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Purpose: Our previous study revealed that the application of a magnetic field longitudinal to a carbon-ion beam of 0.1 ≤  B // ≤ 0.6 T enhances the biological effectiveness of the radiation. The purpose of this study is to experimentally verify whether the application of a magnetic field perpendicular to the beam also alters the biological effectiveness. Methods and materials: Most experimental conditions other than the magnetic field direction were the same as those used in the previous study to allow comparison of their results. Human cancer and normal cells were exposed to low (12 keV/μm) and high (50 keV/μm) linear energy transfer (LET) carbon-ion beams under the perpendicular magnetic fields of B ⊥ = 0, 0.15, 0.3, or 0.6 T generated by a dipole magnet. The effects of the magnetic fields on the biological effectiveness were evaluated by clonogenic cell survival. Doses that would result in the survival of 10%, D 10 s, were determined for the exposures and analyzed using Student's t -tests. Results: For both cancer and normal cells treated by low- and high-LET carbon-ion beams, the D 10 s measured in the presence of the perpendicular magnetic fields of B ⊥ ≥ 0.15 T were not statistically different ( p  ≫ .05) from the D 10 s measured in the absence of the magnetic fields, B ⊥ = 0 T. Conclusions: Exposure of human cancer and normal cells to the perpendicular magnetic fields of B ⊥ ≤ 0.6 T did not alter significantly the biological effectiveness of the carbon-ion beams, unlike the exposure to longitudinal magnetic fields of the same strength. Although the mechanisms underlying the observed results still require further exploration, these findings indicate that the influence of the magnetic field on biological effectiveness of the carbon-ion beam depends on the applied field direction with respect to the beam.

Despite continuous improvements in treatment of glioblastoma, tumor recurrence and therapy resistance still occur in a high proportion of patients. One underlying reason for this radioresistance might be the presence of glioblastoma cancer stem cells (GSCs), which feature high DNA repair capability. PARP protein plays an important cellular role by detecting the presence of damaged DNA and then activating signaling pathways that promote appropriate cellular responses. Thus, PARP inhibitors (PARPi) have recently emerged as potential radiosensitizing agents. In this study, we investigated the preclinical efficacy of talazoparib, a new PARPi, in association with low and high linear energy transfer (LET) irradiation in two GSC cell lines. Reduction of GSC fraction, impact on cell proliferation, and cell cycle arrest were evaluated for each condition. All combinations were compared with a reference schedule: photonic irradiation combined with temozolomide. The use of PARPi combined with photon beam and even more carbon beam irradiation drastically reduced the GSC frequency of GBM cell lines in vitro. Furthermore, talazoparib combined with irradiation induced a marked and prolonged G2/M block, and decreased proliferation. These results show that talazoparib is a new candidate that effects radiosensitization in radioresistant GSCs, and its combination with high LET irradiation, is promising.

The purpose of this study was to investigate the efficacy of metformin as a radiosensitizer for use in combination therapy for human hepatocellular carcinoma (HCC). Three human HCC cell lines (Huh7, HepG2, Hep3B) and a normal human hepatocyte cell line were treated with metformin alone or with radiation followed by metformin. In vitro tests were evaluated by clonogenic survival assay, FACS analysis, western blotting, immunofluorescence and comet assay. Metformin significantly enhanced radiation efficacy under high and low Linear Energy Transfer (LET) radiation conditions in vitro. In combination with radiation, metformin abrogated G2/M arrest and increased the cell population in the sub-G1 phase and the ROS level, ultimately increasing HCC cellular apoptosis. Metformin inhibits the repair of DNA damage caused by radiation. The radiosensitizing effects of metformin are much higher in neutron (high LET)-irradiated cell lines than in γ (low LET)-irradiated cell lines. Metformin only had a moderate effect in normal hepatocytes. Metformin enhances the radiosensitivity of HCC, suggesting it may have clinical utility in combination cancer treatment with high-LET radiation.

