C6 Glioma Model Research Articles

Role of cAMP-dependent protein kinase A activity in low-dose endothelial monocyte-activating polypeptide-II-induced opening of blood-tumor barrier.

Our previous studies demonstrated that low-dose endothelial monocyte-activating polypeptide-II (EMAP-II) can selectively increase the permeability of blood-tumor barrier (BTB). In addition, low-dose EMAP-II significantly decreases the cyclic adenosine monophosphate (cAMP) concentration and the protein kinase A (PKA) expression level in tumor tissues in the rat C6 glioma model. In this study, an in vitro BTB model was used to investigate the potential role of cAMP/PKA signaling cascade in EMAP-II-induced BTB hyperpermeability. Our data revealed that low-dose EMAP-II (0.05nM) induced a significant decrease in total intracellular cAMP concentration and PKA activity in rat brain microvascular endothelial cells (RBMECs). Pretreatment with forskolin to increase intracellular cAMP nearly completely blocked the EMAP-II-induced decrease in transendothelial electric resistance and increase in horseradish peroxidase flux across the BTB. Similar pretreatment completely prevented the EMAP-II-induced changes in RhoA/Rho kinase activity, expression and distribution of tight junction-associated protein ZO-1, and myosin light chain phosphorylation, as well as actin cytoskeleton arrangement in RBMECs. Pretreatment with 6Bnz-cAMP to activate PKA significantly attenuated these EMAP-II-induced alterations in RBMECs. In summary, our present study demonstrates that the cAMP/PKA signaling cascade works as a crucial signaling pathway in EMAP-II-induced BTB hyperpermeability.

Interactions of connexin 43 and aquaporin-4 in the formation of glioma-induced brain edema.

Connexin 43 (Cx43) and aquaporin-4 (AQP4) have important roles in the formation of glioma-induced brain edema; however, the association between these two factors in the development of edema has remained to be elucidated. In the present study, immunofluorescence and western blot analysis revealed that in a rat model of intracranial C6 glioma, Cx43 expression levels were low to undetectable and AQP4 expression levels were low in glioma cells. Significantly higher Cx43 and AQP4 levels were detected in the tissue surrounding the glioma. To further investigate the potential interaction between Cx43 and AQP4, normal glial cells and C6glioma cells were cultured in hypotonic medium. Reverse transcription quantitative polymerase chain reaction indicated that AQP4 and Cx43 mRNA expression levels increased as a function of time in normal glial cells and C6 glioma cells in a hypotonic environment. However, the increase observed in normal glial cells was significantly lower than that observed in C6glioma cells. Furthermore, AQP4 expression levels changed prior to alterations in Cx43 expression. Following AQP4 silencing in C6cells, the increase in Cx43 expression was significantly attenuated (P<0.05). In normal cells, Cx43 silencing did not influence AQP4 expression (P>0.05). Therefore, it was hypothesized that AQP4 and Cx43 had two distinct mechanisms underlying brain edema formation within and surrounding the glioma. Cx43 may be a downstream effector of AQP4. The elucidation of this pathway may aid in the development of drugs targeting the interaction between AQP4 and Cx43, providing novel therapeutic possibilities for glioma-induced brain edema.

Combination of photodynamic therapy and temozolomide on glioma in a rat C6 glioma model

For glioma, temozolomide (TMZ) is a commonly used chemotherapy drug and photodynamic therapy (PDT) is an important adjuvant therapy. The aim of this study was to evaluate the effect of their combination for the treatment of glioma. A rat C6 glioma model using male Wistar rats (n=180) weighing 280-300 g was established. Glioma-bearing rats (n=100) were treated with mock, hematoporphyrin monomethyl ether (HMME), laser or PDT. The expression of P-glycoprotein (P-gp) in endothelial cells of the blood-tumor-barrier and in glioma tissues was detected using immunohistochemistry and western blot, respectively. Glioma-bearing rats (n=40) were treated with normal saline, TMZ (60 mg/m(2) for five consecutive days), PDT (630 nm for 10 min) or a combination of TMZ and PDT. TMZ concentration in glioma tissues was detected using liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS) and cell death was observed using transmission microscopy. Concurrently, another batch of 40 glioma-bearing rats was subjected to the same treatment, and the survival of these rats was estimated using Kaplan-Meier analysis. PDT significantly decreased the expression of P-gp in endothelial cells comprising the blood-tumor-barrier and in glioma tissues. The combination of TMZ with PDT significantly increased TMZ concentration in glioma tissues, enhanced glioma cell apoptosis and prolonged the median survival of glioma-bearing rats. The combination of PDT with TMZ shows synergistic effect in rat C6 glioma model, indicating its potential clinical use in glioma treatment.

