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Background and purpose Plasticity of the intestinal stem cell compartment in response to radiation injury is regulated by a stem cell niche. We present here the first experimental observations of a dose-volume effect of the intestinal stem cell niche and of the solitary intestinal lymphoid tissues (SILT). Materials and methods Regeneration of intestinal crypts in mice was studied following irradiation of millimetre-size jejunal sections with single doses of 6 to 24 Gy and compared to total body irradiation (TBI). The statistical distribution of cells per crypt was scored and regressed to a biomathematical model. The number of SILTs was scored for different doses and field sizes and crypt regeneration was correlated with SILT proximity. Results We observed a differential dose–response of the intestinal stem cell niche at the centres of the irradiated sections, but only for field sizes below 10 mm. Irradiation of 5 mm jejunum results in an increase in crypt survival by up to an order of magnitude, compared to TBI. Distributions of cell-per-crypt numbers and comparison to biomathematical modelling suggest that these observations stem from a field size-dependent regeneration rate. The density of SILTs also exhibits a volume-dependent dose–response and increased crypt survival correlates with a proximity to SILTs. Conclusion Our findings present the first observation of a field-size dependent dose–response of the intestinal stem cell niche. Its regeneration process does apparently not rely on distant radiation-sensitive resources of the organism, such as the bone marrow. Yet, our observations suggest that the niche interacts with intact tissue in millimetres distance, leading to faster crypt regeneration. The field-size dependent dose–response of SILTs posits a role of the immune system on the dose-volume effect.

Background and purpose The high plasticity of the intestinal epithelium is maintained by a resilient reserve stem cell population, whose extent and biology are a matter of ongoing debate. The in vivo clonogenic assay (IVCA), presents a well established and efficient analysis of radiation insult to the intestinal crypts. However, we found that inadequate mathematical analysis over the last four decades led to systematic errors and contradictory results in estimates of radio-sensitivity and size of the reserve stem cell pool. Material and methods We devised a refinement of the IVCA via development of a biomathematical model that delivers a full statistical dynamic description of epithelial radiation injury and subsequent regeneration. We validated the model against cellular and crypt distribution statistics obtained from IVCA experiments and through systematic re-analysis of experimental data from 27 publications. Results A full dynamic description of the evolution of stem cell niche population statistics is obtained. A systematic re-analysis reveals a consistent clonogenic content of the crypt of 31 ± 6 cells. The stem cell reserve manifests to be, contrary to prior predictions, radio-resistant: α = ( 0.22 ± 0.04 ) Gy−1. Conclusion We established a precision tool for the quantitative analysis of radiation insult to the intestinal crypts, which we employ to show that the reserve stem cell population is small, radio-resistant, and remarkably immutable against a large variety of interventions. The increased resolution of the model allows not only a reduction of the number of animals by about 75%, but also to quantify experimentally the influence of additional agents on damage and on regeneration of the stem cell niche.

Purpose: Currently, it is unclear if the variation in proton relative biological effectiveness (RBE) at different positions in the spread-out Bragg peak (SOBP) should be taken into account for medical applications. To date, the majority of studies have examined cells irradiated in vitro. In this work RBE estimates have been made for intact tissue irradiated ex vivo. Materials and Methods: Irradiations were performed in the 200 MeV clinical proton beam produced at iThemba LABS (Cape Town, South Africa). The beam was modulated to produce either a 3-cm or a 7-cm SOBP. The number of intestinal crypts regenerated in jejunum sections of mice after single doses of protons was used as the biological end point. For this, a novel ex vivo irradiation technique was implemented to allow measurements at the very end of the SOBP where most variations in proton RBE can be expected. Results: The proton RBE at the very end of the 7-cm SOBP increases by 10% compared with the middle of the SOBP; the same 10% increas...

The present in vitro study aimed to assess the effects of combining the mTOR inhibitor RAD001 and temozolomide (TMZ) together with irradiation by either low-linear energy transfer (LET) radiation (γ-rays) or high-LET radiation (fast neutrons) on the growth and cell survival of the human glioblastoma cell line U-87. We observed a strong decrease in cell proliferation along with a concomitant increase in cell death as a function of the radiation dose. As expected, high-LET radiation was more effective and induced more sustained damage to DNA than low-LET radiation. While RAD001 in association with TMZ induced autophagic cell death, additional combination with either type of radiation did not further increase its occurrence. On the contrary, apoptosis remained at a low level in all experimental groups.

International audience; Purpose: To analyze the dose rate influence in hyper-radiosensitivity (HRS) of human melanoma cells to very low doses of fast neutrons and to compare to the behaviour of normal human skin fibroblasts.Materials and methods: We explored different neutron dose rates as well as possible implication of DNA double-strand breaks (DSB), apoptosis, and energy-provider adenosine-triphosphate (ATP) levels during HRS.Results: HRS in melanoma cells appears only at a very low dose rate (VLDR), while a high dose rate (HDR) induces an initial cell-radioresistance (ICRR). HRS does not seem to be due either to DSB or to apoptosis. Both phenomena (HRS and ICRR) appear to be related to ATP availability for triggering cell repair. Fibroblast survival after neutron irradiation is also dose rate-dependent but without HRS.Conclusions: Melanoma cells or fibroblasts exert their own survival behaviour at very low doses of neutrons, suggesting that in some cases there is a differential between cancer and normal cells radiation responses. Only the survival of fibroblasts at HDR fits the linear no-threshold model. This new insight into human cell responses to very low doses of neutrons, concerns natural radiations, surroundings of accelerators, proton-therapy devices, flights at high altitude. Furthermore, ATP inhibitors could increase HRS during high-linear energy transfer (high-LET) irradiation.

Historically, improvements in radiotherapy have been mainly due to improvements in physical selectivity: beam penetration, collimation, dosimetry, treatment planning; and advances in imaging. Neutrons were the first high-LET (linear energy transfer) radiation to be used clinically and showed improvement in the differential response of radiation resistant tumors and normal tissues. The benefits of fast neutrons (and other forms of high LET radiations) are due to their biological effects: a reduction of the OER, a reduction in the differential cell radiosensitivity related to their position in the mitotic cycle, and a reduction in cellular repair capacity (thus less importance of fractionation). The poor physical selectivity of the early neutron therapy beams introduced a systematic bias in comparison with the photon treatments and created a negative perception for neutron therapy. However, significant improvements in the neutron therapy equipment resulted in a physical selectivity similar to modern MV photon therapy. The tumor types or sites where the best therapeutic results were obtained included inoperable or recurrent salivary gland tumors locally extended prostatic adenocarcinomas, and slowly growing well-differentiated sarcomas. The benefit of neutrons for some other well-defined groups of patients was demonstrated in randomized trials. It was estimated that about 20 % of all radiotherapy patients could benefit from fast neutrons (if neutrons are delivered under satisfactory physical conditions). An important issue for fast neutron therapy is the selection of the types of patients who could most benefit from high-LET radiations. The same issue is raised today with other high-LET radiations (e.g., 12C ions). It is reasonable to assume that the same types of patients would benefit from 12C irradiation. Of course the better physical selectivity of ion beams enhances the treatment possibilities but this is true for both the high-LET and low-LET radiations (i.e., moving from neutrons to 12C ions and from photons to protons, respectively). An important area of research involves developing criteria to identify specific patients suitable for high-LET radiation. One promising technique is to measure the RBE of the cancer cell population in vitro mainly in head and neck tumors. Modern molecular imaging allows the identification of hypoxic or proliferative regions in the tumor. Special MRI examinations are also able to identify hypoxic regions. A promising predictive test recently initiated, is the study of non-repairable double strand breaks but the utility of the technique needs to be confirmed. The extensive experience with fast neutron therapy can greatly assist the transition to high-LET charged-particle therapy.

