Measurement of thiopurine <i>S</i>-methyltransferase activity                in human blood samples based on high-performance liquid chromatography: reference                values in erythrocytes from children

Keizer-Garritsen, Jenneke J.; Brouwer, C.; Lambooy, Lambert H.J.; Riet, Patricia Ter; Bökkerink, J.P.M.; Trijbels, Frans J.M.; Abreu, Ronney A. De

doi:10.1258/000456303321016222

Cited by 28 publications

(28 citation statements)

References 22 publications

(29 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The retention times of 6-MMP in the published RP-HPLC methods ranged between ca. 4 and 27 min [22,27] while in our case it was 9.3 min. The optimized C 8 RP-HPLC-UV method was suitable for the identification of 6-MMP as well as 6-MP, however, some partial overlapping could not be avoided as it can be seen in Fig.…”

Section: Methods Developmentcontrasting

confidence: 45%

“…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.…”

Section: Methods Developmentmentioning

confidence: 99%

“…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.…”

Section: Indroductionmentioning

confidence: 99%

See 2 more Smart Citations

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

Pecher

Dokupilová

Zelinková

et al. 2017

Journal of Pharmaceutical and Biomedical Analysis

View full text Add to dashboard Cite

Determination of thiopurine S-methyltransferase activity by hydrophilic interaction liquid chromatography hyphenated with mass spectrometry, Journal of Pharmaceutical and Biomedical Analysishttp://dx.doi.org/10. 1016/j.jpba.2017.05.016 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. 1 Highlights New HILIC-HPLC-MS method was developed for determination of TPMT activity  HILIC-HPLC-MS combination was approved by excellent performance parameters  HILIC-HPLC-MS was compared to traditional RP-HPLC-UV as well as advanced RP-HPLC-MS  Conventional approaches were surpassed in terms of selectivity, LOD, analysis time  HILIC-HPLC-MS is favorable for routine assay of 6-MMP/6-MP ratio in RBC lysates List of abbreviations6-MMP -6-methylmercaptopurine, 6-MP -6-mercaptopurine, 6-TG -6-thioguanine, AZA -azathioprine, Cl-P -6-chloropurine, DTT -dithiothreitol, IBD -inflammatory bowel diseases, RBC -red blood cells, SAM -S-adenosyl-L-methionine, TGN -thioguanine nucleotides, TPMT -thiopurine-S-methyl transferase. IndroductionThiopurine drugs, namely azathioprine (AZA), 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG), are immunosuppressive agents that are, due to their low cost and high effectiveness, widely used in the treatment of inflammatory bowel diseases (IBD), but also in other autoimmune diseases, some hemopoetic disorders and some solid organ transplant [1][2][3][4][5]. However, the occurrence of adverse drug reactions limits the use of these drugs. Up to 15 % of IBD patients discontinue this type of treatment due to adverse events that include nausea, skin reactions, pancreatitis, hepatotoxicity or myelotoxicity [6,7].Thiopurine drugs are prodrugs, so their biological activity is preceded by extensive metabolism. Briefly, after absorption, AZA is metabolized in the liver to 6-MP which then can be metabolized to active thioguanine nucleotides (TGN) and to inactive methylated product 6-methylmercaptopurine (6-MMP) by the thiopurine-S-methyl transferase (TPMT) enzyme [8]. TPMT (EC 2.1.1.67), a key enzyme involved in the thiopurine drug metabolism, is subject to common genetic polymorphism that leads to trimodial distribution of the TPMT activity in population [9]. Individuals with lower TPMT activity might have an increased risk of developing thiopurine-induced myelosuppression. On the contrary, individuals with high TPMT activity have lower concentrations of active TGN metabolites resulting in reduced therapeutic efficacy of the drug [10]. In addition, high TPMT activity leads to accumulation of the hepatotoxic methylated metabolites [11]. Therefore, some scientific committees advise to determine the TP...

