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Retinoblastoma (Rb) is a rare form of pediatric cancer that develops from retina cells. Bilateral and some unilateral forms of Rb are associated with heterozygous germline mutations of the (retinoblastoma 1) RB1 gene. RB1 mutations are also associated with a significant risk of secondary malignancy like head and neck tumors. Hence, to date, even if Rb patients are less subjected to radiotherapy to treat their primary ocular tumors, their healthy tissues may be exposed to significant doses of ionizing radiation during the treatment against their secondary malignancies with a significant risk of adverse tissue reactions (radiosensitivity) and/or radiation-induced cancer (radiosusceptibility). However, the biological role of the Rb protein in response to radiation remains misunderstood. Since the ataxia telangiectasia mutated (ATM) protein is a key protein of radiation response and since untransformed skin fibroblasts are a current model to quantify cellular radiosensitivity, we investigated here for the first time the functionality of the ATM-dependent signaling and repair pathway of the radiation-induced DNA double-strand breaks (DSB) in irradiated skin fibroblasts derived from Rb patients The major biomarkers of the DSB repair and signaling, namely clonogenic cell survival, micronuclei, nuclear foci of the phosphorylated forms of the X variant of the H2A histone (γH2AX), the phosphorylated forms of the ATM protein (pATM) and the meiotic recombination 11 nuclease (MRE11) were assessed in untransformed skin fibroblasts derived from three Rb patients. Skin fibroblasts from Rb patients showed significant cellular radiosensitivity, incomplete DSB recognition, delay in the ATM nucleo-shuttling and exacerbated MRE11 nuclease activity. Treatment with statin and bisphosphonates led to significant complementation of these impairments. Our findings strongly suggest the involvement of the ATM kinase in the radiosensitivity/radiosusceptibility phenotype observed in Rb cases. [ABSTRACT FROM AUTHOR]

MacCune–Albright syndrome (MAS) is a rare autosomal dominant osteo-hormonal disorder. MAS is characterized by a severe form of polyostotic fibrous dysplasia, 'café-au-lait' pigmentation of the skin and multiple endocrinopathies. MAS was shown to be caused by mosaic missense somatic mutations in the GNAS gene coding for the alpha-subunit of the stimulatory G-protein. MAS is also associated with radiation-induced malignant tumors, like osteosarcoma, fibrosarcoma and chondrosarcoma but their origin remains misunderstood. In parallel, bisphosphonates treatment was shown to improve the MAS patients' outcome, notably by increasing bone density but, again, the molecular mechanisms supporting these observations remain misunderstood. Here, by using fibroblast and osteoblast cell lines derived from 2 MAS patients, the major radiobiological features of MAS were investigated. Notably, the clonogenic cell survival, the micronuclei and the γH2AX, pATM and MRE11 immunofluorescence assays were applied to MAS cells. It appears that cells from the 2 MAS patients are associated with a moderate but significant radiosensitivity, a delayed radiation-induced nucleoshuttling of the ATM kinase likely caused by its sequestration in cytoplasm, suggesting impaired DNA double-strand breaks (DSB) repair and signaling in both fibroblasts and osteoblasts. Such delay may be partially corrected by using bisphosphonates combined with statins, which renders cells more radioresistant. Our findings represent the first radiobiological characterization of fibroblasts and osteoblasts providing from MAS patients. Although the number of studied cases is reduced, our findings suggest that the MAS cells tested belong to the group of syndromes associated with moderate but significant radiosensitivity. Further investigations are however required to secure the clinical transfer of the combination of bisphosphonates and statins, to reduce the disease progression and to better evaluate the potential risks linked to radiation exposure. [ABSTRACT FROM AUTHOR]

Purpose: Humans are increasingly exposed to ionizing radiation (IR). Both low (<100 mGy) and high doses can cause stochastic effects, including cancer; whereas doses above 100 mGy are needed to promote tissue or cell damage. 10–15% of radiotherapy (RT) patients suffer adverse reactions, described as displaying radiosensitivity (RS). Sensitivity to IR's stochastic effects is termed radiosusceptibility (RSu). To optimize radiation protection we need to understand the range of individual variability and underlying mechanisms. We review the potential mechanisms contributing to RS/RSu focusing on RS following RT, the most tractable RS group. Conclusions: The IR-induced DNA damage response (DDR) has been well characterized. Patients with mutations in the DDR have been identified and display marked RS but they represent only a small percentage of the RT patients with adverse reactions. We review the impacting mechanisms and additional factors influencing RS/RSu. We discuss whether RS/RSu might be genetically determined. As a recommendation, we propose that a prospective study be established to assess RS following RT. The study should detail tumor site and encompass a well-defined grading system. Predictive assays should be independently validated. Detailed analysis of the inflammatory, stress and immune responses, mitochondrial function and life style factors should be included. Existing cohorts should also be optimally exploited. [ABSTRACT FROM AUTHOR]