The effect of combining RDC-11 (a new ruthenium organometallic complex) and fast neutrons (high-LET radiations) has been evaluated in RDM4 and U-87 tumor cell lines. After co-treatment, a substantial reduction of cell growth was recorded, which appears not to be correlated with the induction of apoptosis.

High linear energy transfer (LET) radiation have the ability to kill cancer cells resistant to conventional radiotherapy. On the other hand, protocols combining radiotherapy and chemotherapy are effective in eradicating certain inoperable cancers. In this study, we investigated the cytotoxicity of a co-treatment with fast neutrons and cisplatin in a human glioblastoma cell line, U-87. Cells cultured in vitro were irradiated with p(65)+Be neutrons in the presence of cisplatin. Cell survival and the induction of apoptosis and premature senescence were assessed at different time intervals thereafter, using a variety of methods. A marked reinforcement of the cytotoxicity was obtained when irradiation and cisplatin were associated. This reflected both an amplification of the apoptotic process and the induction of premature cell senescence. The efficiency of a combination between fast neutrons and cisplatin in inducing cell death in U-87 is more than additive. The present data concur with those we previously reported in a mouse lymphoma and suggest the potential utility of platinum compounds as adjuncts to future cancer therapy protocols using high-LET radiation.

L'utilisation de rayonnements a transfert lineique d'energie (TLE) eleve, tels que les neutrons rapides et les ions carbone (hadrontherapie) offre de prometteuses perspectives en radiotherapie. Bien qu'en combinant la radiotherapie et la chimiotherapie, d'importants benefices therapeutiques peuvent etre obtenus pour traiter le cancer, peu d'investigations ont ete consacrees a ce jour aux effets de traitements associant une irradiation avec des rayonnements a TLE eleve et des medicaments anticancereux. Dans ce travail, nous avons etudie ces effets apres une irradiation par des neutrons rapides de 65 MeV combines avec du cisplatine, in vitro sur des cellules RDM4 (lymphome T murin). Les cellules ont ete irradiees a des doses variant de 2 a 8 Gy avec ou sans ajout de cisplatine juste avant l'irradiation a des concentrations entre 0,3 et 12,5 μM. Les deux traitements ont ete appliques concomitamment. La proliferation et l'induction d'apoptose ont ete determinees a intervalles reguliers apres les traitements. Une cytotoxicite plus importante a ete observee lors de l'association de l'irradiation et du cisplatine. De plus, la cytotoxicite induite par ce co-traitement resulte non seulement de l'apoptose mais aussi d'autres formes de mort cellulaire.

The objective of this study was to characterize the apoptotic pathways activated by fast neutrons in the human lymphoblastoid cell line TK6 and in its p53 -/- derivative. Our results demonstrate that while p53 is not required for neutron-induced apoptosis, as previously shown, it does affect the kinetics of apoptosis and the molecular pathways leading to the activation of effector caspases. Indeed, rapid p53-dependent apoptosis was associated with the activation of caspase 9, 8, 3, and 7 and the cleavage of BID by caspase 8. In contrast, the slow-occurring p53-independent apoptotic process, mediated by caspase 7, took place without BID cleavage and loss of transmembrane mitochondrial potential. Altogether, our findings highlight an essential role for caspase 8-mediated BID cleavage, in the course of p53-dependent apoptosis triggered by fast neutrons in lymphoid cells. They also demonstrate that this mechanism is not involved in p53-independent apoptosis.

BACKGROUND AND PURPOSE: The primary objective of this study was to establish in vivo the relationship between 2-2-nitro-1H-imidazol-1yl-N-(2,2,3,3,3-pentafluoropropyl)-acetamide (EF5) adduct formation and intratumoral oxygen concentrations measured by electron paramagnetic resonance (EPR) in a tumor model mimicking a clinical situation. The secondary objective was an attempt to calibrate in situ the immunofluorescence (IF) signal with EPR oximetry. MATERIALS AND METHODS: IM syngeneic fibrosarcoma (NFSA) bearing C3H mice were used. Three days after injection of a paramagnetic charcoal into the tumor, the mice were anesthetized, injected with the hypoxic marker EF5, and monitored every 20 min for 3 h with a low-frequency EPR spectrometer. Animals were allowed to breath either under 21 or 100% O(2). Tumors were then harvested, frozen, cut into sections including the charcoal and processed for EF5 adducts detection using monoclonal antibodies. Slices were viewed with a fluorescence microscope and 190x140 micrometer areas surrounding the charcoal were digitized and analyzed with the NIH-Image and Adobe Photoshop software. The fluorescence intensity (FI) was measured in the whole pictures and in strips of 10 micrometer around the charcoal. RESULTS: EF5 binding increased with decreasing pO(2), most substantially at pO(2) below 5 mm Hg. Baseline (ambient air) pO(2) reached 3.2+/-2.1 mm Hg in NFSA tumors. It increased to 9.8+/-3.2 mm Hg under 100% O(2). A statistically significant correlation was observed on an individual tumor basis between the FI in the first 10 micrometer strip around the charcoal and the pO(2) determined by EPR oximetry (Wilcoxon signed rank test: P

The involvement of the tumor suppressor p53 gene in the sensitivity of many cell types towards low linear energy transfer (LET) radiation is now well established. However, little information is available on the relationship between p53 status of tumor cells and their ability to undergo apoptosis following exposure to high-LET radiation. Here we present the results of experiments carried out with the human lymphoblastoid cell line TK6 and its p53 knock-out counterpart NH32. Cells were irradiated at doses ranging from 0.25 to 8 Gy with fast neutrons (65 MeV), carbon ions (95 MeV/nucleon), and X rays (15 MV). For both cell lines, the occurrence of apoptosis, determined by the quantification of hypodiploid particles as well as the activation of several caspases, was compared with their sensitivity towards high-LET radiation. Results indicate that p53 is involved in the response of TK6 cells to fast neutrons and carbon ions, as measured by cell proliferation and occurrence of apoptosis. However, p53-deficient cells are still able to undergo apoptosis following irradiation. This suggests that heavy ions and fast neutrons induce cellular damage that is not under the control of p53. The involvement of executioner caspases in high-LET radiation induced apoptosis was also evaluated by use of specific inhibitors.Key words: fast neutrons, carbon ions, apoptosis, p53, lymphoblastoid cell line.