Thrombose veineuse et cancer

For glioma, temozolomide (TMZ) is a commonly used chemotherapy drug and photodynamic therapy (PDT) is an important adjuvant therapy. The aim of this study was to evaluate the effect of their combination for the treatment of glioma.A rat C6 glioma model using male Wistar rats (n = 180) weighing 280–300 g was established. Glioma-bearing rats (n = 100) were treated with mock, hematoporphyrin monomethyl ether (HMME), laser or PDT. The expression of P-glycoprotein (P-gp) in endothelial cells of the blood–tumor-barrier and in glioma tissues was detected using immunohistochemistry and western blot, respectively. Glioma-bearing rats (n = 40) were treated with normal saline, TMZ (60 mg/m2 for five consecutive days), PDT (630 nm for 10 min) or a combination of TMZ and PDT. TMZ concentration in glioma tissues was detected using liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS) and cell death was observed using transmission microscopy. Concurrently, another batch of 40 glioma-bearing rats was subjected to the same treatment, and the survival of these rats was estimated using Kaplan–Meier analysis.PDT significantly decreased the expression of P-gp in endothelial cells comprising the blood–tumor-barrier and in glioma tissues. The combination of TMZ with PDT significantly increased TMZ concentration in glioma tissues, enhanced glioma cell apoptosis and prolonged the median survival of glioma-bearing rats.The combination of PDT with TMZ shows synergistic effect in rat C6 glioma model, indicating its potential clinical use in glioma treatment.

O7.06 * MAGNETIC NANOPARTICLES CONJUGATED TO A RECOMBINANT HEAT SHOCK PROTEIN HSP70 FOR TARGETING EXPERIMENTAL GLIOBLASTOMA

Superparamagnetic nanoparticles (SPIONs) due to their unique properties could be applied for theranostic approach in neuro-oncology. Accumulation and retention of the nanoparticles in the tumour can be directly imaged by MRI proving its application as contrast agents. Due to high heating capacity SPIONs could also be applied for thermotherapy of brain tumours. To further increase the biocompatibility and targeted delivery of SPIONs we have functionalized the surface of particles with recombinant heat shock protein Hsp70. Nuclear magnetic resonance (NMR) spectra and magnetic relaxation times of the synthesized conjugates T1, T2, T2* were measured with the help of an NMR spectrometer (CXP-300, Bruker) in a magnetic field of 7.1 T. Hsp70-SPIONs relaxivity corresponded to the properties of negative contrast agents with a hypointensive change of resonance signal in MR imaging. In series of in vitro experiments we observed enhanced internalization of the SPION-Hsp70 conjugates by C6 glioma cells in comparison to control SPIONs. The contrast-enhancing ability of SPION-Hsp70 conjugates was analyzed in the model of intracranial C6 glioma in rodents. The rat images were acquired with the help of a Bruker Avance II NMR spectrometer equipped with a microtomographic accessory at a magnetic field of 11 T. In vivo analysis of the conjugates distribution in the model of intracranial glioma revealed the retention of the functionilzed particles inside the tumour site. Negative contrast tumor enhancement in the T2-weighted MR images was higher in the case of Hsp70-SPIONs in comparison to non-modified SPIONs. Histological analysis of the brain sections confirmed the retention of the Hsp70-SPIONs in the glioma tumor but not in the adjacent normal brain tissues. Immunofluoresence studies confirmed that retention in the glioma of nanoparticles was due to the receptor-mediated endocytosis of the SPION-Hsp70 conjugates via the CD40 receptors. The uptake of SPION-Hsp70 by tumour cells permitted MRI contrast enhancement after systemic delivery while sparring surrounding normal brain tissues.

Targeted delivery of cisplatin by сonnexin 43 vector nanogels to the focus of experimental glioma C6.

The aim of this study was to create a nanocontainer conjugated with monoclonal antibodies to connexin 43 (Cx43) that is actively expressed at the periphery of C6 glioma and in the astroglia roll zone. Stable vector nanogels with high (up to 35%) cisplatin load were synthesized. The antitumor effects of Cx43-modified cisplatin-loaded nanogels, free cisplatin, and nonspecific drugs were carried out on C6 glioma model. Vector nanogels reduced systemic toxicity of cisplatin, effectively inhibited tumor growth, and significantly prolonged the lifespan of animals with experimental tumors.

