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Doctoral Thesis
DOI
https://doi.org/10.11606/T.42.2011.tde-20032012-144514
Document
Author
Full name
Juan Gonzalo Aliaga Gamarra
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2011
Supervisor
Committee
Nucci, Gilberto de (President)
Huenuman, Nilton Erbet Lincopan
Malpartida, Humberto Miguel Garay
Oliveira, Antonio Carlos
Ramos, Augusto Savio Peixoto
Title in Portuguese
Estudo para a caracterização genotípica e fenotípica da atividade enzimática da subfamilia citocromo P450 CYP2D6 de voluntários sadios.
Keywords in Portuguese
Farmacogenética
Fenótipos
Genótipos
Metabolismo
Polimorfismo
Abstract in Portuguese
As enzimas CYP450 são as principais enzimas metabolizadoras de fármacos. Elas são codificadas por genes que apresentam polimorfismos gênicos que lhes confere características fenotípicas diversas. Estes fenótipos são: Metabolizadores Lentos ou PM, Metabolizadores Normais ou EM, Metabolizadores Intermediários ou IM e Metabolizadores Ultra-rápidos ou UM. A Farmacogenética é a ciência que estuda estas variações gênicas e sua relação com a resposta terapêutica no organismo. Entre as enzimas CYP450 se encontram as enzimas CYP2D6, que são responsáveis pelo metabolismo de 25% dos fármacos clinicamente prescritos. O objetivo principal deste estudo foi a identificação dos polimorfismos mais importantes deste gene: CYP2D6*1, CYP2D6*2, CYP2D6*3, CYP2D6*4, CYP2D6*5, CYP2D6*6, CYP2D6*10, CYP2D6*17 e CYP2D6*41 pelos métodos de Genotipagem (PCR Tetra Primer e Seqüenciamento) e Fenotipagem (analisado pelo Índice Metabólico) em 75 voluntários sadios da região de Campinas. Para a caracterização da Fenotipagem foi usada a substância teste Dextrometorfano (DM). Esta foi monitorada por espectrometria de massa mediante a determinação da concentração do seu principal metabólito o Dextrorfano (DX), que foi extraída das amostras de urina. Os resultados foram comparados entre estas duas metodologias e apresentaram alta correlação. Os resultados obtidos são a identificação das freqüências dos alelos *1, *3 e *4 pelo método PCR Tetra Primer (30.66%, 1.3% e 14%, respectivamente). O método de seqüenciamento detectou também outros alelos que não foram detectados pela PCR Tetra Primer. A avaliação do número de cópias do gene CYP2D6 também foi avaliada, detectando em um voluntário 3 cópias do gene CYP2D6, característica de metabolizadores Ultra-rápidos. Podemos afirmar que os métodos usados forneceram perfis dos polimorfismos de maneira rápida e prática.
Title in English
Study to genotype and phenotype characterization of enzymatic activity of subfamily cytochrome P450 CYP2D6 of health volunteers.
Keywords in English
Genotypes
Metabolism
Pharmacogenetics
Phenotypes
Polymorphism
Abstract in English
The CYP450 enzymes are the major drug metabolizing enzymes. They are encoded by genes that show genetic polymorphisms which gives them several phenotypic characteristics. These phenotypes are Poor Metabolizers or PM, or Extensive Metabolizers or EM, Intermediate Metabolizers or IM, and finally, Ultra-rapid Metabolizers or UM. Pharmacogenetics is the science that studies these genetic variations and its relationship to therapeutic response in the body. One of CYP450 enzymes is CYP2D6 enzyme, which are responsible for the metabolism of 25% of clinically prescribed drugs. The main objective of this study was to identify the most important polymorphisms of this gene: CYP2D6 * 1, CYP2D6 * 2, CYP2D6 * 3, CYP2D6 * 4, CYP2D6 * 5, CYP2D6 * 6, CYP2D6 * 10, CYP2D6 * 17 and CYP2D6 * 41 by genotyping methods (PCR Tetra Primer and Sequencing) and phenotyping (by metabolic rate monitoring) in 75 healthy volunteers in Campinas region.To characterize the phenotyping was used to test substance Dextromethorphan (DM). This was monitored by mass spectrometry by determining the concentration of its major metabolite the Dextrorphan (DX), which was extracted from urine samples. The results were compared between these two methods and showed high correlation. We can obtain the identification of allelic frequencies of alleles * 1, * 3 and * 4 by Tetra Primer PCR (30.66%, 1.3% and 14% respectively). The sequencing method has also detected other alleles that were not detected by PCR Tetra Primer. The assessment of the number of copies of the CYP2D6 gene was also assessed. This method detected a volunteer which carrying three copies of CYP2D6 gene, characteristic of Ultra-rapid metabolizers. We can say that the methods used in this study provide polymorphism profiles quickly and conveniently.
 
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Publishing Date
2012-05-25
 
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