Explora??o in silico da a??o farmacol?gica da Hexadecilfosfocolina em ameba de vida livre Naegleria fowleri

Autor:	Oliveira, Victor Valadares de
Přispěvatelé:	Dornas, F?bio Pio, Oliveira, Eduardo de Jesus, Oliveira, Danilo Bretas de, Universidade Federal dos Vales do Jequitinhonha e Mucuri (UFVJM)
Jazyk:	portugalština
Rok vydání:	2022
Předmět:	Mecanismos moleculares de a??o farmacol?gica Molecular mechanisms of pharmacological action Central nervous system protozoal infections Amebicidas Simula??o de acoplamento molecular Amebicides Molecular docking simulation Naegleria fowleri Infec??es protozo?rias do sistema nervoso central
Zdroj:	Repositório Institucional da UFVJM Universidade Federal dos Vales do Jequitinhonha e Mucuri (UFVJM) instacron:UFVJM
Popis:	O presente trabalho foi realizado com apoio da Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior ? Brasil (CAPES) ? C?digo de Financiamento 001. Submitted by Jos? Henrique Henrique (jose.neves@ufvjm.edu.br) on 2022-10-27T20:12:16Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) victor_valadares_oliveira.pdf: 5857009 bytes, checksum: 1328f8b3b833857ebeebda7b09ab1171 (MD5) Approved for entry into archive by Jos? Henrique Henrique (jose.neves@ufvjm.edu.br) on 2022-10-27T20:26:03Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) victor_valadares_oliveira.pdf: 5857009 bytes, checksum: 1328f8b3b833857ebeebda7b09ab1171 (MD5) Made available in DSpace on 2022-10-27T20:26:03Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) victor_valadares_oliveira.pdf: 5857009 bytes, checksum: 1328f8b3b833857ebeebda7b09ab1171 (MD5) Previous issue date: 2022 Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (CAPES) A ameba de vida livre Naegleria fowleri ? o agente etiol?gico da meningoencefalite amebiana prim?ria. Uma infec??o do sistema nervoso central de evolu??o fulminante com taxa de mortalidade de aproximadamente 97%. A hexadecilfosfocolina ? um f?rmaco utilizado no tratamento dessa doen?a, um an?logo de fosfolip?dio com propriedades anti-tumoral e anti-protozo?ria. Apesar de amebicida, o mecanismo molecular subjacente ao efeito citop?tico em Naegleria fowleri n?o ? conhecido. An?lises in silico foram feitas a fim de esclarecer isso. Enzimas de v?rios organismos cujas atividades catal?ticas s?o afetadas por intera??o molecular direta com a hexadecilfosfocolina foram identificadas em revis?o bibliogr?fica. Tais enzimas foram utilizadas na detec??o de enzimas de Naegleria fowleri atrav?s de buscas por alinhamento de sequ?ncias na base de dados AMOEBADB. As estruturas tridimensionais das enzimas recuperadas foram obtidas diretamente do PDB ou do reposit?rio do Swiss-Model. Prote?nas sem modelos prontos e n?o resolvidas experimentalmente foram modeladas pelo m?todo de homologia de sequ?ncias. Os modelos foram validados atrav?s das m?tricas GMQE e QMEANDisCO do Swiss-Model, al?m dos softwares Errat, Prove, Verify 3D e PROCHECK. O Autodock Vina foi empregado em simula??es de ancoramento molecular entre as enzimas detectadas e a hexadecilfosfocolina, empregando enzimas r?gidas e ligante flex?vel. As energias de afinidade das melhores poses de liga??o variaram entre -6.0 e -4.5 kcal/mol. Foram realizadas infer?ncias do mecanismo de a??o citop?tica baseando-se na avalia??o das redes metab?licas hipoteticamente envolvidas, todas dispon?veis nas bases KEEG e MetaCyC. Atrav?s das an?lises realizadas sugere-se que a hexadecilfosfocolina possa exercer efeito citop?tico sobre c?lulas de Naegleria fowleri ao inibir principalmente as enzimas CTP: fosfocolina-citidililtransferase (E.C. Number: 2.7.7.15) e Fosfatidilinositol 3-cinase (E.C. Number: 2.7.1.137), comprometendo os metabolismos de glicerofosfolip?dios e de fosfato de inositol. Al?m disso, ? poss?vel que a Hexadecilfosfocolina interfira na atividade das fosfolipases C (E.C. Number: 3.1.4.11) e D (E.C. Number: 3.1.4.4) que participam do mecanismo de elimina??o descrito da mesma. Foram propostos ensaios experimentais envolvendo a exposi??o de trofozo?tos de Naegleria fowleri a hexadecilfosfocolina, principalmente estudos de cin?tica enzim?tica dos alvos detectados com a finalidade de comprovar os achados feitos. Limita??es metodol?gicas foram levantadas e estrat?gias de resolu??o das mesmas elencadas. Por fim, foi apresentada uma proposta de pesquisa in?dita de um novo protocolo terap?utico contra a meningoencefalite amebiana prim?ria. Disserta??o (Mestrado) ? Programa de P?s-gradua??o em Ci?ncias Farmac?uticas, Universidade Federal dos Vales do Jequitinhonha e Mucuri, 2022. The free-living amoeba Naegleria fowleri is the etiologic agent of primary amoebic meningoencephalitis. This is a central nervous system infection with a fulminant course with a mortality rate of approximately 97%. Hexadecylphosphocholine is a drug used in the treatment of this disease, a phospholipid analogue with anti-tumor and anti-protozoal properties. Despite being amoebicidal, the molecular mechanism underlying the cytopathic effect in Naegleria fowleri is not known. In silico analyzes were performed in order to clarify this. Enzymes from several organisms whose catalytic activities are affected by direct molecular interaction with hexadecylphosphocholine were identified in a literature review. Such enzymes were used to detect Naegleria fowleri enzymes through sequence alignment searches in the AMOEBADB database. The three-dimensional structures of the recovered enzymes were obtained directly from the PDB or from the Swiss-Model repository. Proteins without ready-made and experimentally unresolved models were modeled by the sequence homology method. The models were validated using the Swiss-Model metrics GMQE and QMEANDisCO, in addition to Errat, Prove, Verify 3D and PROCHECK software. Autodock Vina was used in molecular docking simula t ions between the detected enzymes and hexadecylphosphocholine, using rigid enzymes and flexible ligand. The affinity energies of the best binding poses ranged from -6.0 to -4.5 kcal/mol. Inferences of the cytopathic mechanism of action were made based on the evaluation of the hypothetically involved metabolic networks, all available in the KEEG and MetaCyC databases. Through the performed analyzes it is suggested that hexadecylphosphocholine may exert a cytopathic effect on Naegleria fowleri cells by inhibiting mainly the CTP enzymes: phosphocholine-cytidylyltransferase (E.C. Number: 2.7.7.15) and Phosphatidylinositol 3-kinase (E.C. Number: 2.7.1.137), compromising the metabolism of glycerophospholipids and inositol phosphate. In addition, it is possible that hexadecylphosphocholine interferes with the activity of phospholipases C (E.C. Number: 3.1.4.11) and D (E.C. Number: 3.1.4.4) that participate in its described elimination mechanism. Experimental assays involving the exposure of Naegleria fowleri trophozoites to hexadecylphosphocholine were proposed, mainly studies of enzymatic kinetics of the targets detected in order to prove the findings made. Methodological limitations were raised and strategies for their resolution were listed. Finally, an unprecedented research proposal was presented for a new therapeutic protocol against primary amoebic meningoencephalitis.
Databáze:	OpenAIRE
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