Purpose: To investigate the mechanisms regulating the pathways of the bystander transmission in vitro, focusing on the radiation-perturbed signalling (via Interleukine 6, IL-6) of the irradiated cells after exposure to low doses of different radiation types., Materials and Methods: An integrated 'systems radiation biology' approach was adopted. Experimentally the level of the secreted cytokine from human fibroblasts was detected with ELISA (Enzyme-Linked ImmunoSorbent Assay) method and subsequently the data were analyzed and coupled with a phenomenological model based on differential equations to evaluate the single-cell release mechanisms., Results: The data confirmed the important effect of radiation on the IL-6 pathway, clearly showing a crucial role of the ROS (Reactive Oxygen Species) in transducing the effect of initial radiation exposure and the subsequent long-term release of IL-6. Furthermore, a systematic investigation of radiation dose/radiation quality dependence seems to indicate an increasing efficiency of high LET (Linear Energy Transfer) irradiation in the release of the cytokine. Basic hypotheses were tested, on the correlation between direct radiobiological damage and signal release and on the radiation target for this endpoint (secretion of IL-6)., Conclusions: The results demonstrate the role of reactive oxygen and nitrogen species in the signaling pathways of IL-6. Furthermore the systems radiation biology approach here adopted, allowed us to test and verify hypotheses on the behavior of the single cell in the release of cytokine, after the exposure to different doses and different qualities of ionizing radiation.

Purpose: We recently reported that curcumin attenuates ionizing radiation (IR)-induced survival signaling and proliferation in human neuroblastoma cells. Also, in the endothelial system, we have demonstrated that NFκB regulates IR-induced telomerase activity (TA). Accordingly, we investigated the effect of curcumin in inhibiting IR-induced NFκB-dependent hTERT transcription, TA, and cell survival in neuroblastoma cells., Methods and Materials: SK-N-MC or SH-SY5Y cells exposed to IR and treated with curcumin (10-100 nM) with or without IR were harvested after 1 h through 24 h. NFκB-dependent regulation was investigated either by luciferase reporter assays using pNFκB-, pGL3-354-, pGL3-347-, or pUSE-IκBα-Luc, p50/p65, or RelA siRNA-transfected cells. NFκB activity was analyzed using an electrophoretic mobility shift assay and hTERT expression using the quantitative polymerase chain reaction. TA was determined using the telomerase repeat amplification protocol assay and cell survival using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltertrazolium bromide and clonogenic assay., Results: Curcumin profoundly inhibited IR-induced NFκB. Consequently, curcumin significantly inhibited IR-induced TA and hTERT mRNA at all points investigated. Furthermore, IR-induced TA is regulated at the transcriptional level by triggering telomerase reverse transcriptase (TERT) promoter activation. Moreover, NFκB becomes functionally activated after IR and mediates TA upregulation by binding to the κB-binding region in the promoter region of the TERT gene. Consistently, elimination of the NFκB-recognition site on the telomerase promoter or inhibition of NFκB by the IκBα mutant compromises IR-induced telomerase promoter activation. Significantly, curcumin inhibited IR-induced TERT transcription. Consequently, curcumin inhibited hTERT mRNA and TA in NFκB overexpressed cells. Furthermore, curcumin enhanced the IR-induced inhibition of cell survival., Conclusions: These results strongly suggest that curcumin inhibits IR-induced TA in an NFκB dependent manner in human neuroblastoma cells., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Cerebellum is a vital organ responsible for the motor coordination and recently it has been reported to be involved in cognitive function. Reactive oxygen species are implicated in neurodegeneration and cognitive disorders because of higher vulnerability of neuronal tissues. Therefore, the present study aimed at investigating the role of melatonin against high-LET (linear energy transfer) (56)Fe particle irradiation-induced oxidative damage and apoptosis in the mouse cerebellum. Radiation-induced oxidative damage was examined using a neuronal-specific terminal deoxynucleotidyl transferase-mediated nick end-labeling (TUNEL), quantitative histopathology, DNA damage (comet assay), carbonyl content and 4-HAE + MDA (4-hydroxyalkenal + malondialdehyde) status of the cerebellum. Radiation exposure augmented the number of TUNEL positive cell, DNA migration in the comet tail and carbonyl and 4-HAE + MDA level in the cerebellum. Melatonin pretreatment significantly inhibited the oxidative damage to biomolecules as well as cerebellar apoptosis. Melatonin-treated irradiated mice showed higher counts of intact Purkinje cells as compared to vehicle-treated irradiated mice. In addition, radiation induced augmentation of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and a decline in the total antioxidant capacity in serum; these changes were also ameliorated by melatonin pretreatment. The present results provide evidence supporting the antioxidant and neuroprotective function of melatonin.