BACKGROUND AND PURPOSE: To determine the influence of the number of fractions (or the dose per fraction) on the proton relative biological effectiveness (RBE). MATERIALS AND METHODS: Intestinal crypt regeneration in mice was used as the biological endpoint. RBE was determined relative to cobalt-60 gamma rays for irradiations in one, three and ten fractions separated by a time interval of 3.5h. Proton irradiations were performed at the middle of a 7-cm Spread Out Bragg Peak (SOBP). RESULTS: Proton RBEs (and corresponding gamma dose per fraction) at the level of 20 regenerated crypts per circumference were found equal to 1.15+/-0.04 (10.0 Gy), 1.15+/-0.05 (4.8 Gy) and 1.14+/-0.07 (1.7 Gy) for irradiations in one, three and ten fractions, respectively. Alpha/beta ratios as derived from direct analysis of the 'quantal radiation response data' were found to be 7.6 Gy for gamma rays and 8.2 Gy for protons. Additional proton irradiations in ten fractions at the end of the SOBP were found to be more effective than at the middle of the SOBP by a factor of 1.14 (1.05-1.23). CONCLUSION: Proton RBE for crypt regeneration was found to be independent of fractionation up to ten fractions. One can expect that it remains unchanged for higher number of fractions as the lethalities for doses smaller than 3 Gy are exclusively due to direct lethal events. As a tendency for increased effectiveness at the end of the SOBP is reported in the majority of the studies, for clinical applications it would be advisable to allow for by arranging a sloping depth dose curve in the deeper part of the target volume. Finally, it must be noticed that most of in vitro and in vivo RBE values for protons are larger than the current clinical RBE (RBE=1.10).

To assess the capacity of heavy ions to induce apoptosis in lymphocytes, mice have been irradiated with accelerated carbon ions (95 MeV/nucleon) at doses ranging from 0.1 to 4 Gy. Their spleens were removed 24 h later and gently dissociated to prepare a single cell suspension. Mononuclear cells were then maintained in culture at 37°C, and the occurrence of apoptosis in these cells was analysed 24 h later. Lymphocytes were also irradiated in vitro, in the presence of Ac-DEVD-CHO, a potent caspase-3 and -7 inhibitor. Results from three experiments performed at the Grand Accelerateur National d'Ions Lourds (GANIL, Caen, France) are reported here. They indicate that carbon ions induce a marked, dose-dependent, reduction of the spleen weight and cellularity. However, in sharp contrast with spleen cells prepared from X-ray irradiated mice, only a slight increase of apoptosis is evidenced in cultured lymphocytes from mice irradiated with heavy ions. The significance of such results is discussed. So far, few data exist concerning the biological effects of heavy ions, in particular their capacity to induce apoptosis in lymphocytes; the present study provides useful clues for further investigations.Key words : lymphocytes, apoptosis, heavy ions, splenic model, caspases.

Radiation therapy with carbon ions as well as missions into outer space have boosted the interest for high-LET particle radiobiology. Optimization of treatments in accordance with technical developments, as well as the radioprotection of cosmonauts during long missions require that research in these domains continue. Therefore suitable radiation fields are needed. Fast neutrons and carbon ions exhibit comparable LET values and similar radiobiological properties. Consequently, the findings obtained with each radiation quality could be shared to benefit knowledge in all concerned domains. The p(66+Be) neutron therapy facilities of iThemba LABS (South Africa) and the p(65)+Be neutron facility of Louvain-la-Neuve (Belgium) are in constant use to do radiobiological research for clinical applications with fast neutrons. These beams – which comply with all physical and technical requirements for clinical applications – are now fully reliable, easy to use and frequently accessible for radiobiological investigations. These facilities thus provide unique opportunities to undertake radiobiological experimentation, especially for investigations that require long irradiation times and/or fractionated treatments.

The effects of high-linear energy transfer (LET) radiations on lymphoid tissues and lymphocytes are not well understood. As a first approach to delineate these effects, the present work was conducted to assess the effects of high-LET radiations on murine spleen cells ex vivo and in vitro. BALB/c mice were irradiated whole-body with 65 MeV neutrons or 15 MV X rays at doses ranging from 0.2 to 3 Gy. Spleens were removed 1 day postirradiation and weighed, and single cell suspensions were prepared and cultured for several days. Apoptosis occurring in vitro was determined at different times by flow cytometry analysis of cells labeled with propidium iodide. It was found that irradiation with fast neutrons reduced spleen weight and cellularity to a greater extent than photons. Considering the spleen cellularity as end point, the relative biological effectiveness (RBE) of fast neutrons was 2. However, for both modes of irradiation, apoptosis of recovered spleen cells in vitro increased as a function of dose and the duration of culture. The level of apoptosis occurring at various times postirradiation was found to be identical for high- and low-LET radiations. Taken together, these results suggest that external as well as cellular factors might differentially modulate the sensitivity of lymphocytes to fast neutrons and photons.

Experiments were performed with human plasma irradiated in vitro or in vivo in order to evaluate the extent to which clastogenic factors might disturb the adaptive response to DNA-damaging factors currently studied in our laboratory. The studies were carried out with plasma isolated from whole blood given 4 Gy of X-rays in vitro and with plasma from people receiving local radiotherapy at a total dose of about 60 Gy gamma rays. Addition of irradiated plasma to culture medium did not result in a statistically significant increase in structural aberrations in chromosomes of non-irradiated normal blood.

BACKGROUND AND PURPOSE: Thorough knowledge of the RBE of clinical proton beams is indispensable for exploiting their full ballistic advantage. Therefore, the RBE of the 200-MeV clinical proton beam produced at the National Accelerator Centre of Faure (South Africa) was measured at different critical points of the depth-dose distribution. MATERIAL AND METHODS: RBEs were determined at the initial plateau of the unmodulated and modulated beam (depth in Perspex = 43.5 mm), and at the beginning, middle and end of a 7-cm spread-out Bragg peak (SOBP) (depths in Perspex = 144.5, 165.5 and 191.5 mm, respectively). The biological system was the regeneration of intestinal crypts in mice after irradiation with a single fraction. RESULTS: Using 60Co gamma-rays as the reference, the RBE values (for a gamma-dose of 14.38 Gy corresponding to 10 regenerated crypts) were found equal to 1.16 +/- 0.04, 1.10 +/- 0.03, 1.18 +/- 0.04, 1.12 +/- 0.03 and 1.23 +/- 0.03, respectively. At all depths, RBEs were found to increase slightly (about 4%) with decreasing dose, in the investigated dose range (12-17 Gy). No significant RBE variation with depth was observed, although RBEs in the SOBP were found to average a higher value (1.18 +/- 0.06) than in the entrance plateau (1.13 +/- 0.04). CONCLUSION: An RBE value slightly larger than the current value of 1.10 should be adopted for clinical application with a 200-MeV proton beam.

