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This thesis focuses on how physical and biological factors influence the outcome of exposures to γ/X-rays. That the dose rate changes during real life exposure scenarios is well-known, but radiobiological data from exposures performed at increasing or decreasing dose rates is lacking. In paper I, it was found that an exposure where the dose rate decreases exponentially induces significantly higher levels of micronuclei in TK6 cells than exposures at an increasing or constant dose rate. Paper II describes the construction and validation of novel exposure equipment used to further study this “decreasing dose rate effect”, which is described in paper III. In paper I we also observed a radioprotective effect when cells were exposed on ice. This “temperature effect” (TE) has been known for decades but it is still not fully understood how hypothermia acts in a radioprotective manner. This was investigated in paper IV, where a multiparametric approach was used to investigate the underlying mechanisms. In paper V the aim was to investigate the role of biomarkers and clinical parameters as possible risk factors for late adverse effects to radiotherapy (RT). This was studied in a rare cohort of head-and-neck cancer patients that developed mandibular osteoradionecrosis (ORN) as a severe late adverse effect of RT. Biomarker measurements and clinical factors were then subjected to multivariate analysis in order to identify ORN risk factors. The results suggest that the patient’s oxidative stress response is an important factor in ORN pathogenesis, and support the current view that patient-related factors constitute the largest source of variation seen in the frequency of late adverse effects to RT. In summary, this thesis provides new and important insights into the roles of biological and physical factors in determining the consequences of γ/X-ray exposures. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 3: Submitted. Paper 5: Manuscript.</p>

Ionizing radiation (IR) triggers many signaling pathways stemming from DNA damage, and, independently, from extra-nuclear events. To generate radio-mimetic DNA double-strand breaks (DSBs) without and minimizing the effects on extra-nuclear radiation targets, human (p53+) glioma and carcinoma cells containing bromodeoxyuridine (BrdU)- substituted DNA were treated with Hoechst 33258 followed by long wave-length UV (UV-A) (BrdU photolysis). BrdU photolysis resulted in well-controlled, dose-dependent generation of DSBs equivalent to 0.2 - 20 Gy of IR, as detected by pulse-field gel electrophoresis, accompanied by dose-dependent H2AX phosphorylation at ser-139 and ATM phosphorylation at ser-1981, indicating ATM activation. Furthermore, BrdU photolysis increased phosphorylation of Chk2 (at thr-68) and p53 (at ser-15). p53 phosphorylation was reduced by the ATM inhibitor caffeine, and H2AX phosphorylation was greatly reduced in AT cells, confirming that phosphorylation was primarily ATM-dependent. We also examined the effects of BrdU photolysis on the major cellular signaling ERK pathways. Interestingly, low-dose (≤ 2 Gy-equivalents) BrdU photolysis stimulated ERK1/2 phosphorylation whereas higher doses (≥ 5 Gy eq.) resulted in Em1/2 dephosphorylation. ERK1/2 phosphorylation was ATM-dependent, whereas dephosphorylation was ATM-independent and DSBs dose-dependent. Thus ERK1/2 appear to be both positively and negatively regulated by ATM depending on the severity of the insult to DNA. In summary, few DSBs trigger ATM-dependent ERK1/2 pro-survival signals whereas more DSBs result in ERK1/2 dephosphorylation consistent with a switch from pro-survival to anti-survival signaling that might affect DSBs repair.

