[Stereotactic body radiation therapy for hepatocellular carcinoma: Results from a retrospective multicentre study]. / Radiothérapie en conditions stéréotaxiques du carcinome hépatocellulaire : résultats d'une étude rétrospective multicentrique.

PURPOSE: The purpose of this paper was to describe local control, overall survival, progression-free survival and toxicity of CyberKnife®-based stereotactic body radiation therapy of hepatocellular carcinoma. MATERIAL AND METHODS: Records of all the patients treated for hepatocellular carcinoma at the Eugene-Marquis cancer centre, Rennes and the Bretonneau hospital, Tours (France), between November 2010 and December 2016, were reviewed. Radiation therapy was performed as a salvage treatment, while awaiting liver transplantation or if no other treatment was possible. RESULTS: One hundred and thirty-six patients were consecutively included in the study. The median follow-up was 13months. Median total dose prescribed, fractionation and overall treatment time were respectively 45Gy, three fractions and 5 days. Overall survival, progression-free survival and local control rates at 1year and 2years were 79.8 % and 63.5 %, 61.3 % and 39.4 %; 94.5 % and 91 %. Two grade 3 acute toxicity events and two grade 4 late toxicity events corresponding to a duodenal ulcer have been reported. Seven patients underwent classic radiation-induced hepatitis and 13 patients showed non-classical radiation-induced hepatitis. Barcelona Clinic Liver Cancer stage, World Health Organisation grade and planning target volume were correlated with overall survival in univariate Cox analysis. CONCLUSION: Stereotactic body radiation therapy is effective and well-tolerated for inoperable hepatocellular carcinoma or as a bridge to liver transplantation. Toxicity is mainly related to cirrhotic background and requires a selection of patients and strict dose constraints.

[Documentation management: from theory to practice]. / La gestion documentaire: de la théorie à la pratique.

The management of the documentation is one of the key points regarding the efficacy and the performance of the quality management of health centres. It offers to all professionals the possibility to be informed on the procedures in use, leading to a pool of documents for improvement of organisations and for securing the critical steps of the patient management. In this paper, we will describe the optimal organisation of the documentation according to Haute autorité de santé (HAS) and ISO recommendations, then we will discuss in concrete terms the potential methods usable for the production of a tool well adapted to our routine practice, in order to achieve the objectives for security.

[The French preliminary experience of the use of a seed-projector for exclusive iodine 125 prostate brachytherapy: feasibility and acute toxicity]. / L'expérience française de l'utilisation d'un projecteur de source pour la curiethérapie prostatique exclusive par implants permanents d'iode radioactif: faisabilité et données de toxicité aiguë.

PURPOSE: To analyse a new technique for prostate brachytherapy with permanent Iodine implants characterized by the use of a seed projector after a 3D dosimetric peroperative treatment planning (FIRST technique). PATIENTS AND METHOD: 395 patients have been treated in France with this technique in six radiotherapy centres between November 2002 and December 2005 for a localized prostate cancer. RESULTS: Thirteen patients (3.3%) developped a urinary retention, and respectively 7.8 and 26.5% an acute RTOG grade 3 and 2 toxicity. The 6-weeks IPSS score was equal or lower to 15 in 73% with a 11 median IPSS value. A failure of the loading with the seed-projector, leading to a manual loading of the seeds, occurred in 9 patients (2.3%) in two centres, directly related to the loading procedure with the seed-projector in 5 cases. The median duration of the procedure was reduced by 30 minutes for the patients treated in 2005. CONCLUSIONS: This multicenter study establishes the feasibility of the routine use of a seed projector for permanent iodine 125 prostate implants with an initial tolerance similar to the best results published for other implants techniques.

[Study of a new commercial film for high energy photon dosimetry]. / Etude d'un film récemment commercialisé pour la dosimétrie des faisceaux de photons de haute énergie.

Dosimetric properties of a new film, the Extended Dose Range-EDR2, manufactured by Kodak, have been studied. We have established the response of the film versus dose and compared it with that of X-OMAT V films. We found a linear response with dose, for the range from 0.5 to 4 Gy. No dependence of this curve with beam quality and with depth is observed. EDR2 films are useful for dosimetric study of high-energy photon beam, especially when high dose gradient occurs such as for Intensity Modulated Radiotherapy Treatment.