Treatment of hepatocellular carcinoma (HCC) is a major concern for physicians as its response to chemotherapy and radiotherapy remains generally poor, due, in part, to intrinsic resistance to either form of treatment. We previously reported that an irradiation with fast neutrons, which are high-linear energy transfer (LET) particles, massively induced autophagic cell death in the human HCC SK-Hep1 cell line. In the present study, we tested the capacity of the mammalian target of rapamycin (mTOR) inhibitor RAD001 to augment the cytotoxicity of low and high-LET radiation in these cells. As mTOR is a key component in a series of pathways involved in tumor growth and development, it represents a potential molecular target for cancer treatment. Results indicate that RAD001, at clinically relevant nanomolar concentrations, enhances the efficacy of both high- and low-LET radiation in SK-Hep1 cells, and that the induction of autophagy may account for this effect. However, fast neutrons were found to be more efficient at reducing tumor cell growth than low-LET radiation.

Les courbes de survie cellulaire dependent de facteurs «biologiques» propres au materiel irradie (type cellulaire, souche, origine, position dans le cycle mitotique, degre d'oxygenation, temperature,...) et de facteurs «physiques» lies a la qualite du rayonnement (TEL faible ou eleve) et a ses conditions d'application (debit de dose eleve, irradiation continue a faible debit, fractionnement,...). Nous etudierons l'influence de chaque facteur et examinerons les differents modeles et concepts utilises pour l'interpretation des courbes et l'analyse de leurs formes. Leur role en radiotherapie sera egalement rappele

Radiobiological data indicate that high-LET radiations are more effective than low-LET radiations in treating hypoxic, well differentiated, and slowly growing tumors. The clinical outcomes of fast neutron therapy still serve as the basis for the selection of ion beam therapy (IBT). Patient selection is a complex issue and robust predictive tests are needed and in development but not presently validated. Randomized clinical trials comparing modern high- and low-LET radiotherapy machines and treatment planning systems should be organized. For this, protocols and outcomes should be described, reported, and analyzed applying internationally accepted terminology, concepts, and approaches.

Hepatocellular carcinoma (HCC) represents the sixth most common cancer worldwide and a major health problem since the choice of treatment is limited due to chemo- and radio-resistance. It was previously reported that high linear energy transfer (LET) radiation induced massive autophagic cell death in the human HCC SK-Hep1 cell line in vitro. This study analyzed the effects of high-LET radiation on the same HCC tumor model, orthotopically transplanted into nude mice. For this purpose, after surgical xenograft in the liver, animals were irradiated with fast neutrons and cell death occurring in the tumors was assessed with various techniques, including electron microscopy and probe-based confocal laser endomicroscopy. Results indicate that considerable autophagy and only limited apoptosis took place in the tumor xenografts after high-LET irradiation. These data confirm the previous in vitro results, suggesting that autophagy may act as a predominant mode of cell death in the efficacy of high-LET radiation.

The aim of the present study was to determine the cytotoxic consequences of high-linear energy transfer (LET) irradiation in the presence of oxaliplatin on hepatocellular carcinoma (HCC) cells in vitro. We attempted to correlate the induction of apoptosis and autophagy with the formation of DNA double-strand breaks (DSBs). SK-Hep1 cells were irradiated by 65 MeV neutrons in the presence of oxaliplatin and/or the poly(ADP-ribose) polymerase (PARP) inhibitor PJ34. DSBs were measured by the formation of gammaH2AX foci. Results show that in SK-Hep1 cells exposed to fast neutrons in the presence of oxaliplatin, DSBs occurred and persisted with time after irradiation. While apoptosis remained low in co-treated cells, autophagy was considerably increased after irradiation and augmented by the addition of oxaliplatin. Thus, autophagic cell death appears to play a prominent role in the cytotoxicity of the combined treatment and may be linked to the generation of heavy damage to DNA.

Purpose The relative biological effectiveness (RBE) in the presence or absence of CHK2 was estimated at the Korean National Cancer Center Proton Therapy Center (NCCPTC). Methods and Materials The proton beam was fixed at 210 MeV with 6-cm spread-out Bragg peaks (SOBPs) because this is expected to be the most frequently used clinical setting. X-rays were obtained using a 6-MV conventional linear accelerator. The RBE was estimated from the survival of jejunal crypt in C3H/He and Chk2 -/- mice. Results The estimated RBEs of the NCCPTC at the middle of the SOBP were 1.10 and 1.05 in the presence and absence of CHK2, respectively. The doses that reduced the number of regenerated crypt per jejunal circumference to 20 (D 20 ) in C3H/He mice were 14.8 Gy (95% confidence interval [CI], 13.7–15.9) for X-rays and 13.5 Gy (95% CI, 14.5–15.5) for protons. By contrast, the doses of D 20 in Chk2 -/- mice were 15.7 Gy (95% CI, 15.0–16.4) and 14.9 Gy (95% CI, 14.0–15.8) for X-rays and protons, respectively. Conclusions The RBE of the NCCPTC is clearly within the range of RBEs determined at other facilities and is consistent with the generic RBE value of 1.10 for 150- to 250-MeV beams. The mutation of Chk2 gave rise to radioresistance but exhibited similar RBE.

Modern protocols of concomitant chemo/radiotherapy provide a very effective strategy to treat certain types of tumors. High-linear energy transfer (LET) radiations, on the other hand, have an increased efficacy against cancer with low radiosensibility and critical localization. We previously reported that oxaliplatin, a third generation platinum drug, was able to reinforce the cytotoxicity of an irradiation by fast neutrons towards human glioblastoma U-87 cells in culture. We show here that such a combination has the capacity to enhance the number of double strand breaks in DNA and to induce autophagy in these cells. Xenografts experiments were further performed in nude mice subcutaneously transplanted with U-87 cells. When injected shortly before a single irradiation by fast neutrons, oxaliplatin causes a marked reduction of tumor growth compared with the irradiation alone. Overall, our data indicate the unique cytotoxic mechanism of a combined high-LET irradiation and oxaliplatin treatment modality and suggest its potential application in anticancer therapy.