Fundação de Amparo a Pesquisa do Estado de Minas Gerais / A radioterapia é uma das principais abordagens terapêuticas utilizadas no tratamento do câncer e é indicada, principalmente, em casos onde as lesões são inoperáveis. No entanto, uma de suas limitações advém dos próprios efeitos biológicos da radiação, além de outros fatores tais como a radiorresistência inerente a alguns tipos tumorais, tais como os cerebrais. Então, a aplicação concomitante de agentes antineoplásicos com radioterapia vem sendo praticada na clínica de modo a maximizar o efeito benéfico do último e ao mesmo tempo minimizar os efeitos colaterais da exposição à radiação ionizante. Dentro deste contexto geral é importante a pesquisa de novos compostos que possam ser selecionados como protótipos para o desenvolvimento de agentes que possuam os menores efeitos adversos possíveis. As tiossemicarbazonas são uma classe de compostos sintéticos que apresenta um amplo perfil farmacológico e já demonstraram atividade antitumoral. Também já foi relatado que a complexação destes compostos com cátions metálicos pode ser capaz de torná-los mais eficazes. O objetivo do presente trabalho foi avaliar o potencial radiossensibilizador de uma tiossemicarbazona derivada de N(4)-metil-toluil-2-acetilpiridina e seu complexo com cobre sobre linhagens celulares de glioblastoma multiforme, o qual de todos os tumores cerebrais é o que apresenta a maior agressividade e o maior índice de morbidade. Foi avaliada a sensibilidade de linhagens celulares de glioblastoma com diferentes status de p53 RT2 e U87 (p53 selvagem) e T98 (p53 mutante) à radiação gama de uma fonte de 60Co. Os resultados de citotoxicidade indicaram que as linhagens em questão apresentam uma radiossensibilidade similar, p53-independente. Os efeitos citotóxicos da tiossemicarbazona Lac e seu complexo CuLac foram avaliados e os resultados indicaram que ambas possuem excelente efeito citotóxico na ordem de 10-8 M em todas as linhagens avaliadas, portanto menores que drogas como a hidroxiuréia, cisplatina e etoposídeo (IC50 entre 10-4 e 10-6 M em média). O tratamento com as tiossemicarbazonas seguido de 6 Gy de radiação gama se mostrou mais eficiente que o tratamento com radiação isolada em todas as linhagens. A complexação com cobre não alterou de modo significativo o efeito antitumoral da tiossemicarbazona livre sobre as linhagens testadas. Análises de fotomicrografias ópticas indicaram que todos os tratamentos ocasionaram alterações morfológicas, tais como arredondamento celular, redução do volume citoplasmático e surgimento de vesículas na membrana citoplasmática. O conjunto de dados indicam que a tiossemicarbazona Lac e seu complexo de cobre possuem potente efeito antitumoral e também induzem radiossensibilização nas linhagens testadas. / Radiation therapy is one of the main therapeutical approaches used for the treatment of cancer and is indicated, mostly, in cases which the lesions are inoperable. However, one of its limitations comes from its own biological effects, besides other factors such as the radioresistance inherent to some types of tumors like the cerebral ones. Therefore, the concurrent aplication of antineoplasic agentes with the radiation therapy has been used in the clinical pratice with the objective of maximize the benefical effects of the latter and at the same time minimize the side effects of the exposure to ionizing radiation. In this general context is important the research for new compounds that can be selected as prototypes for the development of agents that possess the least adverse effects as possible. The thiosemicarbazones are a class of synthetic compounds that present a broad pharmacological profile and have demonstrated antitumoral activity. Also has been reported that the coordination of these compounds to metallic cations may be capable of make them more effective. The objective of the present work was to evaluate the radiosensitizing potential of a thisemicarbazone derived from N(4)-methyl-tolyl-2-acetylpyridine and its copper complex against glioblastoma multiforme cell lines, which of all brain tumors types, are the most agressive and have the highest morbidity. The sensitivity of glioblastoma cell lines with different p53 status RT2 and U87 (p53 wild type) and T98 (p53 mutant) to gamma radiation from a 60Co source was assessed. Citotoxicity results indicated that the cell lines in study presented a similar radiosensitivity, p53-independent. The citotoxic effects of the thiosemicarbazone Lac and its complex CuLac were assayed and the results indicated that both possess na excelent effect at the order of 10-8 M in all cell lines evaluated, therefore smaller than drugs such as hydroxyurea, cisplatin and etoposide (IC50 between 10-4 and 10-6 M). The treatment with the thiosemicarbazones followed by a 6 Gy gamma radiation dose showed to be more effective than the radiation alone in all cell lines. Coordination to Cooper had not changed significantly the antitumoral effect of the free thiosemicarbazone agains the cell lines tested. Morphological analysis of optical photomicrographies indicated that all treatment caused alterations, such as cell rounding, diminishing of the cytoplasmic volume and rising of vesicles at the cytoplasmic membrane. Together these data indicate that the thiosemicarbazone Lac and its metallic complex possess potent antitimoral effect amd also induce radiosensitivity in the tested cell lines.

