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Master's Dissertation
DOI
10.11606/D.9.2019.tde-28012019-115311
Document
Author
Full name
Alfredo Danilo Ferreira de Souza
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2018
Supervisor
Committee
Parise Filho, Roberto (President)
Almeida, Wanda Pereira
Vargas, Jeanine Giarolla
Kawano, Daniel Fabio
Title in Portuguese
Otimização de novos inibidores da di-idrofolato redutase de Mycobaterium tuberculosis (mtDHFR): Docking molecular, sí­ntese, avaliação da inibição enzimática e da atividade antimicobacte
Keywords in Portuguese
Bioisosterismo
mtDHFR
Otimização de fragmentos
Química farmacêutica
Tuberculose
Abstract in Portuguese
A tuberculose (TB) é considerada uma das principais doenças infecciosas e apresenta fatores críticos como a relação com o HIV/AIDS, tratamento longo e a resistência a múltiplos fármacos. A enzima di-hidrofolato redutase das micobactérias (mtDHFR) é um alvo pouco explorado e apresenta grande potencial para o desenvolvimento de novos fármacos contra TB. Estudos preliminares obtiveram fragmentos com baixa afinidade à mtDHFR, entretanto com potencial para otimização. Com isso, o fragmento foi usado como protótipo para a proposição de 22 análogos. Os compostos foram planejados utilizando informações sobre ligantes e a estrutura tridimensional de mtDHFR, além do biososterismo como estratégia norteadora. Os ensaios de docking molecular com a mtDHFR revelaram que os análogos propostos tiveram escores interessantes e, além disso, a inserção de substituintes demonstrou favorecer a ligação à enzima, o que corroborou o planejamento. Com isso, sintetizou-se 22 análogos planejados e o protótipo MB872, por meio de protocolos de alquilação, hidrólise e cicloadição 1,3 dipolar para os compostos com anéis triazol e tetrazol. Os compostos foram obtidos com rendimentos de bom a ótimo (60 ~ 90%) e suas estruturas foram elucidadas por RMN 1H e 13C. Os resultados do ensaio de inibição enzimática corroboraram com os dados de docking, uma vez que a presença do grupo carboxílico revelou ser importante para a atividade. Além disso, alguns dos compostos revelaram atividades interessantes, entre 8 a 40 µM, sendo que o mais ativo apresentou IC50 de 7 µM. Ensaios de cinética enzimática com o análogo mais ativo indicou uma inibição não competitiva com o substrato natural da enzima, uma vez que os valores de Km se mantiveram constantes, enquanto Vmax decaiu (0,22 µM e 0,43 - 0,34 ΔFU/min, respectivamente). Os análogos sintetizados foram mandados para ensaio in vitro para avaliar a atividade frente a micobactéria.
Title in English
Optimization of new dihydrofolate reductase inhibitors of mycobacterium tuberculosis (mtdhfr): molecular docking, synthesis, evaluation of enzyme inhibition and antimycobacterial activity
Keywords in English
Bioisosterism
Fragments optimization
mtDHFR
Pharmaceutical chemistry
Tuberculosis
Abstract in English
Tuberculosis (TB) is an important infectious disease and presents critical factors such as the relationship with HIV / AIDS, long treatment and resistance to multiple drugs. The enzyme dihydrofolate reductase from mycobacteria (mtDHFR) is a poorly explored and presents great potential to be a target for new drugs against TB. Preliminary studies have obtained fragments with low affinity to mtDHFR, but with potential to become lead compounds. Therefore, the fragment was used as a prototype for 22 analogues proposed in this work. The compounds were designed using bioisosterism, information about ligands and the three-dimensional structure of mtDHFR. Molecular docking assays with mtDHFR revealed satisfactory scores for anlogues. Furthermore, the insertion of substituents seemed to increase the affinity with the enzyme. Thereby, twenty two analogues and prototype were synthesized using alkylation, hydrolysiss and 1,3-dipolar cycloaddition methods. The compounds were obtained in good yields (60 ~ 90%) and their structures were elucidated with 1H and 13C NMR spectroscopy. The enzymatic affinity assay corroborates docking results, because the presence of carboxyl group showed to be important for the activity. Furthermore, some of the compounds revealead interesting activities, ranging 8 to 40 µM. The most active showed IC50 of 7 µM and enzyme kinetics assays indicated noncompetitive inhibition with natural enzyme substrate. The synthesized analogs were sent for in vitro assay to assess mycobacteria activity.
 
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Publishing Date
2019-01-29
 
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