Human Thiopurine S-Methyltransferase Pharmacogenetics: Variant Allozyme Misfolding and Aggresome Formation

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 102; no. 26; pp. 9394 - 9399
• Main Authors: Wang, Liewei, Nguyen, Tien V., McLaughlin, Richard W., Sikkink, Laura A., Ramirez-Alvarado, Marina, Weinshilboum, Richard M., Conney, Allan H.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 28.06.2005; National Acad Sciences
• Subjects: Aggregation; Alleles; Animals; Antibodies; Bacterial Proteins - chemistry; Biological Sciences; Catalysis; Cell aggregates; Chromatography; Circular Dichroism; COS Cells; Cysteine Proteinase Inhibitors - pharmacology; Cytosol; Enzymes; Escherichia coli - metabolism; Genetic Variation; Genetics; Histone Deacetylase 6; Histone Deacetylases - chemistry; Homozygote; Humans; Hydroxylamines - pharmacology; Kinetics; Leupeptins - pharmacology; Methyltransferases - genetics; Methyltransferases - pharmacology; Microscopy, Fluorescence; Microtubules - metabolism; Monomers; Pharmacogenetics; Pharmacogenetics - methods; Polymorphism, Genetic; Protein Folding; Protein Structure, Tertiary; Proteins - chemistry; Purines - chemistry; Quinolines - pharmacology; Recombinant Proteins - metabolism; Temperature; Transfection; Ubiquitin - chemistry; Vinblastine - pharmacology
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0502352102

Thiopurine S-methyltransferase (TPMT) catalyzes the S-methylation of thiopurine drugs. TPMT genetic polymorphisms represent a striking example of the potential clinical value of pharmacogenetics. Subjects homozygous for TPMT*3A, the most common variant allele for low activity, an allele that encodes a protein with two changes in amino acid sequence, are at greatly increased risk for life-threatening toxicity when treated with standard doses of thiopurines. These subjects have virtually undetectable levels of TPMT protein. In this study, we tested the hypothesis that TPMT*3A might result in protein misfolding and aggregation. We observed that TPMT*3A forms aggresomes in cultured cells and that it aggregates in vitro, functional mechanisms not previously described in pharmacogenetics. Furthermore, there was a correlation among TPMT half-life values in rabbit reticulocyte lysate, aggresome formation in COS-1 cells, and protein aggregation in vitro for the three variant allozymes encoded by alleles that include the two TPMT*3A single-nucleotide polymorphisms. These observations were compatible with a common structural explanation for all of these effects, a conclusion supported by size-exclusion chromatography and CD spectroscopy. The results of these experiments provide insight into a unique pharmacogenetic mechanism by which common polymorphisms affect TPMT protein function and, as a result, therapeutic response to thiopurine drugs.