Purpose: Xeroderma Pigmentosum (XP) is a rare, recessive genetic disease associated with photosensitivity, skin cancer proneness, neurological abnormalities and impaired nucleotide excision repair of the UV-induced DNA damage. Less frequently, XP can be associated with sensitivity to ionizing radiation (IR). Here, a complete radiobiological characterization was performed on a panel of fibroblasts derived from XP-group D patients (XPD). Materials and methods: Cellular radiosensitivity and the functionality of the recognition and repair of chromosome breaks and DNA double-strand breaks (DSB) was evaluated by different techniques including clonogenic cell survival, micronuclei, premature chromosome condensation, pulsed-field gel electrophoresis, chromatin decondensation and immunofluorescence assays. Quantitative correlations between each endpoint were analyzed systematically. Results: Among the seven fibroblast cell lines tested, those derived from three non-relative patients holding the p.[Arg683Trp];[Arg616Pro] XPD mutations showed significant cellular radiosensitivity, high yield of residual micronuclei, incomplete DSB recognition, DSB and chromosome repair defects, impaired ATM, MRE11 relocalization, significant chromatin decondensation. Interestingly, XPD transduction and treatment with statins and bisphosphonates known to accelerate the radiation-induced ATM nucleoshuttling led to significant complementation of these impairments. Conclusions: Our findings suggest that some subsets of XPD patients may be at risk of radiosensitivity reactions and treatment with statins and bisphosphonates may be an interesting approach of radioprotection countermeasure. Different mechanistic models were discussed to better understand the potential specificity of the p.[Arg683Trp];[Arg616Pro] XPD mutations. [ABSTRACT FROM AUTHOR]

Purposes: The term 'radiosensitivity' appeared for the first time at the beginning of the 20th century, few years after the discovery of X-rays. Initially used by French and German radiologists, it illustrated the risk of radiation-induced (RI) skin reactions. From the 1950s, 'radiosensitivity' was progressively found to describe other features of RI response such as RI cancers or cataracts. To date, such confusion may raise legal issues and complexify the message addressed to general public. Here, through an historical review, we aimed to better understand how this confusion appeared. Methods: To support our historical review, a quantitative and qualitative wording analysis of the 'radiosensitivity' occurrences and its derived terms was performed with Google books, Pubmed, Web of ScienceTM databases, and in all the ICRP publications. Conclusions: While 'radiosensitivity' was historically related to RI adverse tissue events attributable to cell death, the first efforts to quantify the RI risk specific to each organ/tissue revealed some different semantic fields that are not necessarily compatible together (e.g. adverse tissue events for skin, cataracts for eyes, RI cancer for breast or thyroid). To avoid such confusion, we propose to keep the historical definition of 'radiosensitivity' to any clinical and cellular consequences of radiation attributable to cell death and to introduce the term 'radiosusceptibility' to describe the RI cancers or any feature that is attributable to cell transformation. [ABSTRACT FROM AUTHOR]

