Mutantes do receptor B2 de cininas : análise da interação droga-receptor e sua aplicabilidade no estudo do angioedema hereditário

• Autor(a) principal: Silva, Rafael Filippelli da [UNIFESP]
• Data de Publicação: 2017
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Repositório Institucional da UNIFESP
• dARK ID: ark:/48912/0013000019pjk
• Texto Completo: https://sucupira.capes.gov.br/sucupira/public/consultas/coleta/trabalhoConclusao/viewTrabalhoConclusao.jsf?popup=true&id_trabalho=5170326; http://repositorio.unifesp.br/handle/11600/50524
• topic: Hereditary angioedema; Bradykinin; Receiver B; Mutants; Biased aqonismo; Angioedema hereditário; Bradicinina; Receptor B; Mutantes; Aqonismo tendencioso

Introduction: bradykinin (8K) is an important pressure regulating peptide arterial, nociception, inflammation and is associated with the pathophysiology of angioedema (HAE). The main clinical aspects of HAE are mediated by 8K, which exercise by the activation of receptor 82 (82R), a member of the family of G protein coupled receptors (GPCR). The objective of the present study was to evaluate if 82R mutations found in HAE patients could alter the profile pharmacological effect of 8K signal transduction, leading to relevant for the pathophysiology of the disease. Materials and methods: we use G-and-3-arrestin biosensors to perform 8RET analyzes and evaluate functional profiles of mutant 82Rs. All the experiments were carried out in transiently transfected HEK 293T cells, either with the wild-type or with the human 82R mutants. Competitive binding assays using 8K radiolabeled (3H-8K) were performed to estimate the 8K affinity for mutant receptors. The 8RET assays, used to evaluate the activation of G protein and the recruitment of β-arrestin 1 and 2, were performed using a method adapted from that described by Quoyer et al. (2013). Results: the binding assays showed that 8K retains a similar affinity at mutant receptors R14C, W344C, G354E and V376M, when compared to the wild-type receptor. In analysis, 8K triggered the coupling of G protein and mobilization intracellular calcium with similar potency and efficacy at R14C and G354E compared to WT. On the other hand, these receptors were not able to to recruit! 3-arrestinas. In the W344C and V376M mutants, the potency and efficacy of 8K was reduced on activation of all of the signaling pathways tested. Conclusions: These findings provide strong evidence for a biased agonist profile endogenous 8K G protein by activating mutants R14C and G354E 82R identified in patients with HAE. On the other hand, mutants W344C and V376M showed a decrease in the pharmacological responses induced by 8K. We believe that the current data contribute to revealing the mechanisms molecules involved in HAE and thus opening up new perspectives and approaches to study and treat this disease.