Purpose: Recent studies from this laboratory with (212)Pb-trastuzumab have shown the feasibility of targeted therapy for the treatment of disseminated peritoneal disease using (212)Pb as an in vivo generator of (212)Bi. The objective of the studies presented here was improvement of the efficacy of alpha-particle radioimmunotherapy using a chemotherapeutic agent., Experimental Design: In a series of experiments, a treatment regimen was systematically developed in which athymic mice bearing i.p. LS-174T xenografts were injected i.p. with gemcitabine at 50 mg/kg followed by (212)Pb radioimmunotherapy., Results: In a pilot study, tumor-bearing mice were treated with gemcitabine and, 24 to 30 h later, with 5 or 10 muCi (212)Pb-trastuzumab. Improvement in median survival was observed at 5 microCi (212)Pb-trastuzumab in the absence (31 days) or presence (51 days) of gemcitabine: 45 and 70 days with 10 microCi versus 16 days for untreated mice (P < 0.001). Multiple doses of gemcitabine combined with a single (212)Pb radioimmunotherapy (10 microCi) administration was then evaluated. Mice received three doses of gemcitabine: one before (212)Pb-trastuzumab and two afterwards. Median survival of mice was 63 versus 54 days for those receiving a single gemcitabine dose before radioimmunotherapy (P < 0.001), specifically attributable to (212)Pb-trastuzumab (P = 0.01). Extending these findings, one versus two treatment cycles was compared. A cycle consisted of sequential treatment with gemcitabine, 10 microCi (212)Pb radioimmunotherapy, then one or two additional gemcitabine doses. In the first cycle, three doses of gemcitabine resulted in a median survival of 90 versus 21 days for the untreated mice. The greatest benefit was noted after cycle 2 in the mice receiving 10 microCi (212)Pb-trastuzumab and two doses of gemcitabine with a median survival of 196.5 days (P = 0.005). Pretreatment of tumor-bearing mice with two doses of gemcitabine before (212)Pb radioimmunotherapy was also assessed with gemcitabine injected 72 and 24 h before (212)Pb-trastuzumab. The median survival was 56 and 76 days with one and two doses of gemcitabine versus 49 days without gemcitabine. The effect may not be wholly specific to trastuzumab because (212)Pb-HuIgG with two doses of gemcitabine resulted in a median survival of 66 days (34 days without gemcitabine)., Conclusions: Treatment regimens combining chemotherapeutics with high-LET targeted therapy may have tremendous potential in the management and care of cancer patients.

In order to obtain a deeper insight into the molecular mechanism controlling the cellular response to high-linear energy transfer (LET) radiation, the number and size of pATM (S1981) and gamma-H2AX foci were compared in cultures of diploid human fibroblasts after exposure to charged particles of varying species, energy and LET at the NIRS-HIMAC-facility (Chiba, Japan). Particle LET ranged from 2.2 to 300 keV/mum, and a low fluence of 7.3 x 10(4) cm(-2) was chosen. Therefore, about 1 out of 7 nuclei was traversed by a particle. Doses and LET were verified with thermoluminescence detectors (LiF:Mg, Ti) evaluated according to the high temperature ratio method. Two hours after irradiation, fibroblasts were fixed and the subcellular distribution of pATM (S1981) and gamma-H2AX was visualised by immunofluorescence or histochemical staining using phosphorylation-specific antibodies. It was found that the number of pATM (S1981) foci per nucleus was higher after exposure to higher-LET particles. Irradiation with the two highest LET beams (Fe-ions, 197 and 300 keV/mum) gave a significant increase in the number of pATM foci, whereas ions with an LET lower than 30 keV/mum yielded similar numbers of pATM foci compared with unirradiated control samples. These data show that the early cellular response to high-LET radiation is modulated by the energy deposition of the particle. Therefore, the correlation between the microdosimetric aspect of energy deposition and biologic consequences at low radiation doses deserves further study.

