THIOPURINE S-METHYLTRANSFERASE DEFICIENCY - 2 NUCLEOTIDE TRANSITIONS DEFINE THE MOST PREVALENT MUTANT ALLELE ASSOCIATED WITH LOSS OF CATALYTIC ACTIVITY IN CAUCASIANS

The autosomal recessive trait of thiopurine S-methyltransferase (TPMT) deficiency is associated with severe hematopoietic toxicity when pati ents are treated with standard doses of mercaptopurine, azathioprine, or thioguanine. To define the molecular mechanism of this genetic poly morphism, we cloned and characterized the cDNA of a TPMT-deficient pat ient, which revealed a novel mutant allele (TPMT3) containing two nuc leotide transitions (G(460)-->A and A(719)-->G) producing amino acid c hanges at codons 154 (Ala-->Thr) and 240 (Tyr-->Cys), differing from t he rare mutant TPMT allele we previously identified (i.e., TPMT2 with only G(238)-->C). Site-directed mutagenesis and heterologous expressi on established that either TPMT3 mutation alone leads to a reduction in catalytic activity (G(460)-->A, ninefold reduction; A(719)-->G, 1.4 -fold reduction), while the presence of both mutations leads to comple te loss of activity. Using mutation specific PCR-RFLP analysis, the TP MT3 allele was detected in genomic DNA from approximate to 75% of unr elated white subjects with heterozygous phenotypes, indicating that TP MT3 is the most prevalent mutant allele associated with TPMT-deficien cy in Caucasians.