Estudo do papel da proteína inibidora do ciclo celular p21waf1/cip1 e seus possíveis mecanismos reguladores na progressão do melanoma
Data
2014
Autores
Colaneri, Gabriela Nana 
Orientadores
Jasiulionis, Miriam Galvonas
Tipo
Dissertação de mestrado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
O cancer pode ser considerado uma doenca do ciclo celular. Descrito inicialmente como potente inibidor de quinases-dependentes de ciclina e, portanto, regulador negativo do ciclo celular, p21waf1/cip1 e uma proteina cuja expressao encontra-se frequentemente alterada em tumores. Paradoxalmente, o aumento de sua expressao tem sido correlacionado com canceres mais agressivos e pior prognostico. Isso, pois p21waf1/cip1 se encaixa na definicao de odualidade antagonisticao, usada para proteinas que possuem funcoes contrarias, dependendo da localizacao e contexto celular. Em nosso laboratorio, foi visto que celulas murinas de melanoma metastatico expressam altos niveis de p21waf1/cip1. O melanoma e um cancer cuja incidencia aumentou nos ultimos 50 anos e, embora seja o menos incidente, e o mais agressivo dos canceres de pele. Sua forma metastatica e altamente letal, chegando a 90% dos casos. Assim, o objetivo deste projeto foi investigar o papel de p21waf1/cip1 ao longo da progressao do melanoma e possivel contribuicao para aquisicao do fenotipo metastatico. Para tal, foi utilizado um modelo murino de genese do melanoma, no qual diferentes linhagens foram obtidas apos submeter melanocitos melan-a a condicoes de estresse sustentado. Os resultados demonstram que a superexpressao de p21waf1/cip1 citoplasmatico aumenta a sobrevivencia de celulas de melanoma metastaticas apos tratamento com dacarbazina, um efeito revertido por seu silenciamento via shRNA. Tambem identificamos altos niveis da expressao de Akt fosforilada e p21waf1/cip1fosforilado no residuo Thr145, responsaveis por induzir a retencao de p21waf1/cip1 no citoplasma. Usando Wortmannin, um inibidor de PI3K, que previne a ativacao de Akt, pode-se restaurar a localizacao nuclear de p21waf1/cip1. Por fim, identificamos altos niveis de p21waf1/cip1 e fosfo- p21 em linhagens de melanoma metastaticas derivadas de pacientes. Os resultados obtidos demonstram que o uso de p21waf1/cip1como alvo terapeutico deve ser encarado com cautela, tendo em vista seu papel antagonico. Assim, espera-se contribuir para a compreensao do papel desta proteina em melanoma, melhorarando a eficacia dos tratamentos para este tumor.
Cancer can be considered a cell cycle disease. Initially described as a potent cyclindependent kinase inhibitor and thus a negative regulator of cell cycle, p21waf1/cip1 is a protein whose expression levels are often altered in tumor cells. Paradoxically, the increase in its expression has been correlated with aggressive cancers and poor prognosis.This proteins fits the definition of “antagonistic duality” , used to define proteins that have opposing effects within the cell, depending on localization and cell context. In our laboratory, it was shown that murine metastatic melanoma cells express high levels of p21waf1/cip1. The incidence of melanoma has increased worldwide over the past 50 years. Although it is the least incident of skin cancers, it is the most aggressive type of cancer in this organ. Its metastatic form is highly lethal, reaching 90% of cases. The aim of this project was to investigate the role of p21waf1/cip1 throughout melanoma progression and contribution to the acquisition of metastatic phenotype. For this purpose, a murine model of melanoma genesis, in which different cell lines were obtained after submitting melana melanocytes to sustained stress conditions was used. The results demonstrate that cytoplasmic p21waf1/cip1 overexpression increases metastatic melanoma cells survival after dacarbazine treatment, which can be reversed by its silencing via shRNA. We aso identified high levels of phosphorylated Akt and phosphorylated p21waf1/cip1, responsible to induce p21waf1/cip1 retention in the cytoplasm. Using Wortmannin, an inhibitor of PI3K, which prevents the activation of Akt, we restored the nuclear localization of p21waf1/cip1 in metastatic melanoma lineage. Finally, we have identified high levels of phosphop21waf1/cip1 and p21waf1/cip1 in patientsderived metastatic melanoma lineages. The results of this study show that the use of p21waf1/cip1as a therapeutic target should be viewed with caution, given its antagonistic role. Thus, it is expected to contribute to the understanding of the role of p21waf1/cip1 in melanoma progression and thereby improve the effectiveness of treatments for this tumor.
Cancer can be considered a cell cycle disease. Initially described as a potent cyclindependent kinase inhibitor and thus a negative regulator of cell cycle, p21waf1/cip1 is a protein whose expression levels are often altered in tumor cells. Paradoxically, the increase in its expression has been correlated with aggressive cancers and poor prognosis.This proteins fits the definition of “antagonistic duality” , used to define proteins that have opposing effects within the cell, depending on localization and cell context. In our laboratory, it was shown that murine metastatic melanoma cells express high levels of p21waf1/cip1. The incidence of melanoma has increased worldwide over the past 50 years. Although it is the least incident of skin cancers, it is the most aggressive type of cancer in this organ. Its metastatic form is highly lethal, reaching 90% of cases. The aim of this project was to investigate the role of p21waf1/cip1 throughout melanoma progression and contribution to the acquisition of metastatic phenotype. For this purpose, a murine model of melanoma genesis, in which different cell lines were obtained after submitting melana melanocytes to sustained stress conditions was used. The results demonstrate that cytoplasmic p21waf1/cip1 overexpression increases metastatic melanoma cells survival after dacarbazine treatment, which can be reversed by its silencing via shRNA. We aso identified high levels of phosphorylated Akt and phosphorylated p21waf1/cip1, responsible to induce p21waf1/cip1 retention in the cytoplasm. Using Wortmannin, an inhibitor of PI3K, which prevents the activation of Akt, we restored the nuclear localization of p21waf1/cip1 in metastatic melanoma lineage. Finally, we have identified high levels of phosphop21waf1/cip1 and p21waf1/cip1 in patientsderived metastatic melanoma lineages. The results of this study show that the use of p21waf1/cip1as a therapeutic target should be viewed with caution, given its antagonistic role. Thus, it is expected to contribute to the understanding of the role of p21waf1/cip1 in melanoma progression and thereby improve the effectiveness of treatments for this tumor.
Descrição
Citação
COLANERI, Gabriela Nana. Estudo do papel da proteína inibidora do ciclo celular p21waf1/cip1 e seus possíveis mecanismos reguladores na progressão do melanoma. 2014. 102 f. Dissertação (Mestrado em Microbiologia, Imunologia e Parasitologia) – Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, 2014.