Ceramide induces activation of the mitochondrial/caspases pathway in Jurkat and SCC61 cells sensitive to gamma-radiation but activation of this sequence is defective in radioresistant SQ20B cells.

PURPOSE: To clarify the molecular mechanisms leading to radiation-induced apoptosis or resistance, the kinetics (1-48 h) and sequence of events triggered in response to 10 Gy irradiation were investigated in three cell lines displaying a gradient of sensitivity to 7-rays. MATERIALS AND METHODS: Ceramide levels were measured by high performance liquid chromatography (HPLC). Mitochondrial function was evaluated in terms of transmembrane potential (delta(psi)m), reactive oxygen species (ROS) and glutathione levels analysed by flow cytometry or HPLC. Caspase activation was assessed by immunoblotting, and apoptosis by flow cytometry. RESULTS: In Jurkat radiosensitive cells and SCC61 adherent cells with intermediate radiosensitivity, the degree of delayed ceramide release was directly related to their propensity to undergo apoptosis. Transduction of the death signal was mediated by a drop in delta(psi)m and glutathione levels, ROS accumulation and activation of effector caspases. Experiments conducted with caspase inhibitors, bongkrekic acid, or DL-PDMP indicated that ceramide triggers mitochondrial collapse, followed by the activation of caspases-9, -8 and -3, and poly(ADP-ribose)polymerase cleavage. In SQ20B radioresistant cells, gamma-radiation did not induce ceramide generation or subsequent activation of the mitochondrial/caspase apoptotic pathway. CONCLUSIONS: Ceramide appears to be a determining factor in the commitment phase of radiation-induced apoptosis. When ceramide is not generated, the whole pathway is ineffective and resistance to apoptosis may result.

[Gross tumor volume and clinical target volume: esophageal tumors]. / Volume tumoral macroscopique et volume-cible anatomoclinique en radiothérapie: tumeurs de l'aesophage.

The oesophagus is divided into four regions: cervical oesophagus, and intrathoracic oesophagus with an upper, mid- and lower thoracic portion. Cancer may occur on each of these regions. Computed tomography of the thorax and superior abdomen and endoscopic ultrasound are necessary for reliable staging. CT simulation allows accurate definition of tumour volume. GTV includes tumour volume and regional lymph nodes. CTV encompasses GTV plus a safety margin and lymph node areas considered to harbour potential microscopic disease. The extent of prophylactic lymph node irradiation depends on the anatomic location of the primary tumour.

Temporal relationships between ceramide production, caspase activation and mitochondrial dysfunction in cell lines with varying sensitivity to anti-Fas-induced apoptosis.

To clarify the chronology of events leading to anti-Fas-induced apoptosis, and the mechanisms of resistance to this death effector, we compared the response kinetics of three tumour cell lines that display varying sensitivity to anti-Fas (based on levels of apoptosis), in terms of ceramide release, mitochondrial function and the caspase-activation pathway. In the highly sensitive Jurkat cell line, early caspase-8 activation, observed from 2 h after treatment, was chronologically associated with an acute depletion of glutathione and the cleavage of caspase-3 and poly-ADP ribosyl polymerase (PARP), followed by a progressive fall in the mitochondrial transmembrane potential (Delta(psi)m), between 4 and 48 h after treatment. Ceramide levels began to increase 2 h after the addition of anti-Fas (with no increase during the first hour), and increased continuously to 640% of control cells at 48 h. In the moderately sensitive SCC61 adherent cells, comparable results were observed, though with lower levels of ceramide and a delay in the response kinetics, with apoptotic cells becoming flotant. Finally, despite early cleavage of caspase-8 at 2 h, and a sustained level of activation until 48 h, no apoptotic response was observed in anti-Fas-resistant SQ20B cells. This was confirmed by a lack of ceramide generation and mitochondrial changes, and by the absence of any detectable cleavage of caspase-3 or PARP. Inhibition of caspase processing, and amplification of endogenous ceramide signalling by pharmacological agents, allowed us to establish the order of cellular events, locating ceramide release after caspase-8 activation and before caspase-3 activation, and demonstrating a direct involvement for ceramide release in mitochondrial dysfunction. Furthermore, these experiments provide strong arguments for the role of endogenous ceramide as a key executor of apoptosis, rather than as a consequence of membrane alterations.