Made available in DSpace on 2016-08-10T10:38:45Z (GMT). No. of bitstreams: 1 Cristiana da Costa Luciano.pdf: 1987242 bytes, checksum: 2f3402c7b9c457e5f89de59897be59ea (MD5) Previous issue date: 2012-11-14 / The head and neck cancer is the fifth most common in Brazil, being the most predominant histology type the squamous cell carcinoma. Radiation therapy is a procedure for treatment with the efficacy variable, and may play an important role in controlling tumor growth. Faced with this therapy, the patient is exposed to ionizing radiation that can cause adverse effects resulting cessation of treatment. The objective was to evaluate the association of polymorphisms of TP53 and ATM genes in patients with head and neck cancer with adverse effects on normal tissues presented as a result of radiotherapy. Materials: The DNA was extracted 54 samples of peripheral blood of patients with head and neck cancer, then the fragments of TP53 and ATM were amplified and subsequently sequenced to check for any polymorphism which may be responsible for the radiosensitivity of patients. Statistical analysis was performed using the SPSS 17.0 software. Results and Discussion: In univariate analysis, patients who had experienced adverse effects RT suspended acute low and high grade with RTOG skin (p = 0.012). Those who had a family history of cancer showed higher adverse acute laryngeal RTOG TGI high (p = 0.040). The exchange C> T at position 11322 of TP53 (intron 3), the ATM and TP53 polymorphisms analyzed and the frequency of acute and chronic adverse effects were not significant (p>05). Conclusions: Based on these results is of utmost importance that alternatives are created to predict the dose to be prescribed during radiotherapy, preventing adverse effects and discontinuation of treatment and also providing better tumor control. / O câncer de cabeça e pescoço, no Brasil, é o quinto mais comum, sendo o tipo histológico mais predominante de carcinoma de células escamosas. A radioterapia é uma das modalidades de tratamento com eficácia variável, podendo desempenhar um papel importante no controle do tumor. Diante essa terapêutica, o paciente está exposto a radiações ionizantes que podem causar efeitos adversos gerando a interrupção do tratamento. O objetivo foi avaliar a associação de polimorfismos dos genes TP53 e ATM em pacientes com câncer de cabeça e pescoço com efeitos adversos sobre os tecidos normais apresentados como resultado da radioterapia. Materiais: O DNA foi extraído de 54 amostras de sangue periférico de pacientes com câncer de cabeça e pescoço, em seguida os fragmentos de TP53 e ATM foram amplificados e posteriormente sequenciados a fim de verificar se os polimorfismos poderiam estar associados à radiossensibilidade dos pacientes selecionados. A análise estatística foi realizada utilizando o software SPSS 17.0. Resultados e Discussão: Por meio de análise univariada, pacientes que tiveram a RT suspensa apresentaram efeitos adversos agudo de baixo e alto grau com RTOG de pele (p=0,012). Aqueles que tinham história familiar de câncer apresentaram maiores efeitos adversos agudo de laringe com RTOG TGI alto (p=0,040). A troca C>T na posição 11322 do gene TP53 (intron 3), os polimorfismos de TP53 e ATM analisados e a frequência de efeitos adversos agudos e crônicos não foram significativos para (p>0,05). Conclusões: Diante dos resultados obtidos é de suma importância que alternativas sejam criadas para predizer a dose a ser prescrita durante a radioterapia, prevenindo os efeitos adversos e a interrupção do tratamento e ainda, promovendo melhor controle tumoral.

Made available in DSpace on 2016-04-28T20:16:27Z (GMT). No. of bitstreams: 1 2008 - Jessika Mara Martins Ribeiro.pdf: 4505620 bytes, checksum: 3ace512cbb45ea3a26cc5d88a93ac2b2 (MD5) Previous issue date: 2008-06-24 / Irradiation is a physical process efficiently used in the conservation of foods. The purpose of this study was to investigate the effects of irradiation on the natural mycoflora of poultry feeds and on strains of Aspergillus spp. Maize flour samples, soy crumb and feed were collected directly from the production line of a poultry farm in Avelar, RJ, and exposed to doses of 0, 3.5, 8 and 15 kGy of gamma radiation. Counting, isolation and identification of the contaminant mycoflora were performed before and after irradiation. The radiosensitivity of strains of reference of Aspergillus spp. was determined in CYA medium and in corn for doses ranging from 0 to 8 kGy. Comparisons between the morphologies of control and irradiated strains were performed by using macroscopy, optical microscopy and transmission electron microscopy. Toxigenic profile determination and genetic evaluation by RAPD were also carried out. Higher