Table sugar samples were irradiated with accelerated carbon ions and fast neutrons. Electron paramagnetic resonance (EPR) analysis performed after the irradiation revealed a complex spectrum similar to that observed after gamma-ray irradiations. The total concentration of the paramagnetic centers induced by accelerated carbon ions and neutrons was proportional to the absorbed dose. Good stability of the produced free radicals was observed for a typical period of sugar storage. Sugar was more sensitive to accelerated carbon ions than to neutrons. The results show that table sugar can be a useful material for dosimetry in the case of a radiation accident. (C) 2009 Elsevier Ltd. All rights reserved.

Due to the RBE variations, the carbon-ion doses (in Gy) are no longer sufficient to monitor adequately the biological effect of these radiations. Therefore, "RBE dose weighting factors" - W(RBE) - allowing for the RBE variations with energy, dose and biological system have to be introduced in the treatment plans in order to provide the physician with interpretable information. This paper compares the methods employed for this purpose at NIRS and GSI, which are specific of the beam delivery system of these institutions. NIRS has a "passive" beam delivery system where the dose distribution in the SOBP is determined by a Ridge filter. The dose distribution - and thus, the shaping of the filter - is chosen according to the clinical situation and determined with respect to W(RBE) factors in order to yield a biologically iso-effective SOBP. W(RBE )factors in the SOBP are at first derived from a RBE/LET function for HSG cells, then normalized to 3 at a LET of 80 keV/mum. The latter value of 3 corresponds to the clinical RBE of NIRS-neutrons, which were found to exhibit the same radiobiological properties as 80 keV/mum carbon-ions. GSI has a "dynamic" beam delivery system ("spot" or "voxel" scanning) making it possible to irradiate irregular volumes and to modulate the radiation intensity according to the radiosensitivity of different tissues and/or different sub-volumes. Due to the "power" and the resulting complexity of the system, W(RBE )factors are determined through an integrated calculation code allowing iterative interaction of both physical and radiobiological parameters. The "Local Effect Model" (LEM) was developed in this view with the aim of deriving carbon-ion W(RBE )factors from the parameters determining the response to photons. Advantages and weaknesses of the respective methods will be discussed.

The combination of high-linear energy transfer (LET) radiation with chemotherapeutic agents may offer new perspectives in cancer treatment. We therefore assessed the consequences of a treatment in which U-87 human glioblastoma cells were irradiated with p(65) + Be neutrons in the presence of oxaliplatin, a third generation platinum anticancer drug having higher apoptosis-inducing activity than cisplatin. Cell survival, apoptosis, cell cycle progression as well as p21 and p53 protein expressions were analyzed. Results show that an enhanced cytotoxic effect was obtained when the two treatments were combined and that, unlike what we previously observed with cisplatin, this was not due to a reinforcement of apoptosis. Altogether, our results also indicate the potential of oxaliplatin for use in association with high-LET radiation against tumors refractory to conventional photon radiotherapy. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

The RBE of the new MIT fission converter epithermal neutron capture therapy (NCT) beam has been determined using intestinal crypt regeneration in mice as the reference biological system. Female BALB/c mice were positioned separately at depths of 2.5 and 9.7 cm in a Lucite phantom where the measured total absorbed dose rates were 0.45 and 0.17 Gy/ min, respectively, and irradiated to the whole body with no boron present. The gamma-ray (low-LET) contributions to the total absorbed dose (low- + high-LET dose components) were 77% (2.5 cm) and 90% (9.7 cm), respectively. Control irradiations were performed with the same batch of animals using 6 MV photons at a dose rate of 0.83 Gy/min as the reference radiation. The data were consistent with there being a single RBE for each NCT beam relative to the reference 6 MV photon beam. Fitting the data according to the LQ model, the RBEs of the NCT beams were estimated as 1.50 +/- 0.04 and 1.03 +/- 0.03 at depths of 2.5 and 9.7 cm, respectively. An alternative parameterization of the LQ model considering the proportion of the high- and low-LET dose components yielded RBE values at a survival level corresponding to 20 crypts (16.7%) of 5.2 +/- 0.6 and 4.0 +/- 0.7 for the high-LET component (neutrons) at 2.5 and 9.7 cm, respectively. The two estimates are significantly different (P = 0.016). There was also some evidence to suggest that the shapes of the curves do differ somewhat for the different radiation sources. These discrepancies could be ascribed to differences in the mechanism of action, to dose-rate effects, or, more likely, to differential sampling of a more complex dose-response relationship.

BACKGROUND AND PURPOSE: The Relative Biological Effectiveness (RBE) of 8 fast-neutron beams, 5 proton beams and 1 carbonion beam was determined using as biological criterion intestinal crypt regeneration in mice, i.e. an in vivo system. These beams are used or planned for clinical cancer therapy applications. In addition, the RBE of 6 epithermal neutron beams, used or planned for Boron Neutron Capture Therapy (BNCT), was determined; no boron was administered. The goal of the program was to improve the exchange of information between the centers, facilitate the interpretation of the results and increase the safety of the clinical applications. MATERIALS AND METHODS: In all visited centers, the same technique was applied in the same conditions by the same radiobiology team. The number of regenerating crypts per circumference was scored 3.5 days after single fraction total body irradiation. The control irradiations were performed locally using cobalt-60 units. The mice were randomized according to radiation quality and dose level. RESULTS: (1) For fast neutron beams, the RBE (Ref. cobalt-60 gamma rays) increases with decreasing energy (from approximately 1.7 for p(65)+Be neutrons to approximately 2.4 for d(14.5)+Be neutrons). In addition, it is specific to each facility and depends on the nuclear reaction (p or d + Be), target and collimation type. (2) For proton beams, the RBEs (Ref cobalt-60 gamma rays) at the reference position (middle of a 7-cm Spread Out Bragg Peak, SOBP) range between 1.08 and 1.18. They might differ by approximately 6-8% according to the mode of beam production or delivery. The RBEs at the end of the SOBP are always 5-10 % higher than at the middle of the SOBP. (3) For the carbon ion beam studied at NIRS in Chiba, Japan, the RBE significantly increases with depth. Relative to gamma rays, it ranges from 1.3 in the initial plateau, 1.6 at the beginning, 1.7 at the middle and 1.9 at the end of a 6-cm SOBP. 4) In BNCT beams, the radiation quality (in particular the relative contribution of the different dose components) varies rapidly with depth and depends strongly on the arrangement of the irradiation set-up (e.g. presence or not of back scattering material). Moreover, the (total) dose rates are highly variable (from 0.05 to approximately 0.5 Gy/min) according to the power of the reactors. Wide range of RBE values (Ref. gamma rays) was thus obtained (RBE = 1.4 - 2.2) at shallow depths of 1.5 - 2.5 cm. DISCUSSION AND CONCLUSION: Intestinal crypt regeneration in mice is an in vivo system perfectly suitable to perform intercomparisons between centers applying different types of non-conventional radiation qualities. It was proven to be reproducible, reliable and accurate, and becomes progressively recognized worldwide as part of the Quality Control (QA) procedures for new beams. It should be stressed that the observed RBE for intestinal crypt cells after a single high dose provide some radiobiological characterization of the radiation quality but cannot be used as the RBE weighting factor in clinical prescriptions.

