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Source: Dissertation Abstracts International, Volume: 70-03, Section: B, page: 1570. Advisers: Michael E. Scheurer; Jan M. Risser. Includes bibliographical references.

The WHO classification of brain tumors is based on histological features and the aggressiveness of the tumor is classified from grade I to IV, where grade IV is the most aggressive. Today, the correlation between prognosis and tumor grade is the most important component in tumor classification. High grade gliomas, glioblastomas, are associated with poor prognosis and a median survival of 14 months including all available treatments. Low grade meningiomas, usually benign grade I tumors, are in most cases cured by surgical resection. However despite their benign appearance grade I meningiomas can, without any histopathological signs, in some cases develop bone invasive growth and become lethal. Thus, it is necessary to improve conventional treatment modalities, develop new treatment strategies and improve the knowledge regarding the basic pathophysiology in the classification and treatment of brain tumors. In this thesis, both proteomics and metabolomics have been applied in the search for biomarkers or biomarker patterns in two different types of brain tumors, gliomas and meningiomas. Proteomic studies were carried out mainly by surface enhanced laser desorption ionization time of flight mass spectrometry (SELDI-TOF-MS). In one of the studies, isobaric tags for relative and absolute quantitation (iTRAQ) labeling in combination with high-performance liquid chromatography (HPLC) was used for protein detection and identification. For metabolomics, gas-chromatography time-of-flight mass spectrometry (GC-TOF-MS) has been the main platform used throughout this work for generation of robust global metabolite profiles in tissue, blood and cell cultures. To deal with the complexity of the generated data, and to be able to extract relevant biomarker patters or latent biomarkers, for interpretation, prediction and prognosis, bioinformatic strategies based on chemometrics were applied throughout the studies of the thesis. In summary, we detected differentiating protein profiles between invasive and non-invasive meningiomas, in both fibrous and meningothelial tumors. Furthermore, in a different study we discovered treatment induce protein pattern changes in a rat glioma model treated with an angiogenesis inhibitor. We identified a cluster of proteins linked to angiogenesis. One of those proteins, HSP90, was found elevated in relation to treatment in tumors, following ELISA validation. An interesting observation in a separate study was that it was possible to detect metabolite pattern changes in the serum metabolome, as an effect of treatment with radiotherapy, and that these pattern changes differed between different patients, highlighting a possibility for monitoring individual treatment response.  In the fourth study of this work, we investigated tissue and serum from glioma patients that revealed differences in the metabolome between glioblastoma and oligodendroglioma, as well as between oligodendroglioma grade II and grade III. In addition, we discovered metabolite patterns associated to survival in both glioblastoma and oligodendroglioma. In our final work, we identified metabolite pattern differences between cell lines from a subgroup of glioblastomas lacking argininosuccinate synthetase (ASS1) expression, (ASS1 negative glioblastomas), making them auxotrophic for arginine, a metabolite required for tumor growth and proliferation, as compared to glioblastomas with normal ASS1 expression (ASS1 positive). From the identified metabolite pattern differences we could verify the hypothesized alterations in the arginine biosynthetic pathway. We also identified additional interesting metabolites that may provide clues for future diagnostics and treatments. Finally, we were able to verify the specific treatment effect of ASS1 negative cells by means of arginine deprivation on a metabolic level.

This thesis shows that p53-deficient mouse glioma brain tumour stem cells (BTSCs), which fail to express pluripotency factors, can be reprogrammed with specific transcription factors to generate iPS cell lines (GNS-iPS) expressing endogenous pluripotency factors (Nanog, Oct4, and Rex1). GNS-iPS cell lines formed embryoid bodies (EBs) in vitro and undifferentiated growths in vivo that phenotypically did not resemble tumours derived from non-reprogrammed BTSCs. EBs formed from one GNS-iPS cell line expressed markers of mesoderm, endoderm, and ectoderm. Tumours produced from GNS-iPS cells had reduced astrocytic marker (GFAP) expression compared to those generated from control iPS cell lines or non-reprogrammed BTSCs. Preliminary results suggest that the reprogrammed cells can be re-differentiated into cells that show neural precursor phenotype. These findings suggest that BTSCs can acquire aspects of the pluripotent state with a defined set of transcription factors, opening the door for further exploration of reprogramming strategies to attenuate the cancer phenotype.

