Background Thiopurine S-methyltransferase is a key enzyme in the metabolization of azathioprine. Reduced activity leads to the accumulation of a toxic metabolite 6-thioguanine. This condition is genetically determined (OMIM *187680) and it occurs in 11% individuals of the Caucasian population. Testing of frequent pathogenic allelic variants before azathioprine therapy is recommended for c.238G>C, c.460G>A, c.626–1G>A and c.719A>G. These variants are most frequent (covers more than 95% cases of reduced activity of TPMT enzyme). Defects in other enzymes involved in azathioprine metabolization are extremely rare.

Hereditary xanthinuria type I is caused by a mutation in the XDH gene (OMIM #278300) and it reduce the activity of xanthine dehydrogenase (XDH). Type II is caused by the mutation in XDH and AOX1 and/or MOCOS gene (OMIM #603592) and lead to inactivity of XDH and aldehyde oxidase (AOX1). Molybdenum cofactor sulfurase (encoded by MOCOS) is essential for activities of XDH and AOX1. In patients with purine enzymopathy the accumulation of 6-thioguanine may also occur.

Materials and methods We examined the occurrence of pathogenic allelic variants c.238G>C, c.460G>A, c.626–1G>A, c.719A>G by PCR amplification and direct sequencing in 62 individuals.

In addition, we analysed remaining exons of the TPMT gene in four patients with the history of myelotoxicity. In one patient with undetectable serum UA levels we performed high-performance liquid chromatography of urine and we subsequently analysed XDH, AOX1 and MOCOS genes.

Results We detected heterozygous allelic variant c.238G>C in one case, heterozygous c.460G>A in six cases, heterozygous c.719A>G in seven cases and homozygous c.719A>G in one case. In one patient with myelotoxicity after azathioprine we discovered an unpublished allelic variant c.85T>C (in silico analysis predicted damaging effect). In one patient we diagnosed xanthinuria type II (xanthine in urine was 137 mmol/mol creatinine - reference <25) and subsequently analysis identified rare homozygous allelic variant c.362C>T in the MOCOS (in silico analysis predicted damaging effect).

Conclusions We detected pathogenic allelic variants in the TPMT in ten patients from 62 individuals. The examination of reduced TPMT enzyme activity should become a standard of care. The relationship between purine enzymopathies and azathioprine metabolization should be further studied.

Acknowledgements Supported by Ministry of Health, Czech Republic - conceptual development of research organisation (Institute of Rheumatology – RÚ, 00023728).