Undifferentiated thyroid carcinoma (UTC) is an aggressive tumor with a poor prognosis due to the lack of an effective treatment. Boron neutron capture therapy (BNCT) is based on the selective uptake of boron by the tumor and its activation by a neutron beam, releasing lithium-7 and an alpha particle that will kill the tumor cells by their high linear energy transfer (LET). In previous studies we have shown a selective uptake of borophenylalanine (10BPA) in a human UTC cell line (ARO) and in NIH nude mice implanted with this cell line. When these animals were injected with BPA and irradiated with an appropriated neutron beam, we observed a 100% of tumor growth control and a 50% of histological cure when the initial tumor volume was 50 mm3 or less. Further studies with BOPP (tetrakis-carborane carboxylate ester of 2,4-bis-(alpha, beta-dihydroxyethyl)-deutero-porphyrin IX) showed that when this porphyrin was injected 5-7 days before BPA, and the animals were sacrificed 60 min after the i.p. injection of BPA, a significant increase in boron uptake by the tumor was found (45-38 ppm with both compounds vs. 20 ppm with BPA alone). The application of BNCT using the combination of boron compounds showed a 100% of complete remission in tumors with initial volumes under 50 mm3. Dogs suffer spontaneous UTC, with a similar biological behavior to the human tumor, and a selective uptake of BPA. These results open the possibility of applying BNCT to UTC.

Objective: To compare the anesthetic efficacy of EMLA cream (eutectic mixture of local anesthetics) with that of LET solution (lidocaine, epinephrine, tetracaine) for pretreating lacerations prior to lidocaine injection., Methods: This was a randomized, double-blind clinical trial in a convenience sample of 60 patients aged 1 to 59 years with traumatic lacerations. Eligible wounds were uncomplicated, clean lacerations < or = 6 hours old. Finger and toe lacerations were excluded. At the time of initial presentation to triage, patients were randomized to LET or EMLA. A nurse applied the topical anesthetic into the laceration with a 5-mL syringe. A physician assessed the laceration edges for the presence of blanching and adequacy of anesthesia to a 27-gauge needlestick. Supplemental lidocaine was then infiltrated through the wound edges and the pain of infiltration was recorded by the patient (or guardian) on a 100-mm visual analog scale marked "most pain" at the high end. A sample of 44 patients had 90% power to detect a 20-mm difference in injection pain (two-tailed alpha = 0.05)., Results: Sixty patients were randomized to LET (29) or EMLA (31). Median age was 8.5 years; 23% were female. Most lacerations were facial and closed with sutures. There was no difference in baseline characteristics between groups. More wounds treated with LET were anesthetic to a needlestick than wounds treated with EMLA (73% vs 40%, p = 0.01); however, there was no between-group difference in the median pain of lidocaine infiltration (LET-12 mm vs EMLA-13 mm, p = 0.89)., Conclusions: Pretreatment of simple lacerations with LET or EMLA at the time of patient presentation results in similar amounts of pain of subsequent local infiltration of lidocaine

The search for effective radioprotectors is of major concern in the medical, military, environmental, and space sciences. Conventional radioprotectors are generally effective only during a single irradiation and display their radioprotective properties only at high, toxic concentrations. In addition, they reduce somatic radiation effects but are poorly efficient in protecting from hereditary stochastic radiation effects. In this respect, the pigment melanin merits attention. Experiments referring to potential melanin effects on the ionising radiation response have been carried out with different biological systems, both in vivo and in vitro. In this paper, we present results on the response to high- and low-linear energy transfer (LET) radiation of a human mammary epithelial cell line, H184B5 F5-1 M/10, supplemented by melanin. The incorporation of auto-oxidative (L-dopa) melanin was linear for concentrations from 3 to 10 micrograms/ml in the growth medium. Concentrations of up to 250 micrograms/ml did not significantly impair the cells proliferative ability. No significant protective effect of melanin on the survival of cultured cells after exposure to alpha-particles (130 keV/micron) or x-rays was observed.