This thesis shows that p53-deficient mouse glioma brain tumour stem cells (BTSCs), which fail to express pluripotency factors, can be reprogrammed with specific transcription factors to generate iPS cell lines (GNS-iPS) expressing endogenous pluripotency factors (Nanog, Oct4, and Rex1). GNS-iPS cell lines formed embryoid bodies (EBs) in vitro and undifferentiated growths in vivo that phenotypically did not resemble tumours derived from non-reprogrammed BTSCs. EBs formed from one GNS-iPS cell line expressed markers of mesoderm, endoderm, and ectoderm. Tumours produced from GNS-iPS cells had reduced astrocytic marker (GFAP) expression compared to those generated from control iPS cell lines or non-reprogrammed BTSCs. Preliminary results suggest that the reprogrammed cells can be re-differentiated into cells that show neural precursor phenotype. These findings suggest that BTSCs can acquire aspects of the pluripotent state with a defined set of transcription factors, opening the door for further exploration of reprogramming strategies to attenuate the cancer phenotype.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2007. / Title from first page of PDF file (viewed Oct. 30, 2007). Includes bibliographical references (p. 165-174).

BACKGROUND: Gliomas are the most common primary brain malignancy, with more than 16,000 patients diagnosed every year (Ostrom, et al., 2015). Outcomes vary widely depending on tumor grade and treatment, and have been steadily improving with the advent of new therapeutics. Glioma patients frequently undergo chemotherapy to remove residual tumor after surgery, and many of these cytotoxic therapies are known to affect rapidly dividing cells such as ovarian follicles (Vassilakopoulou et al., 2016). The negative effects of chemotherapy on fertility have been demonstrated in patients with breast and colorectal cancer (Bines, et al., 1996; Avastin Prescribing Information). Additionally, infertility has been linked with decreased quality of life, primarily in women (O’Moore et al., 1983; Greil, 1997). Fertility treatments are available for women undergoing cancer treatment, however it is unknown whether these treatments are routinely discussed with glioma patients before initiating chemotherapy. OBJECTIVE: The primary goal of this study was to assess whether female glioma patients are being effectively counselled on their possible loss of fertility and their choices for fertility treatment prior to beginning chemotherapy. To this end, it was also important to understand the barriers preventing patients from obtaining information related to their fertility. Another principle goal of this study was to describe the effects of chemotherapy on a sample of women with glioma. Finally, this study sought to understand the priorities of women with glioma in regards to family planning, and to address these priorities in the context of a comprehensive fertility preservation discussion. METHODS: To assess these endpoints, a survey was designed and delivered to patients being treated at the Neuro-oncology clinic of the University of California, San Francisco. Eligible candidates were identified prior to a clinic visit, and patients were asked whether they would like to participate in the survey. Consenting patients then completed the survey at home or in the clinic. Seventy two women completed the survey. Data was analyzed using STATA Software Version 10.0. RESULTS: Analysis of the survey results showed that only 35% of women receiving chemotherapy reported having a discussion regarding fertility preservation prior to beginning treatment. Of those who reported having this discussion, only 80% were aware that chemotherapy could negatively affect their fertility. Many women reported that while fertility preservation was not important to them at the time of diagnosis, it was a priority for them at the time of survey completion. Most women surveyed expressed a desire to have a fertility preservation discussion with a reproductive specialist. CONCLUSIONS: The data obtained in this study suggest a lack of understanding of the negative effects of chemotherapy which may be addressed with a more comprehensive fertility discussion with glioma patients prior to beginning treatment. Although interest in having children tends to decrease after cancer treatment, the majority of respondents still report wanting a child after treatment. The priorities of women in the study reflect a concern for the health of their future offspring which may be best addressed prior to beginning treatment in order to increase their chances of conceiving at a later date.

