Abstract
Thiopurine drugs are metabolized, in part, by S-methylation catalyzed by thiopurine S-methyltransferase (TPMT). Patients with very low or undetectable TPMT activity are at high risk of severe, potentially fatal hematopoietic toxicity when they are treated with standard doses of thiopurines. As human TPMT activity is controlled by a common genetic polymorphism, it is an excellent candidate for the clinical application of pharmacogenetics. Here, we report a new molecular approach developed to detect point mutations in the TPMT gene that cause the loss of TPMT activity. A fluorescently labeled amplified DNA is hybridized with oligonucleotide DNA probes immobilized in gel pads on a biochip. The specially designed TPMT biochip can recognize six point mutations in the TPMT gene and seven corresponding alleles associated with TPMT deficiency: TPMT*2; TPMT*3A, TPMT*3B, TPMT*3C, TPMT*3D, TPMT*7, and TPMT*8. The effectiveness of the protocol was tested by genotyping 58 samples of known genotype. The results showed 100% concordance between the biochip-based approach and the established PCR protocol. The genotyping procedure is fast, reliable and can be used for rapid screening of inactivating mutations in the TPMT gene. The study also provides the first data on the frequency of common TPMT variant alleles in the Russian population, based on a biochip analysis of 700 samples. TPMT gene mutations were identified in 44 subjects; genotype *1/*3A was most frequent.
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Acknowledgements
We thank Edward Kreindlin, Kirill Evseev, and Valentina Chupeeva for manufacture of the microchips and Sergei Surzhikov and Irina Grechishnikova for oligonucleotide synthesis.
This work was supported by the St Jude Children's Research Hospital International Outreach Program and by the American Lebanese Syrian Associated Charities (ALSAC).
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Nasedkina, T., Fedorova, O., Glotov, A. et al. Rapid genotyping of common deficient thiopurine S-methyltransferase alleles using the DNA-microchip technique. Eur J Hum Genet 14, 991–998 (2006). https://doi.org/10.1038/sj.ejhg.5201647
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DOI: https://doi.org/10.1038/sj.ejhg.5201647
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