Phospholipids reacylation and palmitoylcoa control tumour necrosis factor-alpha sensitivity.

From the hypothesis that in TNF-alpha-resistant cells the activity of mitochondrial phospholipase A2 could be reversed by a lysophospholipid acyltransferase, we report that the mitochondrial reacylation of phosphatidylcholine as phosphatidylethanolamine was considerably higher in C6 (TNF-alpha-resistant) than in WEHI-164 (TNF-alpha-sensitive) cells. TNF-alpha did not modify the phospholipids' reacylation in C6, while in WEHI-164 it was increased several-fold. These results suggest that TNF-alpha is not sufficient to restore the barrier permeability in sensitive cells, but may be enough to explain the absence of permeability change in resistant cells. AcylCoA esters, depending on whether the acyl group is unsaturated or saturated (palmitic acid), could control membrane permeability either by participating in the reacylation of phospholipids or keeping the pore in a closed state. The analysis of the endogenous acylCoA ester pools of both cell lines show that the amount of palmitoylCoA is higher in resistant than sensitive cell lines. TNF-alpha treatment does not change these results.

[Adenocarcinomas of the distal esophagus and cardia. Anatomy and volume of irradiation]. / Adénocarcinome du bas oesophage et du cardia. Anatomie et volumes à irradier.

The esophagus is divided into four regions: cervical esophagus, intrathoracic esophagus with upper, mid and lower thoracic portion. Cancer may occur on each of these regions. Computed tomography of the thorax and superior abdomen and endoscopic ultrasound are necessary for reliable staging. CT simulation allows accurate definition of tumor volume. GTV includes tumor volume and regional lymph nodes. CTV encompasses GTV plus safety margin and lymph nodes areas considered to harbor potential microscopic disease. The extent of prophylactic lymph node irradiation depends on the anatomic location of the primary tumor.

Comparison of methods used to study cell death in an adherent tumoral cell line with moderate clonogenic radiosensitivity.

Our objective was to compare different methods for studying programmed cell death in adherent H460 non-small lung cancer cells of moderate clonogenic radiosensitivity. The major effect of gamma-radiation was found to be the release of cells from the substratum. The different methods gave complementary and unexpected information: a) with the TUNEL method, a few non-apoptotic cells were found in the culture medium; b) with the flow cytometry after propidium iodide labeling, some hypodiploid cells which remained attached to the substratum were apoptotic, as demonstrated by the effect of a caspase inhibitor; c) with the annexin V labeling, the detached cells were demonstrated either necrotic or very late apoptotic; d) the mitochondria transmembrane potential (deltapsim), measurements demonstrated that the mitochondria were implicated in cell death induced by gamma-radiation. These data illustrate the need to use several complementary methods in the study of apoptosis in adherent cells exposed to gamma-radiation.

Up-regulation of mitochondrial peripheral benzodiazepine receptor expression by tumor necrosis factor alpha in testicular leydig cells. Possible involvement in cell survival.

Porcine Leydig cells in primary cultures are resistant to tumor necrosis factor alpha (TNFalpha) cytotoxicity. Here we report that these cells can be rendered sensitive to TNFalpha killing by treatment with the translational inhibitor cycloheximide, suggesting the existence of proteins that can suppress the death stimulus induced by the cytokine. In search of these cytoprotective proteins, we focused on the constituents of the mitochondrial permeability transition pore (PT pore), whose opening has been shown to play a critical role in the TNFalpha-mediated death pathway. We found that TNFalpha up-regulated mRNA and protein expression of the mitochondrial peripheral benzodiazepine receptor (PBR), an outer membrane-derived constituent of the pore. A strong correlation was established between the resistance of the cells to TNFalpha killing and the density of PBR-binding sites. Concomitantly, TNFalpha down-regulated Bcl-2 mRNA and protein expression. As Bcl-2 has been shown to be an endogenous inhibitor of the PT pore, we hypothesize that the TNFalpha-induced up-regulation of PBR expression may compensate for the decrease in Bcl-2 levels to prevent the opening of the PT pore.

