with 3 µg of CDDP and irradiated at 78.0 and 78.8 keV (15 Gy). This result represents the most protracted survival reported with the radioresistant F98 glioma (Table 2)Citation . In the two series of experiments, about 34% of rats survived 1 year after treatment and appeared healthy. By using the same F98 tumor model, Kaneko et al. (31) reported a MeST of 31 days for the untreated controls (26 days here) and 55 days for rats treated with CDDP and irradiated with X-rays (206 days here). CDDP was injected i.p., which may lead to limited platinum levels inside tumors (31) . Up to now, boron neutron capture therapy was considered the most efficient therapy when using the F98 model: with the same conditions (number of inoculated cells, day of irradiation) and very similar MeST of untreated controls, Barth et al. (53) reported a MeST of 72 days for the treated rats. Interestingly, boron neutron capture therapy applied to another model, the 9L gliosarcoma, provided an impressive 100% rat survival at 250 days after treatment (4) . With a protocol involving both intracerebral injection of 3 µg of CDDP and 16 Gy of ionizing radiation, Kimler et al. (32) reported a MeST of 40 days for 9L-bearing rats. Note that in all of the above studies, the actual presence of tumor was not systematically verified by imaging.


View this table:
[in this window]
[in a new window]

  		Table 2 Review of in vivo studies dealing with rats bearing brain tumors

 
Taken together, our data demonstrate the efficiency of an innovative anticancer treatment involving both CDDP and synchrotron radiation. To emphasize the actual feasibility of the proposed therapeutic protocol and its transfer to clinical trials, it is noteworthy that several intracerebral drug injections have been performed in patients suffering from brain tumors (54 , 55) . Further investigations are in progress at ESRF, notably to introduce high concentrations of drugs containing platinum atoms into tumors. As an example, carboplatin may be an interesting alternative to cisplatin because of its low nephro- and neuro-toxicity (56) . Hence, our approach, validated by molecular analysis and preclinical trials, demonstrates the possibility of powerful monochromatic X-ray sources as new tools against solid tumors.


   ACKNOWLEDGMENTS
 
We are grateful to Dr. S. Corde (Waikato Hospital, Pembroke St Hamilton, New Zealand), Dr. V. Favaudon and C. Reis (Institut Curie Recherche, U350 INSERM, Orsay, France), K. Gamo (University of California Los Angeles, Los Angeles, CA), Dr. V. Ducros (Département de Biologie Intégée, CHU, Grenoble, France), and Prof. M. Peoc’h (Laboratoire Anatomie et Cytologie Pathologiques, CHU, Saint-Etienne, France) for fully fruitful discussions and help. We thank R. Farion (INSERM U594, Grenoble, France) and T. Brochard, D. Dallery, G. Le Duc, C. Nemoz, I. Troprès, and M. Rénier (ESRF, Grenoble, France) for technical support. We were able to perform these experiments thanks to the ESRF beam shift allocation on ID17 and ID 22.


   FOOTNOTES
 
Grant support: The Association pour la Recherche contre le Cancer, the Electricité de France (Comité de Radioprotection), and the European Synchrotron Radiation Facility (Grenoble, France). M-C. Biston was supported by a fellowship from Région Rhône-Alpes.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Requests for reprints: Correspondence to: Nicolas Foray, U647 INSERM "Rayonnement Synchrotron et Recherche Médicale," ID17, European Synchrotron Radiation Facility BP 220, 38043 Grenoble, France. Phone: 33-4-76-88-25-41; Fax: 33-4-76-88-28-85; E-mail: foray{at}esrf.fr

Received 11/17/03. Revised 1/23/04. Accepted 2/ 5/04.