The rationale for introducing ion beams in cancer therapy is the high level of physical selectivity that can be achieved with ions, equal or even better than with proton beams or modern photon techniques, as well as the potential advantage of high-LET radiations for some tumour types and sites. The radiobiological arguments for high-LET radiation in cancer therapy are reviewed: reduction of OER in the case of hypoxic and poorly-reoxygenating tumours, and the lesser importance of repair phenomena which are a problem in controlling repair-proficient photon-resistant tumours. Fast neutrons were the first type of high-LET radiation used clinically, and were often applied under suboptimal technical conditions. Nevertheless, useful clinical information was derived from the neutron experience. A greater benefit from neutrons than from conventional radiotherapy was found for several tumour sites. The present discussion is limited to the results for salivary gland tumours and prostatic adenocarcinoma. Based on the fast neutron experience, radiobiological arguments, and the added benefit of excellent physical selectivity of ion beams, the potential clinical indications for high-LET ions are discussed: hypoxic, slowly growing and well-differentiated photon-resistant tumours. One of the main remaining issues is the selection of individual patients for high- or low-LET radiation. Since the physical selectivity of ions now matches that obtained with other techniques, the selection of patients will be based only on the radiobiological characteristics of the tumour.

We have previously shown that ionizing radiation-induced apoptosis in human lymphoblastoid cells differs according to their p53 status, and that caspase 8-mediated cleavage of BID is involved in the p53-dependent pathway. In the present study, we investigated the role of Fas signaling in caspase 8 activation induced by fast neutrons irradiation in these cells. Fas and FasL expression was assessed by flow cytometry and by immunoblot. We also measured Fas aggregation after irradiation by fluorescence microscopy. We found a decrease of Fas expression after irradiation, but no change in Fas ligand expression. We also showed that, in contrast to the stimulation of Fas by an agonistic antibody, Fas aggregation did not occur after irradiation. Altogether, our data strongly suggest that fast neutrons induced-apoptosis is Fas-independent, even in p53-dependent apoptosis.

PURPOSE: To determine the relative biologic effectiveness (RBE) of the Paul Scherrer Institute (PSI) scanning proton beam in reference conditions and to evaluate the influence of intestine motion on the proton dose homogeneity. METHODS AND MATERIALS: First, RBE was determined for crypt regeneration in mice after irradiation in a single fraction. Irradiation was performed at the middle of a 7-cm spread out Bragg peak (SOBP; reference position), as well as in the proximal part of the plateau and at the distal end of the SOBP. Control gamma-irradiation was randomized with proton irradiation and performed simultaneously. Second, motion of mouse intestine was determined by radiographs after copper wire markers had been placed on the jejunum and intestinal wall. RESULTS: Proton RBE (reference (60)Co gamma) was equal to 1.16 for irradiation at the middle of the SOBP and to 1.11 and 1.21 for irradiation in the initial plateau and end of the SOBP, respectively. The confidence intervals for these RBE values were much larger than those obtained in the other proton beams we have tested so far. They exceeded +/-0.20 (compared with the usual value of +/-0.07), which resulted from the unusually large dispersion of the individual proton data. The instantaneous positions of the mice intestines varied by +/-2 mm in the course of irradiation. CONCLUSION: The results of this study have shown that the RBE of the PSI proton beam is in total accordance with the RBE obtained at the other centers. This experiment has corroborated that proton RBE at the middle of the SOBP is slightly larger than the generic value of 1.10 and that there is a slight tendency for the RBE to increase close to the end of the SOBP. Also, excessive dispersion of individual proton data may be considered to result from intestine motion, taking into account that irradiation at the PSI is delivered dynamically by scanning the target volume with a pencil proton beam ("spot scanning"). Because 2-mm movements resulted in significant variations in local dose depositions, this should be considered for moving targets. Strategies to reduce this effect for the spot scanning technique have been developed at the PSI for radiotherapy of humans.

The purpose of this study was to assess apoptosis in hematopoietic tumor cells irradiated with carbon ions, in order to define its contribution to the cytotoxicity of these high-LET radiations.RDM4 (murine T lymphoma), MOLT-4, TK6 and WTK1 (human lymphoblastoid) cells were irradiated with 12C or 13C. Apoptosis was assessed by flow cytometry. Cell growth and activities of caspases were determined during the same periods.The ability of carbon ions to induce apoptosis markedly varied according to the cell line. MOLT-4 and TK6 cells underwent apoptosis at 1 Gy within 12 hours post-irradiation, whereas RDM4 and WTK1 showed little apoptosis under the same conditions.These results indicate that apoptosis contributes to the overall cytotoxicity of carbon ions towards hematopoietic tumor cells although other death mechanisms must also account for this cytotoxicity.

Purpose: Plasma citrulline, a nitrogen end product of glutamine metabolism in small-bowel enterocytes, was suggested as a marker of radiation-induced small-bowel epithelial cell loss in mice after single-dose whole-body irradiation. Our objective was to evaluate the feasibility of citrulline as a marker for radiation-induced small-intestinal mucosal atrophy in patients during and after abdominal fractionated radiotherapy. Methods and Materials: Twenty-three patients were studied weekly during treatment and at intervals of 2 weeks and 3 and 6 months after treatment by postabsorptive plasma citrulline concentration and clinical toxicity grading. The interrelationship between these variables and the correlation with small-bowel dose and volume parameters were investigated. Results: During fractionated radiotherapy, citrulline concentration significantly decreased as a function of the radiation dose (p < 0.001) and the volume of small bowel treated (p = 0.001). The plasma citrulline concentration correlated with clinical toxicity during the last 3 weeks of treatment. As a whole, citrulline concentration correlated better with radiation dose and volume parameters than clinical toxicity grading. Conclusions: In patients treated with fractionated radiation therapy for abdominal or pelvic cancer sites, plasma citrulline concentration may be a simple objective marker for monitoring epithelial cell loss, a major event in acute radiation-induced small-bowel toxicity. (C) 2004 Elsevier Inc.

Radiation therapy is a very important and effective treatment modality for almost all types of malignancies. Its importance in the local control of primary or bulky cancers will even increase in the future if more effective chemotherapeutic or other systemic treatments become available to treat metastatic disease.