Tumor necrosis factor-alpha inhibits leydig cell steroidogenesis through a decrease in steroidogenic acute regulatory protein expression.

The aim of the present study was to identify the sites of the inhibitory action of TNFalpha (tumor necrosis factor alpha) on LH/hCG-stimulated testosterone formation. By using cultured porcine Leydig cells as a model, TNFalpha was shown to inhibit testosterone secretion when testicular cells were stimulated with hCG but not when incubated with 22R-hydroxycholesterol (a cholesterol substrate derivative that readily passes through cell and mitochondrial membranes). Such an observation suggested that the cytokine may affect cholesterol transport and/or availability to cytochrome P450scc in the mitochondria. Specifically, we report here that TNFalpha reduced in a dose- and time-dependent manner hCG-induced StAR (steroidogenic acute regulatory protein) levels. The maximal and half-maximal effects were obtained with 20 ng/ml (1.2 nM) and 1.6 ng/ml (0.09 nM) of TNFalpha, respectively. Maximal inhibitory effects of TNFalpha on StAR messenger RNA and protein levels were obtained after 48 h of treatment. Additionally, the presence of TNFalpha receptors P55 in terms of protein (identified through cross-linking experiments) and messenger RNA (identified through RT-PCR analysis) suggested that the effects of the cytokine are directly exerted on the testicular steroidogenic cell type.

Hormone-induced changes in cardiolipin from Leydig cells: possible involvement in intramitochondrial cholesterol translocation.

The rate-limiting and hormonally regulated step in steroid hormone biosynthesis is the delivery of cholesterol from the outer to the inner mitochondrial membrane where cytochrome P450scc resides. Although the exact mechanism of intramitochondrial cholesterol translocation remains unknown, the formation of contact sites between outer and inner mitochondrial membranes appears as a necessary component for cholesterol transfer. Several pieces of evidence suggest that local formation of intermembrane contact is a consequence of a non-bilayer arrangement of polymorphic lipids which are enriched in the junctions. As a step toward clarifying mitochondrial contact sites formation and thus cholesterol translocation in steroidogenic cells, we have undertaken studies to identify the factors which might result in non-bilayer structure to be adopted by mitochondrial phospholipids on stimulation of MA-10 Leydig cells. Our results demonstrate that an increase in the unsaturation of the cardiolipin acyl groups on hormonal stimulation might favor the formation of non-bilayer adhesion points.

A possible involvement of endogenous polyamines in the TNF-alpha cellular sensitivity.

A critical step in the cytotoxic action mechanism of tumor necrosis factor-alpha (TNF-alpha) involves, among mitochondrial dysfunctions, an early change of the inner membrane permeability displaying the characteristics of permeability transition. Cytosolic polyamines, especially spermine, are known to inhibit it. Our results show that spermine is only detectable in the TNF-alpha resistant C6 cells while N1-acetylspermidine is present in the TNF-alpha sensitive WEHI-164 cells, and putrescine and spermidine are found in both. TNF-alpha treatment does not change this distribution but only induces a quantitative alteration in TNF-alpha sensitive cells. Omission of glutamine (energetic substrate) from the culture media alters neither the TNF-alpha responsiveness of both cell lines nor their polyamine distributions, only their quantitative polyamine contents.

Radiotherapy with high dose rate brachytherapy boost and concomitant chemotherapy for Stages IIB and III esophageal carcinoma: results of a pilot study.