OP0019 Immunomodulatory potency of recombinant heat shock protein HSP70 in therapy of malignant brain tumours

<h3>Background</h3> HSP70 is known to stimulate anti-tumour immunity, validating its possible application in immunotherapeutic approaches. In a series of in vitro experiments we demonstrated that HSP70, when added to cancer cells, can penetrate the latter inducing the translocation of endogenous HSP70 to plasma membrane. Interchange of exogenous and intracellular HSP70 induced lymphocyte-mediated anti-tumour activity. In an intracranial C6 glioma model, we administered HSP70 intratumourally. The infusion of the chaperone caused inhibition of tumour growth, as measured by high-field (7.1T) MRI, and significantly prolonged survival. These changes correlated with induction of the tumour-specific innate response mediated by NK cells (production of Granzyme B) and adaptive immune response generated by CD8+ T-lymphocytes (proved by IFN<i>γ</i> ELISPOT assay). Immunological changes were accompanied by glioma infiltration by NK-cells (Ly-6c+) and T-cells (i.e., CD3+, CD4+, CD8+). Following the in vivo experiments, the effects of intratumoural injection of HSP70 were explored in a pilot study for feasibility, safety, and possible systemic anti-tumour immune responses in patients with brain tumours. <h3>Methods</h3> Patients with untreated newly diagnosed tumours (<i>n</i>=12) were enrolled in the study. After tumour resection, five injections of HSP70 were performed into the residual tumour cavity through a catheter. Specific immune responses were evaluated with a delayed hypersensitivity test. <h3>Findings</h3> Intratumoural injections of HSP70 were well tolerated by patients. One patient had a complete clinical response documented by radiological findings and one patient had a partial response. In three patients we observed positive delayed hypersensitivity tests. In peripheral blood there was a shift toward cytokines of the Th1-cell mediated response that corresponded to changes in lymphocyte subpopulations. <h3>Interpretation</h3> The present pilot study shows the feasibility and safety of intratumoural delivery of recombinant HSP70 in patients. On the basis of our studies we propose a novel mechanism of immunomodulatory activity of HSP70.

Rapid-Steady-State-T1 signal modeling during contrast agent extravasation: toward tumor blood volume quantification without requiring the arterial input function.

This study demonstrates how to quantify the tumor blood volume fraction (BVf) using the dynamic Rapid-Steady-State-T1 (RSST1 )-MRI method despite contrast agent (CA) leakage and without arterial input function (AIF) determination. For vasculature impermeable to CAs, the BVf is directly quantified from the RSST1 signal amplitude. In case of CA extravasation, we propose a two-compartment model to describe the dynamic RSST1 signal increase. We applied the mathematical model in a pilot-study on a RG2-glioma model to compare extravasation of two Gd-based CAs. The BVf quantification using the mathematical model in a C6-glioma model (n = 8) with the clinical CA Gd-DOTA was validated using a ΔR2 *-steady-state MRI method with an USPIO and by immunohistochemical staining of perfused vessels labeled with Hoechst-33342 dye in the same rats. BVf in tumor and in healthy brain tissues (0.034 ± 0.005 and 0.026 ± 0.004, respectively) derived from the dynamic RSST1 signal were confirmed by ΔR2 *-steady-state MRI (0.036 ± 0.003 and 0.027 ± 0.002, respectively, correlation coefficient rS = 0.74) and by histology (0.036 ± 0.003 and 0.025 ± 0.004 respectively, rS = 0.87). Straightforward tumor BVf quantification without AIF determination is demonstrated in presence of CA leakage. The method will facilitate angiogenesis assessment in longitudinal neuro-oncologic studies in particular when monitoring the response to antiangiogenic therapies.

Magnetic Resonance Imaging of Rat C6 Glioma Model Enhanced by Using Water-Soluble Gadolinium Fullerene

Water-soluble gadofullerene Gd@Ful with composition Gd3+@C82(OH)X3– (x ~ 20) was synthesized for theranostic study. Nanosuspensions of Gd@Ful were used for magnetic relaxation measurements in vitro and for magnetic resonance imaging of a rat with intracranially implanted C6 glioma. Gd@Ful was shown to reduce proton relaxation times in vitro and provide dual contrast of T1- and T2-weighted images in a rat brain tumor model after paramagnetic intravenous delivery. Magnetic relaxation times of water protons under action of Gd@Ful were strongly shortened due to cluster formation and increase of motional correlation times of protons in the vicinity of the fullerene cage. The Gd@Ful administration promoted the improvement of glioma contrast enhancement at T2-weighted images due to accumulation of paramagnetic substance at the tumor site. The contrast efficiency of Gd@Ful corresponds to the characteristics of negative contrast agent. The Gd@Ful nanosuspension is shown to be a contrast enhancer with high anti-tumor therapeutic potency. The retention of the Gd@Ful in the tumor resulted in a 75 % increase in survival times of the tumor-bearing animals.

