Genotyping of the Most Common Thiopurine Methyltransferase Mutations with the LightCycler

Abstract

Thiopurine derivatives are commonly used for immunosuppressive therapy after organ transplantation (azathioprine) and in the therapy of leukemia (mercaptopurine and thioguanine). Their pharmacological efficacy is based on an in vivo toxification pathway that ultimately leads to 6-thioguanine nucleotides, which act as anti-metabolites and interfere with nucleic acid synthesis [1]. Two detoxification pathways are known, one via xanthine oxidase, a stably abundant enzyme in the Caucasian population, and the second via thiopurine methyltransferase (TPMT, EC.2.1.1.67). The latter enzyme is subject to a genetic polymorphism, that in 11% of the Caucasian population leads to a heretozygous deficiency and in 0.3% to a homozygous deficiency of this enzyme [3, 4, 6]. If patients with a homozygous deficiency of TPMT are given thiopurines at the usual dosage, this will lead to a severe myelosuppression [9, 10] often with life-threatening pancytopenia [5]. Phenotyping of this enzyme is possible [2], but the method is laborious and technically demanding; it is therefore only performed in specialized laboratories. Genotyping of this defect is, on the other hand, hampered by the fact that to date eight mutation are known [11–13] that lead to a deficient phenotype (Fig. 1). Around 90% of TPMT deficiencies can be attributed to one of these known mutations.