   REFERENCES
Top
 ABSTRACT
 Introduction
 Materials and Methods
 Results
 Discussion
 REFERENCES
 

  1. Behin A, Hoang-Xuan K, Carpentier AF, Delattre JY Primary brain tumours in adults. Lancet, 361: 323-31, 2003.[CrossRef][Medline]
  2. Bredel M Anticancer drug resistance in primary human brain tumors. Brain Res Rev, 35: 161-204, 2001.[CrossRef][Medline]
  3. Joel DD, Fairchild RG, Laissue JA, et al Boron neutron capture therapy of intracerebral rat gliosarcomas. Proc Natl Acad Sci USA, 87: 9808-12, 1990.[Abstract/Free Full Text]
  4. Coderre JA, Button TM, Micca PL, et al Neutron capture therapy of the 9L rat gliosarcoma using the p-boronophenylalanine-fructose complex. Int J Radiat Oncol Biol, 30: 643-52, 1994.[Medline]
  5. Norman A, Iwamoto KS, Cochran ST Iodinated contrast agents for brain tumor localization and radiation dose enhancement. Investig Radiol, 26(Suppl. 1): S120-1, discussion S5–8. 1991.
  6. Rose JH, Norman A, Ingram M, et al First radiotherapy of human metastatic brain tumors delivered by a computerized tomography scanner (CTRx). Int J Radiat Oncol Biol Phys, 45: 1127-32, 1999.[CrossRef][Medline]
  7. Dawson P, Penhaligon M, Smith E, Saunders J Iodinated contrast agents as "radiosensitizers.". Br J Radiol, 60: 201-3, 1987.[Abstract/Free Full Text]
  8. Robar JL, Riccio SA, Martin MA Tumour dose enhancement using modified megavoltage photon beams and contrast media. Phys Med Biol, 47: 2433-49, 2002.[CrossRef][Medline]
  9. Mesa AV, Norman A, Solberg TD, Demarco JJ, Smathers JB Dose distributions using kilovoltage X-rays and dose enhancement from iodine contrast agents. Phys Med Biol, 44: 1955-68, 1999.[CrossRef][Medline]
  10. Suortti P, Thomlinson W Medical applications of synchrotron radiation. Phys Med Biol, 48: R1-35, 2003.[CrossRef][Medline]
  11. Dilmanian FA, Button TM, Le Duc G, et al Response of rat intracranial 9L gliosarcoma to microbeam radiation therapy. Neuro-Oncol, 4: 26-38, 2002.[Abstract/Free Full Text]
  12. Slatkin DN, Spanne P, Dilmanian FA, Gebbers JO, Laissue JA Subacute neuropathological effects of microplanar beams of X-rays from a synchrotron wiggler. Proc Natl Acad Sci USA, 92: 8783-7, 1995.[Abstract/Free Full Text]
  13. Adam JF, Elleaume H, Joubert A, et al Synchrotron radiation therapy of malignant brain glioma loaded with an iodinated contrast agent: first trial on rats bearing F98 gliomas. Int J Radiat Oncol Biol Phys, 57: 1413-26, 2003.[CrossRef][Medline]
  14. Tisljar Lentulis G, Feinendegen LE, Bond VP Biological radiation effects of inclusion of moderately heavy nuclei into the tissue and use of soft roentgen rays. Strahlentherapie, 145: 656-62, 1973.[Medline]
  15. Tisljar Lentulis G, Feinendegen LE The bromine enhancement ratio in mammalian cells in vitro and experimental mouse tumours. Curr Top Radiat Res, 12: 526-36, 1978.
  16. O Donoghue JA, Wheldon TE Targeted radiotherapy using Auger electron emitters. Phys Med Biol, 41: 1973-92, 1996.[CrossRef][Medline]
  17. Fairchild R, Bond VP Photon activation therapy. Strahlentherapie, 160: 758-63, 1984.[Medline]
  18. Hofer KG Biophysical aspects of Auger processes. Acta Oncol, 39: 651-7, 2000.[CrossRef][Medline]
  19. Laster BH, Thomlinson WC, Fairchild RG Photon activation of iododeoxyuridine: biological efficacy of Auger electrons. Radiat Res, 133: 219-24, 1993.[Medline]
  20. Shinohara K, Ohara H, Kobayashi K, et al Enhanced killing of HeLa cells pre-labeled with 5-bromodeoxyuridine by monochromatic synchrotron radiation at 0.9 A: an evidence for Auger enhancement in mammalian cells. J Radiat Res, 26: 334-8, 1985.
