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Thiopurine s-methyltransferase (TPMT) plays a key role in the metabolism of the drug; 6-MP in children with acute lymphoblastic leukemia (ALL). The wild-type TPMT*1 allele and mutant alleles are associated with normal and intermediate enzyme activity, respectively. In patients with intermediate or deficient TPMT activity, normal dose of the drug may cause serious side effects such as bone marrow toxicity. The aim of this study was to assay the enzyme activity by HPLC for right ordering of chemotherapy drug doses in the patients. TPMT activity was measured in RBC of healthy adults (n=73) and children (n=10). Also, TPMT (*2, *3A, *3B and *3C) genotype of the samples were assessed by ARMS-PCR and RFLP-PCR. No indication of gender and age differences in the TPMT activity was found. This study showed that the HPLC method was sensitive, accurate and precise and can be used in routine clinical laboratory tests.