Análise da interação de cAmbly na linhagem T98G de câncer cerebral Glioblastoma Multiforme
Arquivos
Data
2023-10-10
Autores
Garavelli, Graciana Yokota 
Orientadores
Chudzinski-Tavassi, Ana Marisa
Tipo
Dissertação de mestrado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
Objetivo: Identificar através da técnica de quimioproteômica possíveis alvos moleculares do peptídeo cAmbly, derivado da proteína recombinante denominada Amblyomin-X, no modelo de Glioblastoma Multiforme. Métodos: Como abordagem principal foi utilizada a técnica de quimioproteômica. Os ensaios envolvem além da padronização da técnica, avaliações prévias de achados relacionados as proteínas identificadas, focando em avaliar possíveis alterações em metabolismo mitocondrial bem como a colocalização da molécula na organela. Avaliação do ciclo celular e proliferação celular foram realizados. Também foi analisada a internalização de cAmbly. Resultados: Foram identificadas o total de 25 proteínas em três experimentos distintos, das quais foram consideradas 15 como possíveis alvos de cAmbly. Dentre elas, destacam-se as proteínas mitocondriais envolvidas no metabolismo energético, com ênfase na piruvato carboxilase que além de ser a principal enzima envolvida no efeito Warburg, possivelmente tem interação com outras vias de interesse. Também se destacam a enzima Nicotinamida-N-metiltransferase e outras proteínas relacionadas ao microambiente tumoral, ao sistema imune e a via de inibição de proteassoma como alvos de interesse. Efeitos em proliferação celular e alterações significativas no ciclo celular não foram observados, assim como mudanças na atividade metabólica mitocondrial. Tais achados indicam que apesar de cAmbly se ligar a possíveis alvos, não se observa efeitos nos contextos estudados, sendo essa uma característica de interesse para sua atuação como cargo delivery. Por fim, houve colocalização de cAmbly na mitocôndria. Conclusões: Os resultados obtidos evidenciaram a internalização de cAmbly, e no contexto estudado não houve alterações metabólicas significativas, embora alguns dos possíveis alvos encontrados estejam envolvidos no processo de metabolismo energético. Tal achado corrobora com os resultados de estudos anteriores sobre cAmbly atuar como molécula cargo delivery. Ademais, é necessário que sejam avaliados experimentalmente os possíveis alvos envolvidos com o microambiente tumoral e a atuação no sistema imune.
Objective: Identity with the chemoproteomics technique, new molecular targets of the cAmbly peptide, a fragment of the recombinant protein named Amblyomin-X, using the Glioblastoma Multiform model. Methods: Chemoproteomics technique was the main approach used in this study. Beyond establishing a standard protocol, additional experiments are performed to investigate the findings, focusing on previous evaluations of proteins that interact with cellular metabolism and the presence of cAmbly in mitochondria. Cell cycle analysis and changes in cell proliferation are evaluated. The internalization of cAmbly was also observed. Results: A total of twenty-five proteins were identified in three different experiments, fifteen of them were considered possible cAmbly targets. Some of the identified mitochondrial proteins participate in energy metabolism, with emphasis on pyruvate carboxylase, the main enzyme involved in the Warburg effect and that interacts with other pathways of interest. The enzyme Nicotinamide-N-methyltransferase and other proteins related to the tumor microenvironment, immune system, and inhibition of proteasome pathway, can also be targets of interest. No effects on cell proliferation, cell cycle arrest and changes in mitochondrial metabolism are observed. Such findings indicate that cAmbly binds to targets, but no effects are observed with this experimental approach, which is a characteristic of interest for a molecule with cargo delivery. Finally, the experiments showed the colocalization of cAmbly in the mitochondria. Conclusions: The results demonstrate the internalization of cAmbly without significant changes in metabolism, although we see probable molecular targets interact with energy metabolism. These findings corroborate previous results about the cargo delivery function of the peptide. More studies are necessary to investigate the interaction of cAmbly in the tumor microenvironment and/or the immune system.
Objective: Identity with the chemoproteomics technique, new molecular targets of the cAmbly peptide, a fragment of the recombinant protein named Amblyomin-X, using the Glioblastoma Multiform model. Methods: Chemoproteomics technique was the main approach used in this study. Beyond establishing a standard protocol, additional experiments are performed to investigate the findings, focusing on previous evaluations of proteins that interact with cellular metabolism and the presence of cAmbly in mitochondria. Cell cycle analysis and changes in cell proliferation are evaluated. The internalization of cAmbly was also observed. Results: A total of twenty-five proteins were identified in three different experiments, fifteen of them were considered possible cAmbly targets. Some of the identified mitochondrial proteins participate in energy metabolism, with emphasis on pyruvate carboxylase, the main enzyme involved in the Warburg effect and that interacts with other pathways of interest. The enzyme Nicotinamide-N-methyltransferase and other proteins related to the tumor microenvironment, immune system, and inhibition of proteasome pathway, can also be targets of interest. No effects on cell proliferation, cell cycle arrest and changes in mitochondrial metabolism are observed. Such findings indicate that cAmbly binds to targets, but no effects are observed with this experimental approach, which is a characteristic of interest for a molecule with cargo delivery. Finally, the experiments showed the colocalization of cAmbly in the mitochondria. Conclusions: The results demonstrate the internalization of cAmbly without significant changes in metabolism, although we see probable molecular targets interact with energy metabolism. These findings corroborate previous results about the cargo delivery function of the peptide. More studies are necessary to investigate the interaction of cAmbly in the tumor microenvironment and/or the immune system.
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Citação
GARAVELLI, Graciana Yokota. Análise da interação de cAmbly na linhagem T98G de câncer cerebral glioblastoma multiforme. 2023. 110 p. Dissertação (Mestrado em Biologia Molecular) - Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, 2023.