Citrulline: a physiologic marker enabling quantitation and monitoring of epithelial radiation-induced small bowel damage.Lutgens LC, Deutz NE, Gueulette J, Cleutjens JP, Berger MP, Wouters BG, von Meyenfeldt MF, Lambin P.MAASTRO, Department of Radiation Therapy and Oncology, University Hospital Maastricht/Maastricht University, Maastricht, The Netherlands. l.lutgens@rtil.nlPURPOSE: Small bowel irradiation results in epithelial cell loss and consequently impairs function and metabolism. We investigated whether citrulline, a metabolic end product of small bowel enterocytes, can be used for quantifying radiation-induced epithelial cell loss. METHODS AND MATERIALS: NMRI mice were subjected to single-dose whole body irradiation (WBI). The time course of citrullinemia was assessed up to 11 days after WBI. A dose-response relationship was determined at 84 h after WBI. In addition, citrullinemia was correlated with morphologic parameters at this time point and used to calculate the dose-modifying factor (DMF) of glutamine and amifostine on acute small bowel radiation damage. RESULTS: After WBI, a time- and dose-dependent decrease in plasma citrulline level was observed with a significant dose-response relationship at 84 h. At this time point, citrullinemia significantly correlated with jejunal crypt regeneration (p < 0.001) and epithelial surface lining (p = 0.001). A DMF of 1.0 and 1.5 was computed at the effective dose 50 (ED50) level for glutamine and amifostine, respectively. CONCLUSIONS: Citrullinemia can be used to quantify acute small bowel epithelial radiation damage after single-dose WBI. Radiation-induced changes in citrullinemia are most pronounced at 3 1/2 to 4 days postirradiation. At this time point, citrullinemia correlates with morphologic endpoints for epithelial radiation damage

YingExperimental determination of the RBE of new isotopes for brachytherapy implants (e.g. iodine-125 and palladium-103) remains a very difficult problem, especially in small animals, where the seeds cannot be implanted easily in the planned geometry in a reproducible way. This technical note describes an original device that makes it possible to irradiate a segment of the intestine in mice for the purpose of determining the RBE for crypt regeneration. The device is a length of tube (3.4 mm and 7 mm internal and external diameter, respectively) whose external surface has been longitudinally grooved and into which the seeds can be squeezed (each groove holds either one or two seeds). The tube is composed of two sections. This seed container can be surgically positioned around an intestinal ansa while the mice are anesthetized. The mean dose rates in the intestine (for eight seeds) were found to be 86.3 +/- 5.9 and 79.0 +/- 5.4 cGy/h for 29.2 MBq (1 U) iodine and 28.6 MBq (1 U) palladium seeds, respectively. So far, more than 100 mice have been irradiated successfully. Full dose-effect relationships can be obtained using the same seeds and applying them successively in different groups of animals (which ensured the accuracy of the relative doses).

PURPOSE: This study aims at providing relative biological effectiveness (RBE) data under reference conditions accounting for the determination of the "clinical RBE" of protons. METHODS AND MATERIALS: RBE (ref. (60)Co gamma-rays) of the 200 MeV clinical proton beam produced at the National Accelerator Centre (South Africa) was determined for lung tolerance assessed by survival after selective irradiation of the thorax in mice. Irradiations were performed in 1, 3, or 10 fractions separated by 12 h. Proton irradiations were performed at the middle of a 7-cm spread out Bragg peak (SOBP). Control gamma irradiations were randomized with proton irradiations and performed simultaneously. A total of 1008 mice was used, of which 96 were assessed for histopathology. RESULTS: RBEs derived from LD50 ratios were found not to vary significantly with fractionation (corresponding dose range, approximately 2-20 Gy). They, however, tend to increase with time and reach (mean of the RBEs for 1, 3 and 10 fractions) 1.00, 1.08, 1.14, and 1.25 for LD50 at 180, 210, 240, and 270 days, respectively (confidence interval approximately 20%). alpha/beta ratios for protons and gamma are very similar and average 2.3 (0.6-4.8) for the different endpoints. Additional irradiations in 10 fractions at the end of the SOBP were found slightly more effective ( approximately 6%) than at the middle of the SOBP. A control experiment for intestinal crypt regeneration in mice was randomized with the lung experiment and yielded an RBE of 1.14 +/- 0.03, i.e., the same value as obtained previously, which vouches for the reliability of the experimental procedure. CONCLUSION: There is no need to raise the clinical RBE of protons in consideration of the late tolerance of healthy tissues in the extent that RBE for lung tolerance was found not to vary with fractionation nor to differ significantly from those of the majority of early- and late-responding tissues.

Specification of radiation quality is an important issue in fast neutron therapy since the biological effectiveness of the beams varies to a large extent with neutron energy. It must meet specific criteria, mainly derived from the accuracy requirement for absorbed dose delivery. A first approach to this problem consists in identifying physical parameters that can be related to Relative Biological Effectiveness (RBE) and which describe the beam production technique (e.g. neutron-producing reaction, p + Be or d + Be, energy of the incident particle). A second is based on microdosimetry, which provides a description of the secondary radiation components to which the biological consequences of irradiations are more directly correlated. A third approach consists in experimental RBE determinations in reference conditions: intestinal crypt regeneration in mice after irradiation to the whole body with single doses is proposed as a standard biological system for radiobiological calibrations of clinical fast neutron beams. Dosimetric, microdosimetric and radiobiological intercomparisons are encouraged since they provide a homogeneous set of data which facilitate the exchange of clinical information. They also constitute a basis for the clinical RBE approach and an overall check of the irradiation procedure. Therefore they should be recommended in every non-conventional radiation therapy facility.

In fast neutron therapy, the relative biological effectiveness (RBE) of a given beam varies to a large extent with the neutron energy spectrum. This spectrum depends primarily on the energy of the incident particles and on the nuclear reaction used for neutron production. However, it also depends on other factors which are specific to the local facility, eg, target, collimation system, etc. Therefore direct radiobiological intercomparisons are justified. The present paper reports the results of an intercomparison performed at seven neutrontherapy centres: Orleans, France (p(34)+Be), Riyadh, Saudi Arabia (p(26)+Be), Ghent, Belgium (d(14.5)+Be), Faure, South Africa (p(66)+Be), Detroit, USA (d(48)+Be), Nice, France (p(65)+Be) and Louvain-la-Neuve, Belgium (p(65)+Be). The selected radiobiological system was intestinal crypt regeneration in mice after single fraction irradiation. The observed RBE values (ref cobalt-60 gamma-rays) were 1.79 +/- 0.10, 1.84 +/- 0.07, 2.24 +/- 0.11, 1.55 +/- 0.04, 1.51 +/- 0.03, 1.50 +/- 0.04 and 1.52 +/- 0.04, respectively. When machine availability permitted, additional factors were studied: two vs one fraction (Ghent, Louvain-la-Neuve), dose rate (Detroit), influence of depth in phantom (Faure, Detroit, Nice, Louvain-la-Neuve). In addition, at Orleans and Ghent, RBEs were also determined for LD50 at 6 days after selective abdominal irradiation and were found to be equal to the RBEs for crypt regeneration. The radiobiological intercomparisons were always combined with direct dosimetric intercomparisons and, when possible in some centres, with microdosimetric investigations.