doses have been found to reduce the number of active colonies, causing elimination of the mycoflora at 8 kGy. A larger radiosensitivity of yeasts was observed in comparison with filamentous fungi. A significant reduction in fungi population occurred at 3.5 kGy to levels below the limit that ensures the hygienic quality of ingredients and poultry feeds. The residual mycoflora was found to decrease with post-irradiation time and included mostly Cladosporium spp., Curvularia spp., Fusarium spp. and Aspergillus spp. and sterility of mycelium prevented further identification of the surviving species of Aspergillus spp. Differences in radioresistance were found among species of Aspergillus and the highest tolerance to radiation was observed for A. parasiticus. Initial morphologic changes were found to be more severe during the first isolation after irradiation than in later ones, with the fungi gradually recovering their normal growth rate. Ultrastructural changes in the irradiated strains were observed mostly in the plasmatic membrane and membranous organelles of nuclei and mitochondria. An increase in the rate of production of toxins by the irradiated strains has been found, however no significant alterations have been observed in their genotypes. Such findings apparently indicate that irradiation stress affected the metabolic response of the fungi, leading to a larger production of toxins. In addition, when appropriate conditions of feeding and growth were restored, the physiologic damages were gradually repaired. Therefore, under such circumstances, irradiated strains may resume growing, thus further deteriorating the substratum. / Irradia??o ? um processo f?sico eficazmente utilizado na preserva??o de alimentos. Este estudo teve como objetivo investigar os efeitos da exposi??o ? radia??o gama sobre a micobiota natural de ra??es av?colas e cepas de Aspergillus spp. Amostras de fub?, farelo de soja e ra??o foram coletadas diretamente da linha de produ??o de uma granja av?cola, no munic?pio de Avelar, RJ e, submetidas ?s doses de 0; 3,5; 8 e 15 kGy de radia??o gama. Foram realizados: contagem, isolamento e identifica??o da micobiota contaminante antes e ap?s a irradia??o. A radiossensibilidade de cepas de refer?ncia do g?nero Aspergillus spp. foi estudada em meio CYA e em milho, com doses de 0 a 8 kGy. Foi comparada a morfologia de cepas controle e irradiadas por macroscopia, microscopia ?ptica e microscopia eletr?nica de transmiss?o. Estudou-se o perfil tox?geno e realizou-se uma avalia??o gen?tica pela t?cnica de polimorfismo de ADN amplificado ao acaso. Observou-se redu??o na contagem com o aumento da dose, tendo sido verificada a elimina??o da micobiota com 8 kGy. Constatou-se tamb?m uma maior radiossensibilidade de leveduras, em rela??o aos fungos filamentosos. Ocorreu efeito do tempo de armazenamento, sendo que as contagens ap?s 45 dias foram inferiores ?s encontradas aos 7 dias ap?s a irradia??o. A dose 3,5 kGy proporcionou significativa redu??o da contagem f?ngica, a n?vel abaixo do limite estipulado para a garantia da qualidade higi?nica de ingredientes e ra??o av?cola. No entanto, h? uma micobiota residual, formada principalmente pelos g?neros: Cladosporium spp., Curvularia spp., Fusarium spp. e Aspergillus spp., esses ?ltimos ap?s serem repicados, apresentaram apenas o desenvolvimento de mic?lio est?ril, o que n?o permitiu a identifica??o da esp?cie. Diferen?as em radiossensibilidade foram observadas entre as cepas de refer?ncia de Aspergillus avaliadas, sendo A. parasiticus o mais radiorresistente em ambos os substratos avaliados: meio CYA e milho. Observou-se menor conidiog?nese e aumento de estruturas de resist?ncia, como os escler?cios. As altera??es morfol?gicas foram mais intensas no isolamento inicial ap?s irradia??o sendo que, gradualmente, os isolados irradiados voltaram a apresentar crescimento semelhante ao padr?o nos repiques sucessivos. Altera??es ultraestruturais nas cepas irradiadas foram observadas principalmente em n?vel de membrana plasm?tica e de organelas, em especial n?cleo e mitoc?ndrias. Essas cepas tiveram aumentada sua produ??o de toxinas. Por outro lado, n?o foram observadas altera??es significativas no gen?tipo, ao menos com os tr?s marcadores utilizados. Esses relatos refor?am a hip?tese de que situa??es de estresse induzem uma maior produ??o de toxina pelos fungos. A irradia??o produz um estresse, afetando metabolicamente o fungo. Contudo, ao serem fornecidos nutrientes e condi??es adequadas de crescimento, essa altera??o fisiol?gica ? superada. Esses achados sugerem que cepas sobreviventes ? irradia??o podem voltar a se desenvolver, deteriorando o substrato, desde que sejam fornecidas condi??es favor?veis ao para seu crescimento.