  21. Larson D, Bodell WJ, Ling C, et al Auger electron contribution to bromodeoxyuridine cellular radiosensitization. Int J Radiat Oncol Biol Phys, 16: 171-6, 1989.[Medline]
  22. Jonsson AC, Jonsson BA, Strand SE, Grafstrom G, Spanne P Cell survival after Auger electron emission from stable intracellular indium exposed to monochromatic synchrotron radiation. Acta Oncol, 35: 947-52, 1996.[Medline]
  23. Kelland LR Overcoming resistance to platinum therapy in patients with advanced cancer. Am J Cancer, 1: 247-55, 2002.
  24. Screnci D, Mc Keage MJ Platinum neurotoxicity: clinical profiles, experimental models and neuroprotective approaches. J Inorg Biochem, 77: 105-10, 1999.[CrossRef][Medline]
  25. Dewit L Combined treatment of radiation and cis-diamminedichloroplatinum (II): a review of experimental and clinical data. Int J Radiat Oncol Biol Phys, 13: 403-26, 1987.[Medline]
  26. Twentyman PR, Kallman RF, Brown JM The effect of time between X-irradiation and chemotherapy on the growth of three solid mouse tumors. III. Cis-diamminedichloroplatinum. Int J Radiat Oncol Biol Phys, 5: 1365-7, 1979.[Medline]
  27. Overgaard J, Khan AR Selective enhancement of radiation response in a C3H mammary carcinoma by cisplatin. Cancer Treat Rep, 65: 501-3, 1981.[Medline]
  28. Lelieveld P, Scoles MA, Brown JM, Kallman RF The effect of treatment in fractionated schedules with the combination of X-irradiation and six cytotoxic drugs on the RIF-1 tumor and normal mouse skin. Int J Radiat Oncol Biol Phys, 11: 111-21, 1985.[Medline]
  29. Teicher BA, Holden SA Antitumor and radiosensitizing activity of several platinum-(+) dye complexes. Radiat Res, 109: 58-67, 1987.[CrossRef][Medline]
  30. Douple EB, Richmond RC, Logan Mark E Therapeutic potentiation in a mouse mammary tumor and an intracerebral rat brain tumor by combined treatment with cis-dichlorodiammineplatinum (II) and radiation. J Clin Hematol Oncol, 7: 585-603, 1977.
  31. Kaneko S, Clendenon NR, Kartha M Experimental study in a rat brain tumor by combined treatment with cis-DDP and irradiation. Neurol Med Chir, 23: 917-23, 1983.[CrossRef]
  32. Kimler BF, Liu C, Evans RG, Morantz RA Combination of aziridinylbenzoquinone and cis-platinum with radiation therapy in the 9L rat brain tumor model. Int J Radiat Oncol Biol Phys, 26: 445-50, 1993.[Medline]
  33. Corde S, Balosso J, Elleaume H, et al Synchrotron photoactivation of cisplatin elicits an extra number of DNA breaks that stimulate RAD51-mediated repair pathways. Cancer Res, 63: 3221-7, 2003.[Abstract/Free Full Text]
  34. Barth RF Rat brain tumor models in experimental neuro-oncology: the 9L, C6, T9, F98, RG2 (D74), RT-2 and CNS-1 gliomas. J Neuro-Oncol, 36: 91-102, 1998.[CrossRef][Medline]
  35. Suortti P, Fiedler S, Bravin A, et al Fixed-exit monochromator for computed tomography with synchrotron radiation at energies 18–90 keV. J. Synchrotron Radiat, 7: 340-7, 2000.
  36. Elleaume H, Charvet AM, Berkvens P, et al Instrumentation of the ESRF medical imaging facility. Methods Phys Res Section A, 428: 513-27, 1999.[CrossRef]
  37. Foray N, Priestley A, Alsbeih G, et al Hypersensitivity of ataxia telangiectasia fibroblasts to ionizing radiation is associated with a repair deficiency of DNA double-strand breaks. Int J Radiat Biol, 72: 271-83, 1997.[CrossRef][Medline]
  38. Benabid AL, Rémy C, Chauvin C Experimental model of rat brain tumors by stereotactic injection of C6 glioma and HTC hepatoma cell lines Walker MD Thomas DGT eds. . Biology of brain tumors, p. 221-6, Martinus Nijhoff Publishers Amsterdam 1986.