Despite many recent technical advances, including the use of accelerated high linear energy transfer (LET) particles, cancer radiotherapy remains poorly effective in controlling certain types of tumors due to the frequent failure of malignant cells to undergo a cell death program after radiation exposure. Inactivating molecular components of signalling networks that are involved in tumor radioresistance thus represents an approach to improve cancer radiotherapy. Growing evidence suggests that one such component is the mammalian target of rapamycin (mTOR), a protein kinase that is commonly hyperactive in human tumors. We therefore evaluated the potent and selective mTOR inhibitor RAD001 (everolimus) for its capacity to enhance the effects of low-LET (X-rays and gamma rays) and high-LET (65MeV neutrons) radiation in two human cancer cell lines: SK-Hep1 (hepatocellular carcinoma) and U-87 (glioblastoma). Cellular growth (SRB assay) and the occurrence of DNA double-strand breaks (DSBs, determined by γH2AX foci) and cell death (apoptosis and autophagy, assessed by several methods) were monitored at various times post-irradiation. Clonogenic survival assays and western blotting of mTOR were also performed. RAD001 (20nM) was added to cell cultures 1 hr before the irradiation and maintained throughout the experiments. Irradiation was administered as a single fraction, at doses ranging from 2 to 16Gy. While highly resistant to low-LET radiation, both cell lines proved less resistant to fast neutrons, in agreement with previous studies. When associated with either type of radiation, RAD001 strongly reduced cell growth and also affected clonogenic survival, albeit marginally. Moreover, RAD001 treatment combined with neutron irradiation was found to induce the highest levels of γH2AX foci persistence and cytotoxicity in both cell lines. However, the type of cell death differed radically between the two cell lines. In the SK-Hep1 line, whereas only a low percent of cells underwent apoptosis after irradiation, the percent of autophagic cells was elevated and further enhanced by RAD001. By contrast, in the U-87 line, apoptosis contributed for a large part of cell death after irradiation, but its occurrence was decreased by the addition of RAD001 while the incidence of autophagic cells was concomitantly increased. Our results indicate that a correlation may exist between the persistence of DSBs and the induction of autophagy. Furthermore, autophagic cell death appears to account, almost completely in SK-Hep1 and partially in U-87, for the overall cytotoxicity induced by the combination of RAD001 and irradiation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1466. doi:1538-7445.AM2012-1466

The hypothesis that cisplatinum (c-DDP) interacts with radiation by inhibiting the cellular repair capacities, was tested by comparing the interaction of c-DDP with low-LET (60Co gamma-rays) and high-LET radiation (d(50) + Be neutrons) in mice. The biological endpoint was lethality, 6 days after total body irradiation (early intestinal tolerance). The dose modifying factor was 1.80 +/- 0.25 for c-DDP plus 60Co gamma-rays, and 1.97 +/- 0.3 for c-DDP plus neutrons. As less repairable damage is induced by fast neutrons than by photons, this suggests that, in this system, the interaction between radiation and c-DDP is not explained by repair inhibition but is purely additive.

The RBE of p(75) + Be neutrons relative to d(50) + Be neutrons has been determined for chromosome aberrations induced in Allium cepa (onion) roots. Two biological criteria were selected: the average number of aberrations (mainly fragments) per cell in anaphase and telophase, and the percentage of aberration-free cells. The influence of sampling time (3 to 7 h incubation) between irradiation and fixation was investigated systematically. This factor did not significantly influence the results. The RBE values of p(75) + Be neutrons compared to those of d(50) + Be neutrons were 0.85 (0.79-0.91) and 0.87 (0.80-0.95) for the first and the second criteria, respectively. In previous experiments for the same beams, we found an RBE of 0.90 (0.86-0.94) for survival of V79 cells (D0 ratio), 0.96 (0.93-0.99) for the intestinal crypt cell system, and 0.83 (0.70-0.96) for Vicia faba growth delay.

In an early phase II trial combining gemcitabine (dFdC) and radiotherapy for lung carcinomas, severe pulmonary toxicity was observed. In this framework, the objective of this study was to investigate the effect of dFdC on the tolerance of the lungs of C3H mice to single-dose irradiation. The thoraxes of C3H mice were irradiated with a graded single dose of 8 MV photons; dFdC (150 mg/kg) or saline (control animals) was administered i.p. 3 or 48 h prior to irradiation. Lung tolerance was assessed by the LD50 at 7-180 days after irradiation. For irradiation alone, the LD50 reached 14.45 Gy (95% CI 13.33-15.66 Gy). With a 3-h interval between administration of dFdC and irradiation, the LD50 reached 13.29 (95% CI 12.26-14.44 Gy); the corresponding value with a 48-h interval reached 13.01 Gy (95% CI 11.92-14.20 Gy). Our data also suggested a possible effect of dFdC on radiation-induced esophageal toxicity. dFdC has a minimal effect on lung tolerance after single-dose irradiation. However, a proper phase I-II trial should be designed before any routine use of combined dFdC and radiotherapy in the thoracic region.

Male C57Bl/Cnb and BALB/c mice were exposed to single and fractionated d(50) + Be neutrons or 137Cs gamma rays at 12 weeks of age and were followed for life-shortening and disease incidence as ascertained by autopsy and histological examinations at the time of spontaneous death. Fractionation schedules used were 10 exposures at 24-h intervals and 8 exposures at 3-h intervals for gamma rays, and 8 exposures at 3-h intervals for neutrons. The data were analyzed by the Kaplan-Meier procedure using as criteria causes of death and possible causes of death. Individual groups were compared by a modified Wilcoxon test according to Hoel and Walburg (J. Natl. Cancer Inst. 49, 361-372 (1972)). No significant difference was found in C57Bl/Cnb and BALB/c male mice between a single gamma-ray exposure and a single neutron exposure. Gamma-ray fractionation was clearly less effective in reducing survival time than a single exposure. In contrast, fractionation of neutrons was slightly, although not significantly, more effective in reducing survival time than a single exposure. The relative biological effectiveness (RBE) for life-shortening for d(50)-Be neutrons compared to gamma rays is of the order of 1 to 2 for a single exposure to neutrons and between 2 and 3 for fractionated neutrons compared to a single exposure to gamma rays. Neutron irradiation caused somewhat more cancer than gamma irradiation, and the RBE for cancer induction may be higher, probably between 2 and 3 in the range of 1 to 3 Gy, although the present data do not allow a more precise assessment.