Phenotype Determination of Thiopurine Methyltransferase in Erythrocytes by HPLC

Abstract

Thiopurine methyltransferase (TPMT) is a cytosolic enzyme that catalyzes the S -methylation of thiopurine drugs, which are used in cancer chemotherapy and as immunosuppressive agents (1). TPMT activity is controlled by a common genetic polymorphism that contributes to interindividual variability in drug response and, consequently, to implications for thiopurine therapeutic efficacy and toxicity (2). Severe myelosuppression has been reported for TPMT-deficient patients treated with standard doses of thiopurines (3)(4)(5), and high TPMT activity has been associated with the rejection of transplanted organs (6). Because of the clinical significance of the TPMT genetic polymorphism, determination of the TPMT phenotype in red blood cells is routinely performed to optimize and individualize thiopurine treatment (5). Variant alleles of the TPMT gene have been characterized and associated with low TPMT activity (7)(8). Recently, Spire-Vayron de la Moureyre et al. (9) reported that genotypic analysis of TPMT allows the correct determination of metabolic capacity for 87% of individuals. A lower correlation was found for individuals with TPMT activity that was close to the antimode value. Thus, phenotypic analysis may be useful and could be performed concomitantly with genotyping tests. TPMT activity has classically been measured using a radiochemical assay (10). Few HPLC methods using nonradiolabeled calibrators with liquid-liquid extraction (11)(12) or solid-phase extraction (13) have been reported. Here, we report a reversed-phase HPLC method that uses a simple and rapid treatment procedure for the determination …