Gliomas são os mais comuns e devastadores tumores primários do sistema nervoso central. Os nucleotídeos da adenina são moléculas sinalizadoras no meio extracelular, envolvidas em importantes condições fisiológicas e patológicas. O ATP, neurotransmissor excitatório, e a adenosina, neuromoduladora, entre outros efeitos, podem induzir proliferação celular em linhagens de gliomas. Os eventos induzidos pelos nucleotídeos extracelulares são controlados pela ação das E-NTPDases, que hidrolisam o ATP até adenosina extracelularmente. Recentes estudos epidemiológicos têm sugerido que os flavonóides derivados da dieta, em particular a quercetina, apresentam um papel benéfico em prevenir ou inibir a tumorigênese. Assim, primeiramente nós avaliamos o efeito antiproliferativo da quercetina em linhagem de glioma humano U138MG. O estudo demonstrou que este flavonóide induziu em cultura de gliomas: (1) diminuição da proliferação e da viabilidade celular; (2) morte celular via necrose e apoptose; (3) parada no ciclo celular na fase G2 e (4) diminuição do índice mitótico. Além disso, nós demonstramos que a quercetina, enquanto promoveu regressão tumoral, protegeu culturas organotípicas hipocampais do dano isquêmico. Em conjunto, esses dados sugerem que a quercetina exibe efeitos antiproliferativos direcionados para as células tumorais e reduzida citotoxicidade para células normais, características altamente desejáveis na quimioterapia. Dados do nosso laboratório demonstram que o metabolismo extracelular das purinas encontra-se alterado em linhagens de gliomas com relação a culturas de astrócitos, sugerindo que mudanças no sistema purinérgico podem ser uma característica dos gliomas que potencialmente podem contribuir para o seu fenótipo de malignidade. Assim, o passo seguinte desse trabalho foi investigar o perfil de secreção dos derivados da adenina, o metabolismo extracelular do AMP e a ação da quercetina sobre o sistema purinérgico. As culturas de glioma apresentaram secreção de ATP, o qual foi detectado em maiores níveis com relação as outras moléculas avaliadas, ADP, AMP, adenosina e inosina. O AMP extracelular foi eficientemente metabolizado pelos gliomas, demonstrando uma ecto-5’-NT/CD73 muito ativa. Adicionalmente, quercetina interagiu com o sistema purinérgico, inibindo não-competitivamente a atividade da ecto-5’-NT/CD73 e modulando negativamente a sua expressão. Nós sugerimos que a inibição da atividade da ecto-5’-NT/CD73 pode resultar em um decréscimo na disponibilidade de adenosina extracelular, uma promotora tumoral. Tal efeito pode estar correlacionado com a inibição da proliferação promovida pela quercetina nessa linhagem de glioma. Nossos dados sugerem que a quercetina pode ter uma função importante na inibição da proliferação dos gliomas, atuando em diferentes vias de sinalização, incluindo o sistema purinérgico. Assim, esse estudo abre novas perspectivas para as potenciais aplicações dos flavonóides na prevenção e tratamento de tumores cerebrais.

O antioxidante resveratrol, uma fitoalexina encontrada, principalmente em uvas e também em quantidades significativas em vários tipos de vinho tinto, é um promissor produto natural, com atividades antitumoral, cardioprotetora e neuroprotetora. O objetivo do presente estudo foi investigar em células de glioma C6 o efeito do resveratrol sobre proliferação celular e alguns parâmetros específicos relacionados a astrócitos (captação de glutamato, glutamina sintetase e secreção de S100B), comumente associados com o papel protetor dessas células. Além disso, foi investigado o efeito genoprotetor do resveratrol em condições de estresse oxidativo induzido por peróxido de hidrogênio (H2O2) em células C6. A proliferação celular foi significativamente reduzida após tratamento com 100 e 250 μM de resveratrol. Uma rápida incubação com resveratrol (1 h) induziu um aumento linear na captação de glutamato e na atividade da glutamina sintetase. O dano ao DNA foi medido pelo ensaio Cometa. Para investigar os efeitos do resveratrol sobre o dano ao DNA induzido por H2O2 utilizou-se dois modelos de indução de estresse oxidativo. A mudança de parâmetros gliais pode contribuir para o papel protetor de astrócitos em condições de injúria cerebral, reforçando o uso deste composto no arsenal terapêutico contra doenças neurodegenerativas e desordens isquêmicas. Resveratrol também foi capaz de prevenir o dano oxidativo ao DNA celular, provavelmente, devido a suas propriedades antioxidantes, isto pode ser importante para proteger o DNA em doenças relacionadas com estresse oxidativo. / The antioxidant resveratrol, a phytoalexin found mainly in grapes and also substantial amounts in several types of red wine, is a promising natural product with anticancer, cardio-protective and neuroprotective activities. The objective of the present study was to investigate in C6 glioma cells, the effect of resveratrol on cell proliferation, cell death and some specific parameters of astrocyte activity (glutamate uptake, glutamine synthetase and secretion of S100B) commonly associated with the protective role of these cells. Furthermore, it was investigated the genoprotective effects of resveratrol under conditions of oxidative stress induced by hydrogen peroxide (H2O2) in C6 cells. Cell proliferation was significantly decreased following treatment with 100 and 250 μM resveratrol. Short-term (1 h) of resveratrol exposure induced a linear increase in glutamate uptake and in glutamine synthetase activity. DNA damage was assessed by the comet assay. For investigate the effects of resveratrol against oxidative stress induced by H2O2 on DNA damage, two models of oxidative stress induction were used. Changes in glial activities can contribute to the protective role of astrocytes in brain injury conditions, reinforcing the use of this compound in the therapeutic arsenal against neurodegenerative diseases and ischemic disorders. Resveratrol was able to prevent oxidative damage to cellular DNA, probably, due to its antioxidant properties, it may be important in diseases for protecting against DNA damage through oxidative stress.