show abstract

Section: Methods Developmentcontrasting

confidence: 45%

Section: Methods Developmentmentioning

confidence: 99%

Section: Indroductionmentioning

confidence: 99%

See 1 more Smart Citation

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

Pecher

Dokupilová

Zelinková

et al. 2017

Journal of Pharmaceutical and Biomedical Analysis

View full text Add to dashboard Cite

show abstract

“…Thiopurine S-methyl transferase activity in RBC was determined by a validated high-performance liquid chromatography (HPLC) as described earlier. 28 …”

Section: Thiopurine S-methyl Transferase (Tpmt)mentioning

confidence: 99%

The effect of allopurinol and low-dose thiopurine combination therapy on the activity of three pivotal thiopurine metabolizing enzymes: Results from a prospective pharmacological study

Seinen

Asseldonk

Boer

et al. 2013

Journal of Crohn's and Colitis

View full text Add to dashboard Cite

Introduction: Thiopurine therapy is often discontinued in inflammatory bowel disease (IBD) patients. The xanthine oxidase (XO) inhibitor allopurinol has previously shown to enhance thiopurine efficacy and to prevent adverse reactions, the mechanism of this beneficial interaction is not completely clarified. The aim of this study is to observe possible effects of allopurinol and low-dose thiopurine combination therapy on the activity of three pivotal thiopurine metabolizing enzymes. Methods: A prospective study of IBD patients failing thiopurine therapy due to a skewed thiopurine metabolism was performed. Patients were treated with allopurinol and azathioprine or mercaptopurine. Xanthine oxidase, hypoxanthine-guanine phosphoribosyl transferase (HGPRT) and thiopurine S-methyl transferase (TPMT) activities, and thiopurine metabolites concentrations were measured during thiopurine monotherapy, and after 4 and 12 weeks of combination therapy. Conclusion: Allopurinol and thiopurine combination-therapy seems to increase HGPRT and decrease XO activity in IBD patients, which at least in part may explain the observed changes in thiopurine metabolite concentrations.

show abstract

“…The sample preparation and HPLC analysis was adopted from a previously described procedure [30] . Briefl y, RBC were isolated from heparinized blood, lysed, and supernatants were collected and stored at -80 ° C until further usage.…”

Section: Tpmt Enzyme Activity Assaymentioning

confidence: 99%

Thiopurine S-Methyltransferase Pharmacogenetics: Genotype to Phenotype Correlation in the Slovenian Population

et al. 2006

View full text Add to dashboard Cite

The toxicity of thiopurine drugs has been correlated to the activity of thiopurine S-methyltransferase (TPMT), whose interindividual variation is a consequence of genetic polymorphisms. We have herein investigated the relevance of some genetic markers for the prediction of thiopurine-related toxicities and to determine the genotype to phenotype correlation in the Slovenian population. The most prevalent mutant allele in the Slovenian population is TPMT*3A (4.1%), followed by TPMT*3C (0.5) and TPMT*3B (0.3), while the TPMT*2 allele was not found in any of the examined samples. TPMT enzyme activity distribution in the subgroup sample was bimodal and as such correlated with genetic data. Using a cutoff value of 9.82 pmol/10⁷ RBC per h, the genetic data correctly predicted TPMT enzyme activity in 91.6% of the examined individuals. Pharmacogenetic TPMT analyses have therefore proved to have significant clinical implications for prediction of individuals’ responses to treatment with thiopurine drugs in order to avoid possible life-threatening therapy-related toxicities.

show abstract

Measurement of thiopurine S-methyltransferase activity in human blood samples based on high-performance liquid chromatography: reference values in erythrocytes from children

Abstract: Background Monitoring 6-thiopurine S-methyltransferase (TPMT; EC 2.1.1.67) activity is especially important when patients are treated with 6-thiopurine drugs, since severe bone marrow toxicity may be induced if patients have deficient TPMT activity.

Cited by 28 publications

References 22 publications

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

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

The effect of allopurinol and low-dose thiopurine combination therapy on the activity of three pivotal thiopurine metabolizing enzymes: Results from a prospective pharmacological study

Thiopurine S-Methyltransferase Pharmacogenetics: Genotype to Phenotype Correlation in the Slovenian Population

Contact Info

Product

Resources

About