<p>Radiotherapy is one of the most common therapeutic methods for treatment of many types of cancer. Despite many decades of development and experience there is much to improve, both in efficacy of treatment and to decrease the incidences of adverse healthy tissue reactions. Around 20 % of the radiotherapy patients show a broad range in the severity of normal tissue reactions to radiotherapy, and dose limits are governed by severe reactions in the most radiosensitive patients (< 5 %). Identification of patients with low, moderate or high clinical radiosensitivity before commencing of radiotherapy would allow individual adaptation of the maximum dose with an overall increase in the cure rate. Characterization of factors that may modify the biological effects of ionizing radiation has been a subject of intense research efforts. Still, there is no assay currently available that can reliably predict the clinical radiosensitivity. The aim of this work has been to investigate the role of oxidative stress in individual radiosensitivity and evaluate novel markers of radiation response, which could be adapted for clinical use.</p><p>8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG), a general marker of oxidative stress, is one of the major products of interaction of ionizing radiation with DNA and the nucleotide pool of the cell. As 8-oxo-dG is highly mutagenic due to incorrect base pairing with deoxyadenosine, various repair mechanisms recognize and remove 8-oxo-dG. The repaired lesions are released from cells to the extracellular milieu (serum, urine and cell culture medium) where they can be detected as markers for free radical reactions with the nucleic acids.</p><p>Significant variations in background levels as well as in radiation induced levels of 8-oxo-dG in urine have been demonstrated in breast cancer patients (paper 1). Two major patterns were observed: high background and no therapy-related increase vs. low background and significant increase during radiotherapy for the radiosensitive and non radiosensitive patients respectively.</p><p>Studies in paper 2 indicated major contribution of the nucleotide pool to the extracellular 8-oxo-dG levels. The results also implicated induction of prolonged endogenous oxidative stress in the irradiated cells. RNA “knock-down” experiments on the nucleotide pool sanitization enzyme hMTH1 in paper 3 lend further experimental evidence to this assumption.</p><p>The applicability of 8-oxo-dG as a diagnostic marker of oxidative stress was demonstrated in paper 4. Studies on dialysis patients revealed a good correlation between inflammatory responses (known to be associated with persistent oxidative stress) and extracellular 8-oxo-dG.</p><p>In summary, our results confirm that extracellular 8-oxo-dG is a sensitive <i>in vivo</i> biomarker of oxidative stress, primarily formed by oxidative damage of dGTP in the nucleotide pool with a potential to become a clinical tool for prediction of individual responses to radiotherapy.</p>

Radiotherapy is one of the most common therapeutic methods for treatment of many types of cancer. Despite many decades of development and experience there is much to improve, both in efficacy of treatment and to decrease the incidences of adverse healthy tissue reactions. Around 20 % of the radiotherapy patients show a broad range in the severity of normal tissue reactions to radiotherapy, and dose limits are governed by severe reactions in the most radiosensitive patients (&lt; 5 %). Identification of patients with low, moderate or high clinical radiosensitivity before commencing of radiotherapy would allow individual adaptation of the maximum dose with an overall increase in the cure rate. Characterization of factors that may modify the biological effects of ionizing radiation has been a subject of intense research efforts. Still, there is no assay currently available that can reliably predict the clinical radiosensitivity. The aim of this work has been to investigate the role of oxidative stress in individual radiosensitivity and evaluate novel markers of radiation response, which could be adapted for clinical use. 8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG), a general marker of oxidative stress, is one of the major products of interaction of ionizing radiation with DNA and the nucleotide pool of the cell. As 8-oxo-dG is highly mutagenic due to incorrect base pairing with deoxyadenosine, various repair mechanisms recognize and remove 8-oxo-dG. The repaired lesions are released from cells to the extracellular milieu (serum, urine and cell culture medium) where they can be detected as markers for free radical reactions with the nucleic acids. Significant variations in background levels as well as in radiation induced levels of 8-oxo-dG in urine have been demonstrated in breast cancer patients (paper 1). Two major patterns were observed: high background and no therapy-related increase vs. low background and significant increase during radiotherapy for the radiosensitive and non radiosensitive patients respectively. Studies in paper 2 indicated major contribution of the nucleotide pool to the extracellular 8-oxo-dG levels. The results also implicated induction of prolonged endogenous oxidative stress in the irradiated cells. RNA “knock-down” experiments on the nucleotide pool sanitization enzyme hMTH1 in paper 3 lend further experimental evidence to this assumption. The applicability of 8-oxo-dG as a diagnostic marker of oxidative stress was demonstrated in paper 4. Studies on dialysis patients revealed a good correlation between inflammatory responses (known to be associated with persistent oxidative stress) and extracellular 8-oxo-dG. In summary, our results confirm that extracellular 8-oxo-dG is a sensitive in vivo biomarker of oxidative stress, primarily formed by oxidative damage of dGTP in the nucleotide pool with a potential to become a clinical tool for prediction of individual responses to radiotherapy.

