Screening of thiopurine S‐methyltransferase mutations by horizontal conformation‐sensitive gel electrophoresis

Alves, Sandra; Prata, Maria João; Ferreira, Fátima; Amorim, António

doi:10.1002/(sici)1098-1004(200003)15:3<246::aid-humu5>3.0.co;2-#

Cited by 28 publications

(22 citation statements)

References 19 publications

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“…In general, drug response is affected by several factors, such as sex, age, environment, drug-drug interactions, and drug-enzyme interactions (4 ). Another important factor for explaining interindividual differences in therapeutic efficacy and adverse reactions is genetic variation in the enzymes in the metabolic pathways for these drugs (1,3,(5)(6)(7)(8)(9)(10).…”

confidence: 99%

“…The sequences of the PCR primers for amplification of exons IV and V were as described by Alves et al (6 ). The TPMT gene was identified by use of the UCSC genome server (http://genome.ucsc.edu), and PCR primers for amplification of exons VII, VIII, and X were designed with PRIMER3, available through the Biology WorkBench (http://workbench.sdsc.edu) (31 ).…”

Section: Pcrmentioning

confidence: 99%

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Pyrosequencing of TPMT Alleles in a General Swedish Population and in Patients with Inflammatory Bowel Disease

Haglund

Lindqvist²,

Almér

et al. 2004

Clinical Chemistry

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Background: Interindividual differences in therapeutic efficacy in patients treated with thiopurines might be explained by the presence of thiopurine S-methyltransferase (TPMT) alleles that encode for reduced TPMT enzymatic activity. It is therefore of value to know an individual's inherent capacity to express TPMT. Method: We developed a pyrosequencing method to detect 10 single-nucleotide polymorphisms (SNPs) in TPMT. A Swedish population (n ‫؍‬ 800) was examined for TPMT*3A, TPMT*3B, TPMT*3C, and TPMT*2. Patients with inflammatory bowel disease (n ‫؍‬ 24) and healthy volunteers (n ‫؍‬ 6), selected on the basis of TPMT enzymatic activity, were investigated for all 10 SNPs to determine the relationship between TPMT genotype and phenotype. Results: In the general population we identified the following genotypes with nonfunctional alleles:

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confidence: 99%

Section: Pcrmentioning

confidence: 99%

Pyrosequencing of TPMT Alleles in a General Swedish Population and in Patients with Inflammatory Bowel Disease

Haglund

Lindqvist²,

Almér

et al. 2004

Clinical Chemistry

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“…The most common alleles contain four or five repeats, occurring with frequencies of 55 and 34%, respectively. Transient transfection experiments demonstrated only a modest modulatory effect of VNTR alleles on promoter activity (Spire-Vayron et al, 1999), though the clinical importance of these findings still remains to be elucidated (Spire-Vayron et al, 1999;Yan et al, 2000;Alves et al, 2001). …”

Section: Variations Of Transcriptionmentioning

confidence: 99%

“…Detection of individual SNPs is then achieved by a variety of methods, including RFLP, allele-specific amplification, direct sequencing, SSCP or DHPLC analysis ; Spire-Vayrone de la Mourneyre et al, 1998;Alves et al, 1999;Seki et al, 2000;Schaeffeler et al, 2001). DNA-based methods to prospectively diagnose TPMT deficiency offer a clinically important strategy to minimize the risk of potentially life-threatening hematopoietic toxicity in patients treated with these medications (Black et al, 1998;Relling et al, 1999b;Scerri, 1999;Coulthard et al, 2000).…”

Section: Determination Of Tpmt Activity and Genotype In Humansmentioning

confidence: 99%

Drug methylation in cancer therapy: lessons from the TPMT polymorphism

2003

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The genetic polymorphism of thiopurine methyltransferase (TPMT) is one of the most developed examples of pharmacogenetics, spanning from molecular genetics to clinical diagnostics for individualizing thiopurine therapy (i.e. azathioprine, mercaptopurine, and thioguanine). Elucidation of the molecular mechanisms and biochemical consequences of TPMT deficiency demonstrates how pharmacogenetic traits can be identified, characterized, and translated to the bedside. Insights gained from studies of the TPMT polymorphism illustrate the potential of pharmacogenomics to optimize cancer therapy by avoiding toxic side effects in genetically distinct subgroups of patients.

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“…For the VNTR typing, we used the HCSGE (horizontal conformation sensitive gel electrophoresis) technique described in Alves et al (2000b). This procedure amplifies the tendency of multiple or single-base mismatches to produce conformational changes in the double-stranded helix, with a consequent increment of the differential migration of DNA homoduplexes and heteroduplexes during electrophoretic separation.…”

Section: Genotypingmentioning

confidence: 99%

Evolution of a VNTR located within the promoter region of the thiopurine methyltransferase gene: inferences from population and sequence data

Alves¹,

Amorim²,

Prata³

2002

Hum Genet

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The promoter region of the human thiopurine methyltransferase (TPMT) gene contains a variable number of tandem repeats (VNTR) with three kind of motifs (A, B, and C) differing by the length of the unit core and nucleotide sequence. We have studied the structural variation within the VNTR alleles in two human populations and in samples from gorillas and chimpanzees. In humans, no intermingling of motifs was detected within the VNTR, and the sequences of the three core motifs remained remarkably unchanged, differences between alleles corresponding essentially to variations in the number of A and B repeats. The variation pattern in humans is consistent with an interpretation in which two contiguous genetic units (repeats A and B) behave evolutionarily according to the stepwise mutation model, as inferred from the population distribution profiles and from the molecular phylogenetic relationships among the VNTR alleles. However, the observation of a strong negative correlation between the numbers of A and B repeats also suggests that the regularity and/or independence of the mutational process has been disrupted to some extent by interactions between the A and B stretches. Selective pressure (the VNTR plays some role, although minor, in the TPMT function) or biased mutation are possible explanations. In gorillas and chimpanzees, several A-, B-, or C-like motifs were detected, but their arrangement within the VNTR alleles did not followed the regular pattern registered in humans and, particularly for the B-like motifs, a considerable sequence hypervariability was registered. Furthermore, the structural differences among non-human alleles were sufficiently numerous to render more plausible the assumption of the infinite allele model.

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Screening of thiopurine S‐methyltransferase mutations by horizontal conformation‐sensitive gel electrophoresis

Cited by 28 publications

References 19 publications

Pyrosequencing of TPMT Alleles in a General Swedish Population and in Patients with Inflammatory Bowel Disease

Pyrosequencing of TPMT Alleles in a General Swedish Population and in Patients with Inflammatory Bowel Disease

Drug methylation in cancer therapy: lessons from the TPMT polymorphism

Evolution of a VNTR located within the promoter region of the thiopurine methyltransferase gene: inferences from population and sequence data

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