Naslov
Molecular methods in pharmacogenetics

Autori
Štefanović, Mario

Izvornik
Periodicum Biologorum (0031-5362) 103 (2001), Supp 1;

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, kongresno priopcenje, znanstveni

Ključne riječi
cytochrome P450; pharmacogenetics; molecular methods

Sažetak
The cytochrome P450s (CYP) and phase II conjugating enzymes play an important role in metabolizing drugs, activation and detoxification of chemical xenobiotics. Variants of genes (alleles) for specific human CYP and phase II enzymes typically occur with variable frequency between individuals and may result in poor metabolizers or extensive metabolizers for specific substrates. Because their response to drug therapy may be greatly altered by defects in genes that code for these enzymes, molecular diagnostics is of great importance for detecting individual metabolic capacity. Many molecular diagnostic techniques are used for genotyping drug-metabolizing enzyme polymorphisms, and they may in general be divided into two categories 1) classical PCR based, low throughput, relatively cheap, time/effort consuming techniques and 2) modern – high throughput, expensive techniques that can be fully automated, being developed by different companies. Commonly used classical methods are : - PCR-restriction fragment length polymorphism (PCR-RFLP), - Single strand conformation polymorphism (SSCP), - allele-specific amplification (ASA) and multiplex ASA, - amplification -refractory-mutation assay (ARMS), - ligase chain reaction (LCR) - heteroduplex analysis (HA) and several others. Some of these methods detect a mutation position and identification while some like SSCP and HA do not. Although all those methods are relatively simple they are time and effort consuming. Even more important is the limiting number of important polymorphisms that can simultaneously be detected and a number of genes that can be screened within particular individual. A recent development has been the introduction of several methods that use cleavage of heteroduplexes, mass spectrometry or hybridization of a reporter probe (Taqman real-time PCR) – for distinguishing two genetic variants. Current genotyping methods are going to be replaced in the future with more sophisticated automated systems that offer high throughput, are not time consuming and simultaneously screen for almost 95-99% if not all known genetic defects within all important drug metabolism enzyme systems. Furthermore, when handling huge numbers of samples there is a need to rely on robotics both to increase productivity and because of the risk of human error. In the future one could expect the common use of DNA microarray chip technology which ensures the capacity to assay a large number of samples in a short time on the principles of miniaturization, multiplexing and automation, providing a set of performance specifications that cannot be achieved with the earlier technologies.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti

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