Permeability of the blood–tumor barrier is enhanced by combining vascular endothelial growth factor with papaverine

This study aims to determine the effects of vascular endothelial growth factor (VEGF), papaverine (PA), and the combination of VEGF and PA on the permeability of the blood-tumor barrier (BTB) and to determine possible molecular mechanisms contributing to the effects. In the rat C6 glioma model, the extravasation of Evans blue (EB) through the BTB was increased significantly by VEGF and PA. VEGF-induced and PA-induced increase of EB extravasation was further increased after combining VEGF with PA infusion. Transmission electron microscopy (TEM) showed that the combination of VEGF and PA not only opened tight junctions (TJ) dramatically but increased the presence of pinocytotic vesicles of brain microvascular endothelial cells (BMECs) significantly. Meanwhile, the downregulation of the TJ-associated proteins occludin and claudin-5 and the upregulation of the caveolae structure proteins caveolin-1 and caveolin-2 caused by the combination of VEGF and PA were observed by Western blot and immunohistochemistry, which were more remarkable than those by the two strategies separately. In addition, after VEGF and PA infusion, the results of radioimmunoassay, Western blot, and enzyme-linked immunosorbent assay (ELISA) revealed a significant increase in expression levels of cGMP and protein kinase G-1 (PKG-1) and the activation of nuclear factor-κB (NF-κB) p65. This study demonstrates that combination of VEGF and PA can increase the permeability of the BTB by a paracellular pathway (downregulation of occludin and claudin-5) and a transcellular pathway (upregulation of caveolin-1 and caveolin-2) and that the cGMP/PKG/NF-κB signal pathway might be involved in the modulation process.

HPPH光动力学治疗大鼠C6脑胶质瘤肿瘤生长实验研究的磁共振表现

HPPH光动力学治疗大鼠C6脑胶质瘤肿瘤生长实验研究的磁共振表现

HPPH光动力学治疗大鼠C6脑胶质瘤肿瘤生长实验研究的磁共振表现

HPPH光动力学治疗大鼠C6脑胶质瘤肿瘤生长实验研究的磁共振表现

大鼠C6脑胶质细胞移植瘤肿瘤模型动态生长核磁共振表现

大鼠C6脑胶质细胞移植瘤肿瘤模型动态生长核磁共振表现

大鼠C6脑胶质细胞移植瘤肿瘤模型动态生长核磁共振表现

大鼠C6脑胶质细胞移植瘤肿瘤模型动态生长核磁共振表现

Concomitant treatment with pertussis toxin plus temozolomide increases the survival of rats bearing intracerebral RG2 glioma

Glioblastoma multiforme is the most frequent primary brain tumor, it has poor prognosis, and it remains refractory to current treatment. The success of temozolomide (TMZ) appears to be limited by the occurrence of chemoresistance. Recently, we report the use of pertussis toxin as adjuvant immunotherapy in a C6 glioma model; showing a decrease in tumoral size, it induced selective cell death in Treg cells, and it elicited less infiltration of tumoral macrophages. Here, we evaluated the cytotoxic effect of pertussis toxin in combination with TMZ for glioma treatment, both in vitro and in vivo RG2 glioma model. We determined cell viability, cell cycle, apoptosis, and autophagy on treated RG2 cells through flow cytometry, immunofluorescence, and Western blot assays. Twenty-eight rats were divided in four groups (n = 7) for each treatment. After intracranial implantation of RG2 cells, animals were treated with TMZ (10 mg/Kg/200 μl of apple juice), PTx (2 μg/200 μl of saline solution), and TMZ + PTx. Animals without treatment were considered as control. We found an induction of apoptosis in around 20 % of RG2 cells, in both single treatments and in their combination. Also, we determined the presence of autophagy vesicles, without any modifications in the cell cycle in the TMZ - PTx-treated groups. The survival analyses showed an increase due to individual treatments; while in the group treated with the combination TMZ - PTx, this effect was enhanced. We show that the concomitant use of pertussis toxin plus TMZ could represent an advantage to improve the glioma treatment.