  39. Bohic S, Simionovici A, Snigirev A, et al Synchrotron induced x-ray fluorescence on single cells. Appl Phys Lett, 78: 3544 2001.[CrossRef]
  40. Corde S, Biston MC, Elleaume H, et al Lack of cell death enhancement after irradiation with monochromatic synchrotron X rays at the K-shell edge of platinum incorporated in living SQ20B human cells as cis-diamminedichloroplatinum (II). Radiat Res, 158: 763-70, 2002.[CrossRef][Medline]
  41. Douple EB, Totten MD, Spencer F Platinum levels in murine tumor following intraperitoneal administration of cisplatin or paraplatin. Natl Cancer Inst Monogr, 6: 129-32, 1988.
  42. Minami T, Okazaki J, Kawabata A, Kuroda R, Okazaki Y Penetration of cisplatin into mouse brain by lipopolysaccharide. Toxicology, 130: 107-13, 1998.[CrossRef][Medline]
  43. Olivi A, Ewend MG, Utsuki T, et al Interstitial delivery of carboplatin via biodegradable polymers is effective against experimental glioma in the rat. Cancer Chemother Pharmacol, 39: 90-6, 1996.[CrossRef][Medline]
  44. Witman G, Cadman E, Kapp D, Wagner F The use of cisplatinum for treatment of malignant glioma. Med Hypotheses, 8: 335-41, 1982.[CrossRef][Medline]
  45. European Organisation for Research and Treatment of Cancer. Cisplatin does not enhance the effect of radiation therapy in malignant gliomas. Eur J Cancer, 27: 568-71, 1991.
  46. Boiardi A, Silvani A, Pozzi A, et al Advantage of treating anaplastic gliomas with aggressive protocol combining chemotherapy and radiotherapy. J Neuro-Oncol, 34: 179-85, 1997.[CrossRef][Medline]
  47. Freeman CR, Kepner J, Kun LE, et al A detrimental effect of a combined chemotherapy-radiotherapy approach in children with diffuse intrinsic brain stem gliomas?. Int J Radiat Biol Phys, 47: 561-4, 2000.
  48. Husain A, He G, Venkatraman ES, Spriggs DR BRCA1 up-regulation is associated with repair-mediated resistance to cis-diamminedichloroplatinum(II). Cancer Res, 58: 1120-3, 1998.[Abstract/Free Full Text]
  49. Jeggo PA Identification of genes involved in repair of DNA double-strand breaks in mammalian cells. Radiat Res, 150: S80-91, 1998.[Medline]
  50. Turchi JJ, Henkels KM, Zhou Y Cisplatin-DNA adducts inhibit translocation of the Ku subunits of DNA-PK. Nucleic Acids Res, 28: 4634-41, 2000.[Abstract/Free Full Text]
  51. Iliakis G The role of DNA double strand breaks in ionizing radiation-induced killing of eukaryotic cells. Bioessays, 13: 641-8, 1991.[CrossRef][Medline]
  52. Badie C, Iliakis G, Foray N, et al Induction and rejoining of DNA double-strand breaks and interphase chromosome breaks after exposure to X rays in one normal and two hypersensitive human fibroblast cell lines. Radiat Res, 144: 26-35, 1995.[Medline]
  53. Barth RF, Yang W, Rotaru JH, et al Boron neutron capture therapy of brain tumors: enhanced survival and cure following blood-brain barrier disruption and intracarotid injection of sodium borocaptate and boronophenylalanine. Int J Radiat Oncol Biol Phys, 47: 209-18, 2000.[CrossRef][Medline]
  54. Gutman RL, Peacock G, Lu DR Targeted drug delivery for brain cancer treatment. J Controlled Release, 65: 31-41, 2000.[CrossRef][Medline]
  55. Walter KA, Tamargo RJ, Olivi A, Burger PC, Brem H Intratumoral chemotherapy. Neurosurgery, 37: 1128-45, 1995.[Medline]
  56. Yang LX, Douple EB, O’Hara JA, Wang HJ Production of DNA double-strand breaks by interactions between carboplatin and radiation: a potential mechanism for radiopotentiation. Radiat Res, 143: 309-15, 1995.[CrossRef][Medline]



