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Authors
Abstract(s)
O glioblastoma multiforme (GBM) é o tumor maligno, do cérebro e do sistema nervoso
central, mais comum. A remoção cirúrgica destes tumores, seguida de radioterapia e
quimioterapia com temozolomida, constituem a abordagem terapêutica mais usada no
tratamento do GBM. No entanto, devido à sua agressividade, a esperança média de vida
destes doentes é de, aproximadamente, 15 a 18 meses após o diagnóstico e, mesmo em
condições favoráveis, a maioria dos pacientes não ultrapassa os 2 anos. Assim, um
conhecimento mais aprofundado deste tipo de tumor e a identificação de alvos
terapêuticos alternativos são necessários para melhorar o prognóstico e as taxas de
sobrevida dos pacientes. O GBM tem características metabólicas particulares. Tal como
na maioria das células cancerígenas, a glicólise prevalece sobre a fosforilação oxidativa
como principal fonte de energia, mesmo quando há oxigénio disponível. Essa alteração
metabólica designada por Efeito de Warburg, favorece a grande demanda de
nucleótidos, aminoácidos e lípidos essenciais para a proliferação celular. Assim, a
reversão deste efeito surge como uma abordagem terapêutica promissora no tratamento
do GBM. Dados recentes, obtidos a partir de ensaios de microarrays e sequenciação de
RNA de GBM, sugerem que estes tumores expressam o recetor olfativo 51E2 (OR51E2),
sendo o lactato, um dos produtos da glicólise, um dos ligandos conhecidos deste recetor.
Uma vez que o OR51E2, tem sido implicado na modulação da proliferação e invasão
noutros tumores, este trabalho pretende investigar a possibilidade do OR51E2 funcionar
como sensor de lactato no microambiente tumoral e, desta forma, modelar o metabolismo
e reverter o Efeito de Warburg. Os resultados deste estudo confirmaram a expressão do
mRNA e da proteína do OR51E2 numa linha de astrócitos normais HASTR/ci35 e em três
linhas de GBM, U-87MG, SNB-19 e U-373MG. Procedeu-se também à comparação dos
níveis de expressão do OR51E2 em amostras humanas de astrocitomas de diferentes
graus (I-IV). Verificou-se que o OR51E2 é expresso, predominantemente, no citosol nas
linhas celulares SNB-19 e U-373MG. Assim, a linha SNB-19 foi usada para os restantes
ensaios. De acordo com o que seria expectável, as SNB-19, quando sujeitas à privação de
oxigénio e glucose, sofrem uma diminuição na viabilidade celular. Para avaliar o possível
envolvimento na resposta destas células à hipoxia, o OR51E2 foi silenciado com siRNA.
Os resultados obtidos mostraram que o silenciamento do recetor não afeta
significativamente a viabilidade celular em condições normais de oxigénio (HG e LG
normóxia). Resultados semelhantes foram obtidos sob privação de oxigénio (HG e LG
hipóxia). Doseou-se também o lactato extracelular, e não se observaram diferenças
significativas entre condições normais de oxigénio (HG e LG normóxia) e de privação de oxigénio (HG e LG hipóxia). Face aos resultados obtidos concluímos que o OR51E2 não
medeia a resposta à hipoxia nesta linha celular de GBM.