Tumor targeting using magnetic nanoparticle Hsp70 conjugate in a model of C6 glioma

Superparamagnetic iron oxide nanoparticles (SPIONs), due to their unique magnetic properties, have the ability to function both as magnetic resonance (MR) contrast agents, and can be used for thermotherapy. SPIONs conjugated to the heat shock protein Hsp70 that selectively binds to the CD40 receptor present on glioma cells, could be used for MR contrast enhancement of experimental C6 glioma. The magnetic properties of the Hsp70-SPIONs were measured by NMR relaxometry method. The uptake of nanoparticles was assessed on the C6 glioma cells by confocal and electron microscopes. The tumor selectivity of Hsp70-SPIONs being intravenously administered was analyzed in the experimental model of C6 glioma in the MRI scanner. Hsp70-SPIONs relaxivity corresponded to the properties of negative contrast agents with a hypointensive change of resonance signal in MR imaging. A significant accumulation of the Hsp70-SPIONs but not the non-conjugated nanoparticles was observed by confocal microscopy within C6 cells. Negative contrast tumor enhancement in the T2-weighted MR images was higher in the case of Hsp70-SPIONs in comparison to non-modified SPIONs. Histological analysis of the brain sections confirmed the retention of the Hsp70-SPIONs in the glioma tumor but not in the adjacent normal brain tissues. The study demonstrated that Hsp70-SPION conjugate intravenously administered in C6 glioma model accumulated in the tumors and enhanced the contrast of their MR images.

Low-dose Endothelial Monocyte-activating Polypeptide-II Increases Permeability of Blood–tumor Barrier by Caveolae-mediated Transcellular Pathway

Low-dose endothelial monocyte-activating polypeptide-II (EMAP-II) can selectively increase blood-tumor barrier (BTB) permeability via the paracellular pathway. The role of the transcellular pathway in this process is unclear. This study was conducted to evaluate the potential involvement of the transcellular pathway in EMAP-II-induced opening of the BTB and to identify the associated mechanisms. Evans blue extravasation test was used to measure changes in BTB permeability after EMAP-II (80ng/kg) administration in a rat model of C6 glioma. Changes in the quantity of pinocytotic vesicles in rat brain microvascular endothelial cells (BMECs) were observed using transmission electron microscopy. Reverse transcription-polymerase chain reaction, Western blotting, and immunohistochemistry assays were performed to detect the expression of the caveolar structural proteins, caveolin-1 and caveolin-2, in BMECs. Alterations in the expression of phospho (p)-Src, p-caveolin-1, and p-caveolin-2 and the activity of RhoA also were measured. Effects of tyrosine kinase inhibition on EMAP-II-induced RhoA/Rho kinase activations and tyrosine kinase, RhoA, or Rho kinase inhibition on EMAP-II-induced caveolin-1 and caveolin-2 phosphorylation were determined by inhibition studies. One hour after EMAP-II administration, the quantity of pinocytotic vesicles in BMECs increased markedly, consistent with changes in BTB permeability. The expression levels of caveolin-1, caveolin-2, p-caveolin-1, and p-caveolin-2 in BMECs also were significantly increased at 1h. The peak expression level of p-Src and the peak activity of RhoA occurred at 0.25 and 0.5h, respectively. Inhibition of tyrosine kinase significantly diminished the activities of RhoA and Rho kinase induced by EMAP-II. In addition, EMAP-II-induced phosphorylation of caveolin-1 and caveolin-2 was completely blocked by inhibition of tyrosine kinase, RhoA, or Rho kinase. We suggest that low-dose EMAP-II can induce BTB hyperpermeability via the transcellular pathway, which is associated with phosphorylation and upregulation of caveolin-1 and caveolin-2 and involves the tyrosine kinase/RhoA/Rho kinase signaling pathway.