This article has been cited by other articles:


Home page
 J R Soc InterfaceHome page
J. E. Pattison, R. P. Hugtenburg, and S. Green
Enhancement of natural background gamma-radiation dose around uranium microparticles in the human body
J R Soc Interface, September 23, 2009; (2009) rsif.2009.0300v1.
[Abstract] [Full Text] [PDF]

Home page
 Radiat Prot DosimetryHome page
M. Ghasemi, M. Shamsaei, M. Ghannadi, and G. Raisali
DOSIMETRIC STUDIES OF MICROPENCIL X-RAY BEAM INTERACTING WITH LABELLED TISSUES BY AU AND GD AGENTS USING GEANT4
Radiat Prot Dosimetry, February 17, 2009; (2009) ncp017v1.
[Abstract] [Full Text] [PDF]

Home page
 Clin. Cancer Res.Home page
J. Rousseau, C. Boudou, R. F. Barth, J. Balosso, F. Esteve, and H. Elleaume
Enhanced Survival and Cure of F98 Glioma Bearing Rats following Intracerebral Delivery of Carboplatin in Combination with Photon Irradiation
Clin. Cancer Res., September 1, 2007; 13(17): 5195 - 5201.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
F. A. Dilmanian, Z. Zhong, T. Bacarian, H. Benveniste, P. Romanelli, R. Wang, J. Welwart, T. Yuasa, E. M. Rosen, and D. J. Anschel
Interlaced x-ray microplanar beams: A radiosurgery approach with clinical potential
PNAS, June 20, 2006; 103(25): 9709 - 9714.
[Abstract] [Full Text] [PDF]

Home page
 Clin. Cancer Res.Home page
G. De Stasio, D. Rajesh, J. M. Ford, M. J. Daniels, R. J. Erhardt, B. H. Frazer, T. Tyliszczak, M. K. Gilles, R. L. Conhaim, S. P. Howard, et al.
Motexafin-Gadolinium Taken Up In vitro by at Least 90% of Glioblastoma Cell Nuclei
Clin. Cancer Res., January 1, 2006; 12(1): 206 - 213.
[Abstract] [Full Text] [PDF]

Home page
 Radiat Prot DosimetryHome page
S. Corde, J.-F. Adam, M.-C. Biston, A. Joubert, A.-M. Charvet, F. Esteve, J.-F. Le Bas, H. Elleaume, and J. Balosso
Sensitivity variation of doped Fricke gel irradiated with monochromatic synchrotron X rays between 33.5 and 80 keV
Radiat Prot Dosimetry, December 1, 2005; 117(4): 425 - 431.
[Abstract] [Full Text] [PDF]

Home page
 Clin. Cancer Res.Home page
W. Yang, R. F. Barth, G. Wu, M. J. Ciesielski, R. A. Fenstermaker, B. A. Moffat, B. D. Ross, and C. J. Wikstrand
Development of a Syngeneic Rat Brain Tumor Model Expressing EGFRvIII and Its Use for Molecular Targeting Studies with Monoclonal Antibody L8A4
Clin. Cancer Res., January 1, 2005; 11(1): 341 - 350.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Biston, M.-C.
Right arrow Articles by Balosso, J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Biston, M.-C.
Right arrow Articles by Balosso, J.
Related Collections
Right arrow Therapeutics and Targets
Right arrow Therapeutics and Targets: Drug Mechanisms and Resistance

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Cancer Research Clinical Cancer Research
Cancer Epidemiology Biomarkers & Prevention Molecular Cancer Therapeutics
Molecular Cancer Research Cancer Prevention Research
Cancer Prevention Journals Portal Cancer Reviews Online
Annual Meeting Education Book Meeting Abstracts Online