PurposeFor 50 years, cellular radiosensitivity has been defined in vitro as the lack of clonogenic capacity of irradiated cells and its mathematical link with dose has been described by the target theory. Among the numerous formulas provided from the target theory, the linear-quadratic (LQ) model empirically describes cell survival as a negative exponential of a second degree polynomial dose-function in which αD is the linear component and βD2is the quadratic one. The LQ model is extensively used in radiobiology (to describe survival curves) and in radiotherapy (the α/β ratio indicates whether tissue reactions can occur early or late after the treatment). However, no biological interpretation of the LQ parameters was proposed to explain together the radiation response in a wide dose range, the radiosensitivity of some genetic syndromes caused by the mutation of cytoplasmic proteins and the hyper-radiosensitivity phenomenon specific to low-dose. The modelFrom a solid amount of experimental data, we hypothesized that the major forms of ataxia telangiectasia mutated (ATM) are cytoplasmic dimers and that ionizing radiation induce ATM monomerization. The resulting ATM monomers diffuse into nucleus to facilitate double-strand-breaks (DSB)recognitionandrepair. Such hypotheses lead to a coherent molecular interpretation of the LQ model by considering the yield of recognized but unrepaired (α-type) DSB and the non-recognized (β-type) DSB. The notion of celltoleranceto unrepaired DSB was introduced by considering that not all DSB are lethal. Cell survival and DSB repair and signaling immunofluorescence data from 42 normal skin fibroblast and 18 tumor human cell lines were used to verify the validity of this biomathematical model proposed. ResultsOur model is validated at different levels by one of the widest spectrum of radiosensitivity. That mathematical developments of the present model imply that β is a Lorentzian function of α was confirmed experimentally. Our model is also relevant to describe the hypersensitivity to low-dose phenomenon. ConclusionsOur model provides a very general picture of human radiosensitivity, independently of the dose, the cell type and the genetic status. [ABSTRACT FROM AUTHOR]

Purpose: To ask whether dose-rate influences low-dose hyper- radiosensitivity and induced radioresistance (HRS/IRR) response in rat colon progressive (PRO) and regressive (REG) cells. Methods: Clonogenic survival was applied to tumorigenic PRO and non-tumorigenic REG cells irradiated with 60Co γ-rays at 0.0025-500 mGy.min−1. Both clonogenic survival and non-homologous end-joining (NHEJ) pathway involved in DNA double-strand breaks (DSB) repair assays were applied to PRO cells irradiated at 25 mGy.min−1 with 75 kV X-rays only. Results: Irrespective of dose-rates, marked HRS/IRR responses were observed in PRO but not in REG cells. For PRO cells, the doses at which HRS and IRR responses are maximal were dependent on dose-rate; conversely exposure times during which HRS and IRR responses are maximal (tHRSmax and tIRRmax) were independent of dose-rate. The tHRSmax and tIRRmax values were 23 ± 5 s and 66 ± 7 s (mean ± standard error of the mean [SEM], n = 7), in agreement with literature data. Repair data show that tHRSmax may correspond to exposure time during which NHEJ is deficient while tIRRmax may correspond to exposure time during which NHEJ is complete. Conclusion: HRS response may be maximal if exposure times are shorter than tHRSmax irrespective of dose, dose-rate and cellular model. Potential application of HRS response in radiotherapy is discussed. [ABSTRACT FROM AUTHOR]

Purpose: By extending to other animal models the experiments on irradiated rabbit testes performed by the German Albers-Schönberg and Frieben and inspired by the preliminary observations of Regaud on spermatogenesis after irradiation, Bergonié and Tribondeau established a link between radiosensitivity and proliferation as a 'law' in 1906. Although it is still popular and taken as one of the founding laws of radiation oncology, it was early considered as a 'first approximation'. Conclusions: More than 100 years later, one tribute paid to these pioneers should be to at least avoid mixing notions like radiosensitivity, proliferation, differentiation, stem cells that they deeply contributed to define and to re-read their works in their original version in order to better understand what the technological and conceptual advances really are today. [ABSTRACT FROM AUTHOR]

Purpose: To assess in vitro mammographic radiation-induced DNA damage in mammary epithelial cells from 30 patients with low (LR) or high (HR) family risk of breast cancer. Materials and methods: Spontaneous and radiation-induced DNA double-strand breaks (DSB) were quantified by using immunofluorescence of the phosphorylated H2AX histone (γH2AX) in different conditions of mammography irradiation (2, 4, 2 ++ 2 mGy). Results: HR patients showed significantly more spontaneous γH2AX foci than LR patients ( p == 0.014). A significant dose-effect was observed, with an exacerbation in HR patients ( p == 0.01). The dose repetition (2 ++ 2 mGy) provided more induced and more unrepaired DSB than 2 mGy and 4 mGy, and was exacerbated in HR ( p == 0.006). Conclusions: This study highlights the existence of DSB induced by mammography and revealed by γH2AX assay with two major radiobiological effects occurring: A low-dose effect, and a LOw and Repeated Dose (LORD) effect. All these effects were exacerbated in HR patients. These findings may lead us to re-evaluate the number of views performed in screening using a single view (oblique) in women whose mammographic benefit has not properly been proved such as HR patients. [ABSTRACT FROM AUTHOR]