Transferrin modified PEG-PLA-resveratrol conjugates: In vitro and in vivo studies for glioma

Glioblastoma is one of the most malignant brain tumors with a poor prognosis. In this study, we examined the effects of transferrin (Tf)-modified poly ethyleneglycol-poly lactic acid (PEG-PLA) nanoparticles conjugated with resveratrol (Tf-PEG-PLA-RSV) to glioma therapy in vitro and in vivo. The cell viability of Tf-PEG-PLA-RSV on C6 and U87 glioma cells was determined by the MTT assay. In vivo biodistribution and antitumor activity were investigated in Brain glioma bearing rat model of C6 glioma by i.p. administration of RSV-polymer conjugates. We found that the average diameter of each Tf-PEG-PLA-RSV is around 150nm with 32 molecules of Tf on surface. In vitro cytotoxicity of PEG-PLA-RSV against C6 and U87 cells was higher than that of free RSV, and further the modification of Tf enhanced the cytotoxicity of the RSV-polymer conjugates as a result of the increased cellular uptake of the RSV-modified conjugates by glioma cells. In comparison with free RSV, RSV conjugates could significantly decrease tumor volume and accumulate in brain tumor, which resulted in prolonging the survival of C6 glioma-bearing rats. These results suggest that Tf-NP-RSV had a potential of therapeutic effect to glioma both in vitro and in vivo and might be a potential candidate for targeted therapy of glioma and worthy of further investigation.

Lentivirus-mediated CD/TK fusion gene transfection neural stem cell therapy for C6 glioblastoma

A suicide gene can convert nontoxic prodrugs into toxic products to kill tumor cells. In this study, our aim was to transfect lentivirus-mediated CD/TK fusion gene into Wistar rat's neural stem cells (NSC) and then implant the NSC into a C6 glioma model to observe a C6 glioma growth inhibition effect. Primary NSC and stable transfection CD/TK fusion gene cell lines were established. To observe the tumor size and rat survival period in different groups, C6 glioma cell apoptosis and cell viability rate were applied to analyze the tumor inhibition effect of the neural stem cells' transfected CD/TK fusion gene. C6 cell viability showed that CDglyTK-NSC + GCV/5-Fc (group 1) was lower than CDglyTK-NSC (group 2), NSC + GCV/5-Fc (group 3), and control (group 4) from day 2 (p < 0.05), and the apoptosis rate was higher in group 1 compared with that of other groups (50.6%, p < 0.05) either in vitro or in vivo (35.47%, p < 0.05); both cell viability and apoptosis had no significance in the other three groups. In vivo, tumor size in group 1 was 7.76 ± 1.37 mm(3), which is smaller than the others (group2 27.28 ± 4.11 mm(3), group3 27.94 ± 2.08 and 28.61 ± 2.97 mm(3); p < 0.05). The other groups' tumor size was not significant (p > 0.05). Survival time of rats treated with CDglyTK-NSC + GCV/5-Fc (group 1) was significantly longer than that of the other groups (p < 0.05; group 1 48.86 ± 1.97, group 2 28.67 ± 3.75, group 3 31.5 ± 1.27, group 4 29.3 ± 1.33). We also showed that the transfected C6 cells had a migratory capacity toward gliomas in vivo. Transfected CD/TK fusion gene neural stem cells combined with propyl-guanosine and 5-flucytosine double prodrug significantly inhibit the development of glioma.

The prospective application of a hypoxic radiosensitizer, doranidazole to rat intracranial glioblastoma with blood brain barrier disruption

BackgroundGlioblastoma is one of the intractable cancers and is highly resistant to ionizing radiation. This radioresistance is partly due to the presence of a hypoxic region which is widely found in advanced malignant gliomas. In the present study, we evaluated the effectiveness of the hypoxic cell sensitizer doranidazole (PR-350) using the C6 rat glioblastoma model, focusing on the status of blood brain barrier (BBB).MethodsReproductive cell death in the rat C6 glioma cell line was determined by means of clonogenic assay. An intracranial C6 glioma model was established for the in vivo experiments. To investigate the status of the BBB in C6 glioma bearing brain, we performed the Evans blue extravasation test. Autoradiography with [14C]-doranidazole was performed to examine the distribution of doranidazole in the glioma tumor. T2-weighted MRI was employed to examine the effects of X-irradiation and/or doranidazole on tumor growth.ResultsDoranidazole significantly enhanced radiation-induced reproductive cell death in vitro under hypoxia, but not under normoxia. The BBB in C6-bearing brain was completely disrupted and [14C]-doranidazole specifically penetrated the tumor regions. Combined treatment with X-irradiation and doranidazole significantly inhibited the growth of C6 gliomas.ConclusionsOur results revealed that BBB disruption in glioma enables BBB-impermeable radiosensitizers to penetrate and distribute in the target region. This study is the first to propose that in malignant glioma the administration of hydrophilic hypoxic radiosensitizers could be a potent strategy for improving the clinical outcome of radiotherapy without side effects.