Glioblastoma multiforme (GBM) is the most common malignant tumor of the brain and central nervous system. Surgical removal of these tumors, followed by radiotherapy and chemotherapy with temozolomide, are the most used therapeutic approach in the treatment of GBM. However, due to its aggressiveness, the average life expectancy of these patients is approximately 15 to 18 months after diagnosis and, even under favorable conditions, most patients do not exceed 2 years. Thus, a deeper knowledge of this type of tumor and the identification of alternative therapeutic targets are needed to improve the prognosis and survival rates of patients. GBM has particular metabolic characteristics. As in most cancer cells, glycolysis prevails over oxidative phosphorylation as the main source of energy, even when oxygen is available. This metabolic alteration, called the Warburg Effect, favors the great demand for nucleotides, amino acids and lipids essential for cell proliferation. Thus, the reversal of this effect appears as a promising therapeutic approach in the treatment of GBM. Recent data, obtained from microarray assays and GBM RNA sequencing, suggest that these tumors express the olfactory receptor 51E2 (OR51E2), with lactate, one of the products of glycolysis, one of the known ligands of this receptor. Since OR51E2 has been implicated in the modulation of proliferation and invasion in other tumors, this work intends to investigate the possibility of OR51E2 functioning as a lactate sensor in the tumor microenvironment and, in this way, modeling the metabolism and reversing the Warburg Effect. The results of this study confirmed the expression of OR51E2 mRNA and protein in a HASTR/ci35 normal astrocyte line and in three GBM lines, U-87MG, SNB-19 and U-373MG. We also compared the expression levels of OR51E2 in human samples of astrocytomas of different grades (I-IV). It was found that OR51E2 is predominantly expressed in the cytosol in the SNB-19 and U-373MG cell lines. Thus, the SNB-19 line was used for the remaining tests. As expected, SNB-19, when subjected to oxygen and glucose deprivation, suffer a decrease in cell viability. To assess the possible involvement in the response of these cells to hypoxia, OR51E2 was silenced with siRNA. The results obtained showed that receptor silencing does not significantly affect cell viability under normal oxygen conditions (HG and LG normoxia). Similar results were obtained under oxygen deprivation (HG and LG hypoxia). Extracellular lactate was also measured, and no significant differences were observed between normal oxygen conditions (HG and LG normoxia) and oxygen deprivation (HG and LG hypoxia). In view of the results obtained, we conclude that OR51E2 does not mediate the response to hypoxia in this GBM cell line.
Glioblastoma multiforme (GBM) is the most common malignant tumor of the brain and central nervous system. Surgical removal of these tumors, followed by radiotherapy and chemotherapy with temozolomide, are the most used therapeutic approach in the treatment of GBM. However, due to its aggressiveness, the average life expectancy of these patients is approximately 15 to 18 months after diagnosis and, even under favorable conditions, most patients do not exceed 2 years. Thus, a deeper knowledge of this type of tumor and the identification of alternative therapeutic targets are needed to improve the prognosis and survival rates of patients. GBM has particular metabolic characteristics. As in most cancer cells, glycolysis prevails over oxidative phosphorylation as the main source of energy, even when oxygen is available. This metabolic alteration, called the Warburg Effect, favors the great demand for nucleotides, amino acids and lipids essential for cell proliferation. Thus, the reversal of this effect appears as a promising therapeutic approach in the treatment of GBM. Recent data, obtained from microarray assays and GBM RNA sequencing, suggest that these tumors express the olfactory receptor 51E2 (OR51E2), with lactate, one of the products of glycolysis, one of the known ligands of this receptor. Since OR51E2 has been implicated in the modulation of proliferation and invasion in other tumors, this work intends to investigate the possibility of OR51E2 functioning as a lactate sensor in the tumor microenvironment and, in this way, modeling the metabolism and reversing the Warburg Effect. The results of this study confirmed the expression of OR51E2 mRNA and protein in a HASTR/ci35 normal astrocyte line and in three GBM lines, U-87MG, SNB-19 and U-373MG. We also compared the expression levels of OR51E2 in human samples of astrocytomas of different grades (I-IV). It was found that OR51E2 is predominantly expressed in the cytosol in the SNB-19 and U-373MG cell lines. Thus, the SNB-19 line was used for the remaining tests. As expected, SNB-19, when subjected to oxygen and glucose deprivation, suffer a decrease in cell viability. To assess the possible involvement in the response of these cells to hypoxia, OR51E2 was silenced with siRNA. The results obtained showed that receptor silencing does not significantly affect cell viability under normal oxygen conditions (HG and LG normoxia). Similar results were obtained under oxygen deprivation (HG and LG hypoxia). Extracellular lactate was also measured, and no significant differences were observed between normal oxygen conditions (HG and LG normoxia) and oxygen deprivation (HG and LG hypoxia). In view of the results obtained, we conclude that OR51E2 does not mediate the response to hypoxia in this GBM cell line.
Description
Keywords
Glioblastoma Multiforme Hipóxia Metabolismo da Glucose Or51e2 Proliferação Celular