Purpose: To ask whether highly metastatic sublines show more marked low-dose hyper-radiosensitivity (HRS) response than poorly metastatic ones. Materials and methods: The progressive (PRO) subline showing tumourigenicity and metastatic potential and the regressive (REG) subline showing neither tumourigenicity nor metastatic potential were both isolated from a parental rat colon tumour. Clonogenic survival, micronuclei and apoptosis, cell cycle distribution, DNA single- (SSB) and double-strand breaks (DSB) induction and repair were examined. Results: HRS phenomenon was demonstrated in PRO subline. Before irradiation, PRO cells show more spontaneous damage than REG cells. After 0.1 Gy, PRO cells displayed: (i) More DNA SSB 15 min post-irradiation, (ii) more unrepaired DNA DSB processed by the non-homologous end-joining (NHEJ) and by the RAD51-dependent recombination pathways, (iii) more micronuclei, than REG cells while neither apoptosis nor p53 phosphorylation nor cell cycle arrest was observed in both sublines. Conclusions: HRS response of PRO subline may be induced by impairments in NHEJ repair that targets G1 cells and RAD51-dependent repair that targets S-G2/M cells. The cellular consequences of such impairments are a failure to arrest in cell cycle, the propagation of damage through cell cycle, mitotic death but not p53-dependent apoptosis. Tumourigenic cells with high metastatic potential may preferentially show HRS response. [ABSTRACT FROM AUTHOR]

Purpose: Human diseases associated with acute radiation responses are rare genetic disorders with common clinical and biological features including radiosensitivity, genomic instability, chromosomal aberrations, and frequently immunodeficiency. To determine what molecular assays are predictive of cellular radiosensitivity whatever the genes mutations, the existence of a quantitative correlation between cellular radiosensitivity and unrepaired DNA double-strand breaks (DSB) repair defects was examined in a collection of 40 human fibroblasts representing 8 different syndromes. Materials and methods: A number of techniques such as pulsed-field gel electrophoresis, plasmid assay and immunofluorescence with antibodies against MRE11, MDC1, 53BP1 and phosphorylated forms of H2AX, DNA-PK were applied systematically. Results and conclusions: Survival fraction at 2 Gy was found to be inversely proportional to the amount of unrepaired DSB, whatever the genes mutations and the assay applied. However, no single assay discriminates the full range of human radiosensitivity. Particularly, nuclear foci formed by the phosphorylation of H2AX do not predict well moderate radiosensitivities. Our findings suggest the existence of an ATM-dependent interplay between the activation of DNA-PK and MRE11. A classification of diseases according their cellular radiosensitivity, their molecular response to radiation and the functional assays permitting their evaluation is proposed. [ABSTRACT FROM AUTHOR]

Purpose: By allowing the visualization of the proteins inside cells, the immunofluorescence technique has revolutionized our view of events that follow radiation response. Particularly, the formation of nuclear foci, their kinetic of appearance and disappearance, and the association-dissociation of protein partners are useful endpoints to better understand the effects of ionizing radiation. Recently, the technique based on the phosphorylation of the histone 2A family, member X (H2AX) has generated a plethora of reports concerning the interaction between the major proteins involved in DNA repair and stress signaling pathways. However, some unavoidable overlaps of excitation and emission wavelength spectra (the so-called bleed-through phenomenon) of the available fluorescent markers are still generating discrepancies and misinterpretations in the choreography of DNA damage response. Biases are particularly strong with the fluorescein isothiocyanate (FITC)-rhodamine couple, tetramethyl rhodamine iso-thiocyanate (TRITC), the most extensively used markers. Method and results: Here, two representative examples of biased co-immunofluorescence with pH2AX proteins that form radiation-induced nuclear foci or not are presented. A brief review of literature points out differences in kinetic of appearance and association-dissociation of radiation-induced pH2AX and MRE11 foci. Conclusion: Through this report, we would like authors to consider more carefully protein co-localizations by performing systematically, before any co-immunofluorescence, immunofluorescence of each protein separately to avoid bleed-through artifacts. [ABSTRACT FROM AUTHOR]