Threats from man-made radionuclides include waste issues, increasing number of power plants, underground bomb testing, nuclear weapons, and “dirty bombs”. Until recently the ionizing radiation protection system focused on protecting humans with an implied protection of biota. However, goals of sustainable development and precautionary principles for human activity are leading to an inclusion of plant and animal populations in the protection system. From this perspective, the present thesis examines wetlands that function as sinks for the radionuclide 137Cs, and describes calculated and measured radiation doses to residing biota. Also, multi-level effects from exposure to low-dose rate ionizing radiation were studied. Accumulation of 137Cs after the Chernobyl accident fallout was studied in a riparian wetland with a mean activity concentration of 1 200 kBq m-2 in Sweden (paper I). A mass balance budget of 137Cs showed that the sedimentation of new material was balanced by the decay process of 137Cs in parts of the wetland (paper I). Frogs were identified as organisms of concern in this wetland. Internal radiation doses, based on whole body measurements of frogs, were estimated to be lower than external doses based on soil samples (paper II). Current dose models for biota resulted in a wide range of doses depending on different levels of conservatism in the models. Therefore, in situ measurements with frog-phantoms were found to provide valuable dose information (paper III). Measured doses using frog-phantoms were lower than calculated doses using several dose models. Although a dose conversion factor by FASSET was found to be useful for comparison with measurements in the field. A higher dose was measured to the phantom surface in comparison to inner parts, i.e. the sensitive skin of frogs receives the highest dose. Estimated and measured radiation doses to frogs were below suggested dose rate limits. Low-dose rate 137Cs exposure of eggs and tadpoles from three amphibian species, Scaphiopus holbrookii, Bufo terrestris, and Rana catesbeiana, showed no increased levels of strand breaks in red blood cells, and no effects on development, survival or growth up to metamorphosis (paper IV). The ecological factor larval density had a stronger effect on metamorphic traits than low-dose rate radiation. Higher levels of strand breaks were detected after an acute dose in R. catesbeiana than after a chronic dose supporting a dose rate limit for protection of amphibians rather than a dose limit (paper IV). Based on current knowledge, frogs in the contaminated wetland are probably not exposed to radiation doses from 137Cs that are harmful for the population. However, variations in sensitivity between populations and species, and adaptive responses have been shown for amphibians exposed to other stressors. This supports further research on effects of chronic low-dose rates of ionizing radiation on amphibians.

The tumour suppressor p53-binding protein 1 (53BP1) is phosphorylated following DNA double strand breaks (DSBs); however, little is understood about the upstream signaling pathways that control this phosphorylation. Additionally, it is not known how these processes combine with 53BP1 to control the survival of cells following DNA damage such as that imparted by ionizing radiation (IR), which is the basis of radiotherapy. In this thesis, I have shown that 53BP1 is phosphorylated specifically in S-phase cells, but not relocalized to intranuclear foci, in response to severe oxygen stress. This occurs with only partial dependence on the ATM kinase (Chapter 2). Following IR, I find that both ATM and DNA-PKcs contribute to intranuclear phosphorylated 53BP1 foci, but that this phosphorylation is independent of proximal signaling molecules that control the localization of 53BP1 to initial DSBs (Chapter 3). Furthermore, I show that 53BP1 loss confers sensitivity to IR and this can be further augmented by inhibition of ATM and DNA-PKcs kinases suggesting that there are both 53BP1-dependent and -independent pathways of survival from IR (Chapter 4). These findings may have important implications for molecular pathology and personalized medicine as 53BP1 has recently been found to be activated or lost in subsets of human tumours. I have collaborated to initiate the development of a novel system to interrogate the implications of 53BP1 loss as traditional siRNA approaches in human cancer cells were not feasible (Chapter 5 and Appendix 2). This system can be used in vivo as tumour xenografts to further understand how 53BP1 and the tumour microenvironment interact endogenously and in response to IR. I also present the possibility and proof of concept for the use of 53BP1 as a biomarker in primary human prostate cancer tissue where little is known about 53BP1 biology (Chapter 5).