This study was initiated to investigate a possible strategy to alter an enzyme deficiency in a mouse model. The enzyme investigated is a multifunctional nucleoside kinase from Drosophila melanogaster (Dm-dNK). This enzyme has special features in that it has higher enzymatic activity than any other known nucleoside kinases and still has similar substrate specificity as the human nucleoside kinases. The deficiency where the Dm-dNK transgenic mice model will be used is a TK2 deficient model with severe phenotype caused by mitochondrial DNA depletion. The Dm-dNK transgenic mice model will be used as a way to rescue the TK2 deficient mice. The results from the present study show that Dm-dNK expression in mice results in a substantial increase of thymidine phosphorylation in several investigated tissues. The mice were otherwise normal as judged by life span, weight and behavior. The mitochondrial DNA was also detected at normal levels. In conclusion, the Dm-dNK mouse model is promising as a way to rescue the severe phenotype of the TK2 deficient mice.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2007. / Härtill 4 uppsatser.

Thèse (Ph. D.)--Université Laval, 2007. / Titre de l'écran-titre (visionné le 5 mai 2008). Bibliogr.

Thèse (M.Sc.)--Université Laval, 2001. / Bibliogr.: f. 86-96. Publié aussi en version électronique.

Thesis (Ph. D.)--Chemistry and Biochemistry, Georgia Institute of Technology, 2008. / Wartell, Roger, Committee Member ; Bunz, Uwe, Committee Member ; Doyle, Donald, Committee Member ; Fahrni, Christoph, Committee Member ; Schuster, Gary, Committee Chair.

by Jose Victor Lopez. / Typescript. / Thesis (M.S.)--Florida State University, 1988. / Bibliography: leaves 98-109.

Made available in DSpace on 2014-10-09T12:53:04Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T13:58:24Z (GMT). No. of bitstreams: 0 / Dissertação (Mestrado) / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP

Made available in DSpace on 2014-10-09T12:53:04Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T13:58:24Z (GMT). No. of bitstreams: 0 / Análogos da timidina têm sido marcados com diferentes radioisótopos devido ao seu potencial em monitorar a proliferação incontrolável de células. Considerando que o radioisótopo tecnécio-99m mantém ainda uma posição privilegiada devido às suas propriedades químicas e nucleares, este trabalho constituiu-se do desenvolvimento de uma nova técnica de marcação da timidina com o 99mTc, mediante o emprego de complexos organometálicos. Os objetivos do trabalho foram: a síntese do complexo organometálico carbonil-tecnécio-99m; marcação da timidina com este complexo precursor; avaliação da estabilidade; e avaliações radioquímicas e biológicas com animais sadios e portadores de tumor. A preparação do complexo precursor, utilizando o gás CO foi de fácil execução, assim como a marcação da timidina com este precursor, obtendo-se uma pureza radioquímica ≥ 97% e ≥ 94%, respectivamente. Sistemas cromatográficos com bons níveis de confiabilidade foram utilizados, podendo qualificar e quantificar as espécies radioquímicas. O resultado do estudo in vitro da lipofilicidade revelou que o a timidina radiomarcada é hidrofílica, com um coeficiente de partição (log P) de -1,48. O complexo precursor e a timidina radiomarcada apresentaram boa estabilidade radioquímica em até 6 h em temperatura ambiente. A estabilidade com soluções de cisteína e histidina apresentaram perdas entre oito e onze pontos percentuais para concentrações de até 300 mM. Os ensaios de biodistribuição em camundongos sadios indicaram que a timidina radiomarcada apresentou uma rápida depuração sangüínea e baixa captação nos demais órgãos, com predominância de excreção da droga pelo sistema urinário e hepatobiliar. A captação tumoral foi baixa, apresentando valores de 0,28 e 0,18 %DI/g para tumor de pulmão e mama, respectivamente. Os resultados obtidos sugerem mais investigações em outros modelos tumorais ou a modificação da estrutura da molécula orgânica que atua como ligante. / Dissertação (Mestrado) / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP

Thymidine kinase 1 (TK1) is a cancer biomarker which has diagnostic and prognostic potential in a variety of malignancies. TK1 is significantly elevated in the serum and tumor tissue of most malignancies. This increase in TK1 can be detected in the very early stages of malignancy, including in pre-malignant disease with an increased risk for progression. Several studies have demonstrated that elevated TK1 is found in serum months before any clinical symptoms of malignancy. It has also been demonstrated that TK1 is elevated months before clinical recurrence of malignancy. This work first sought to demonstrate the early nature of TK1 expression in breast tumor tissue and pre-malignant tissue. We found that TK1 is elevated in breast hyperplasia tissue and breast carcinoma tissue. In this study we also identified some cases of ‘normal’ tumor margins (considered normal by current pathological standards) which also had elevated TK1 expression. Conversely, true normal breast tissue from noncancerous individuals had no reported elevation in TK1 expression. This study illustrated that TK1 is elevated in pre-malignant breast hyperplasia tissue, as well as some 'normal' tumor margins. TK1 expression was significantly elevated in lung, prostate, colon, esophagus, stomach, liver, and kidney tissues. This work further investigated TK1 expression in a variety of malignant tissue including the two leading causes of cancer mortality in men: lung and prostate cancer. In our study, TK1 was significantly elevated in lung and prostate cancer but not significantly elevated in prostate hyperplasia tissue. TK1 expression also increased with increasing grade in prostate carcinoma tissue. Overall, this work demonstrated that TK1 is a good universal marker of malignancy and is elevated in early cancer development. Despite the potential for TK1 as both a screening and monitoring treatment tool, there have been significant challenges associated with developing a clinically relevant method of TK1 detection. This work proposes one clinically relevant method of detection, namely a TK1 ELISA. Using preoperable lung cancer patients and normal controls, we developed a sensitive and specific ELISA which shows highly statistically significant differences in serum TK1 levels between stage 1 and stage 2 lung cancer compared with normal controls. In fact, this TK1 ELISA is more sensitive and accurate than the traditional TK radioassay, which was unable to detect differences in TK1 between early stage lung cancer and normal patients. Although elevated TK1 is not lung cancer specific, we reported significantly elevated TK1 levels in lung cancer sputum. Screening of sputum and serum for TK1 may be one method for the early detection of lung cancer. Overall, we report TK1 has promising diagnostic potential in a variety of malignancies. We also propose one sensitive and specific method to detect TK1 levels which may easily be adapted to meet current clinical applications. We hope this work will help propel TK1 forward into clinical view in the coming years.

L’identification de marqueurs pronostiques et prédictifs du cancer du sein est un facteur important pour une meilleure compréhension du processus évolutif et le développement de thérapies ciblées. Les récepteurs des oestrogènes (RE) représentent ainsi à la fois un marqueur pronostique mais aussi prédictif du traitement par le tamoxifène ou les anti-aromatases. Cependant, un certain nombre de patientes vont évoluer en dépit de traitements anti-hormonaux adaptés. L’objectif de notre travail, a été d’évaluer la méthode de mesure des RE, l’apport des protéases dans la distinction de profils tumoraux pronostiques et prédictifs. Nous avons démontré l’influence du mode de mesure des RE et en particulier de l’expression quantitative sur l’interprétation pronostique et sur une meilleure détermination du bénéfice du traitement en fonction du niveau d’expression des RE. Nous avons montré l’intérêt de l’évaluation des protéases tissulaires uPA, PAI-I et cathépsine-D, pour caractériser l’hétérogénéité des tumeurs en complément des RE. Particulièrement, chez les patientes RE+, des taux élevés de cathépsine-D et de PAI-1 étaient un indicateur de mauvais pronostic. Nous avons développé un nomogramme combinant RE et le statut ganglionnaire à 3 types de protéases : PAI-1, cathépsine-D et la thymidine kinase, pour déterminer la probabilité de survie à 2 et 5 ans. De plus, ces protéases évaluées dans les tumeurs infectées par l’Epstein-Barr virus (EBV), témoignaient de tumeurs biologiquement agressives avec des taux plus élevés de thymidine kinase. Notre travail a contribué à améliorer l’identification de profils des tumeurs en fonction des RE et des protéases et de caractériser les tumeurs viro-induites. / The identification of prognostic and predictive markers is important for a better understanding of the evolutionary process and the development of targeted therapies. Thus estrogen receptors (ER) represent both an important prognostic marker but also predictive of therapies using tamoxifen or aromatase inhibitors. However, a number of patients will evolve despite hormonotherapy. The objective of our work was to evaluate the method for measuring ER, the contribution of proteases in the distinction of prognostic and predictive tumor profiles. In our work, we demonstrated the influence of the mode of measure of ER and in particular its quantitative expression on the prognostic interpretation and a better determination of benefit of treatment depending on the level of expression of ER. We show the interest of the evaluation of tissue proteases uPA, PAI-I and cathepsin-D, to characterize the heterogeneity of tumors in addition to ER. Specifically, in ER + patients, high levels of cathepsin-D and PAI-1 are an indicator of poor prognosis. We developed a nomogram combining ER and nodal status, to 3 types of proteases: PAI-1, cathepsin-D and thymidine kinase, to determine the probability of survival at 2 and 5 years. In addition, these proteases are evaluated in tumors infected with the Epstein-Barr virus (EBV) and shows high rates of thymidine kinase in EBV + BC, reflecting biologically aggressive tumors. Our work has helped to improve the identification of profiles of tumors according to ER and proteases and characterize virus-associated tumors.