PURPOSE: Radiotherapy (RT) and concomitant chemotherapy (CT) is the standard treatment for non resectable esophageal cancer. Usual total radiation dose is 50 Gy. In order to enhance local control rate a Phase II study was initiated to evaluate the feasibility of a combined treatment with an external radiation dose of 60 Gy and three cycles of concomitant CT, using the three main active drugs (CDDP, 5 FU and MMC), followed by a high dose rate (HDR) brachytherapy delivering 10 Gy. METHODS AND MATERIALS: Fifty-three patients, 48 men and 5 women, were entered in this study. Stages were evaluated with CT scan and with endoscopic sonography. Fifteen were Stage IIB, 38 Stage III. Treatment consisted of conventional fractionated RT to a total dose of 60 Gy delivered with 2 Gy per fraction, one fraction per day and five fractions per week. The CT regimen was a combination of Cisplatinum (CDDP) 20 mg/m2 and 5 Fluorouracil (5FU) 600 mg/m2 continuous infusion, from days 1-4 Mitomycin C (MMC) was given at 6 mg/m2 on day 1. Three cycles were administered on days 1, 22, and 43. Brachytherapy was delivered one week after the end of external radiation therapy. RESULTS: Full radiation therapy dose was delivered for 94% of the patients. CT compliance, evaluated on the mean relative dose-intensity was 85% for CDDP, 81% for 5FU and 51% for MMC. Overall grade 3 and 4 WHO toxicity rates were 23% and 7%, respectively. Haematologic toxicity was the most limiting factor. One patient died from treatment toxicity. Local control rate at one year was 74%. Three-year actuarial survival rate was 27%. Distant metastasis was the main cause of treatment failure. Swallowing score was good for 75% of the patients. Stage, performance status and weight loss were prognostic factors. CONCLUSION: This regimen with high dose RT, HDR brachytherapy and concomitant CT is feasible; however, a high level of haematologic toxicity was observed with the CDDP, 5FU and MMC regimen. Despite a poor compliance with CT, treatment results are very encouraging for patients with locally advanced disease.

Increase of mitochondrial PLA2-released fatty acids is an early event in tumor necrosis factor alpha-treated WEHI-164 cells.

We have previously reported that TNF induced changes in mitochondrial enzymes, one of which, succinate-dehydrogenase, is specifically activated in various TNF-sensitive cell lines. In an attempt to further characterize the mechanism of trans-membrane signalling at the mitochondrial level, we have oriented our investigation to the study of phospholipase A2 activity localized in this organelle isolated from TNF-treated WEHI-164 cells. Under physiological conditions, this enzyme has a very low basal activity near the resting state, while under TNF treatment its activity is dramatically increased. This event is induced by TNF concentrations which also cause cytolysis; however, the activity of this enzyme is increased several hours before maximum cytotoxicity occurs. The activation of the mitochondrial phospholipase A2 is not arachidonoyl or fatty acid selective, as is the case for the cytosolic species. Phospholipase A2 and succinate-dehydrogenase display higher activities simultaneously under TNF treatment. This change in activity is linked to morphologic modifications. Mitochondria in particular display an orthodox state characterized by a large and clear matrix space and few crests.

Study of O-sialylation of glycoproteins in C6 glioma cells treated with retinoic acid.

When treated with retinoic acid in vivo, C6 glioma cells show an enhancement of CMP-Neu5Ac:Gal beta 1-3 GalNAc-R alpha-2,3 sialyltransferase activity. A 300 kDa glycoprotein was detected by lectin affinoblotting in retinoic acid-treated C6 cells which stained weakly or not at all in control cells. Comparative studies with different lectins demonstrated that this glycoprotein contains alpha 2,3 Neu5Ac Gal-GalNAc O-glycan moieties. Cultures in the presence of an inhibitor of O-glycan synthesis (N-acetylgalactosaminide alpha-O-benzyl) demonstrated that enhancement of staining of the 300 kDa glycoprotein was not due to the increase of the alpha 2,3 sialytransferase but to the de novo synthesis of the polypeptide chain of this glycoprotein.

Study of retinoic acid effect upon retinoic acid receptors beta (RAR-beta) in C6 cultured glioma cells.

Using monoclonal antibodies against the RAR-alpha and RAR-beta retinoic receptors, we demonstrated that these receptors were present together in C6 glioma cells as two isoforms of 50 and 55 kDa. For RAR-beta, the 50 kDa isoform predominated (60 to 80% of the total of the two isoforms). After a treatment for 48 h with retinoic acid 10 microM, the 55 kDa form was enhanced, while no effect was observed either on RAR-alpha isoforms from C6 cells and on both RAR-alpha and RAR-beta forms from neuroblastoma SKN SH SY5Y used as a control. Using purified neuronal and glial rat brain nuclei, we showed that the 55 kDa isoform from RAR-beta predominated in glial cells. These results suggest that retinoic acid treatment of C6 cells led to a partial differentiation, the enhancement of the heavy form of RAR-beta being a marker of this phenomenon.