Augmenting Clinical Interpretability of Thiopurine Methyltransferase Laboratory Evaluation

Abstract

Objective: Individuals with decreased thiopurine methyltransferase (TPMT) activity are at risk of adverse effects of thiopurine administration whereas its increased activity may inactivate drugs faster. We evaluated genotype-phenotype correlations in patients with suspected hematological malignancies and inflammatory bowel disease from our region based on findings of nonlinear TPMT enzyme kinetics previously unreported. Patients and Methods: The study group comprised 267 individuals. They were screened for the most common variants of low TPMT activity. TPMT activity was measured in erythrocytes using the HPLC rate-blanked method. Results: Thirty-three patients (12.4%) were heterozygous (26 were TPMT*1/*3A, 5 TPMT*1/*2, 2 TPMT *1/*3C) and 1 was a compound heterozygote (*2/*3A). Normal and low normal TPMT activities substantially overlapped in wild-type and heterozygous individuals, whereas high activities were found in 29 wild-type genotyped patients. Extreme and life-threatening toxicity was observed in the compound heterozygote patient. Conclusion: Activity measurement performed at diagnosis provides clinicians with information on immediate pharmacokinetic-related adverse events and/or hypermetabolism, and genotyping may indicate the rate of pharmacodynamic thioguanine nucleotide accumulation due to slower overall thiopurine metabolism.

Determination of thiopurine S -methyltransferase activity by hydrophilic interaction liquid chromatography hyphenated with mass spectrometry

“…While most of the published methods used a C 18 RP-column [18,19,[22][23][24][25][26][27][28][29], we preferred a C 8 RP-column in order to improve separation of 6-MMP from the matrix constituents. This is critical when using non-selective UV detection.…”

“…Moreover, the radiochemical method is more time-consuming than currently preferred HPLC methods [16]. Over the years more suitable methods for TPMT phenotyping were developed, namely mass spectrometry [14], capillary electrophoresis combined with UV detection [17] or, most commonly, HPLC coupled with UV/DAD [16,[18][19][20][21][22][23][24][25][26][27] or fluorescence detection [28]. Although separations with non-selective UV detection have been preferred due to their simplicity and low cost, increasing demands on the reliability of data and more detail sample profiles stimulated development of more advanced methods.…”

The Examination of a TPMT Gene Before Administration of Azathioprine in Rheumatology Practice and Identification of a Novel Variant p.W29R

“…Genetic testing is routinely performed before starting therapy with AZA, but it is also possible to determine the activity of the TPMT enzyme in erythrocytes using high-performance liquid chromatography 19 . This examination was performed on selected patients at a collaborating clinic.…”