TY - JOUR
T1 - Role of 5′-nucleotidase in thiopurine metabolism
T2 - Enzyme kinetic profile and association with thio-GMP levels in patients with acute lymphoblastic leukemia during 6-mercaptopurine treatment
AU - Brouwer, Connie
AU - Vogels-Mentink, Trude M.
AU - Keizer-Garritsen, Jenneke J.
AU - Trijbels, Frans J.M.
AU - Bökkerink, Jos P.M.
AU - Hoogerbrugge, Peter M.
AU - Van Wering, Elisabeth R.
AU - Veerman, Anjo J.P.
AU - De Abreu, Ronney A.
N1 - Funding Information:
This study was supported by a grant of the Dutch Cancer Society (KUN97-1485). The authors thank the board members of the Dutch Childhood Oncology Group (DCOG) for approval of this study in the setting of the DCLSG-ALL-9-protocol. The contributions of A. van der Does-van den Berg, director of the DCOG, are greatly acknowledged, as are the coworkers of the DCOG for excellent technical assistance in preparing the blood samples and extensive registration of treatment data. The cooperating centers and hospitals are also acknowledged for supplying patients' material.
PY - 2005/11
Y1 - 2005/11
N2 - Thiopurines are used for treatment of several diseases. Cytotoxicity is caused by the derived compounds 6-thioguanine nucleotides (TGNs) and methyl-6-thioinosine monophosphate (methylthio-IMP). The 6-thiopurine mononucleotides 6-thio-IMP (thio-IMP), 6-thio-GMP (thio-GMP) and methylthio-IMP can be catabolized by purine 5′-nucleotidase. It has been shown that the various 5′-nucleotidases are key enzymes for (6-thio)-purine metabolism. We aimed to investigate whether the overall 5′-nucleotidase (5′NT) activity is correlated with the efficacy and toxicity of 6-thiopurine nucleotides. Substrate affinity of 5′NT for IMP, GMP, AMP, thio-IMP, thio-GMP and methylthio-IMP was studied in human lymphocytes. For each of the substrates, the pH for optimal overall enzyme activity has been determined at a pH range between 6 and 10. At the optimal pH, assays were performed to establish Km and Vmax values. Optimal pH values for the various substrates were between 7 and 8.5. Km values ranged from 33 to 109 μM, Vmax ranged from 3.99 to 19.5 nmol/106 peripheral mononuclear cells (pMNC) h, and Vmax/Km ratios ranged from 105 to 250. The results did not show a distinct preference of 5′NT activity for any of the tested thiopurine nucleotides. The enzyme kinetic studies furthermore revealed substrate inhibition by thio-IMP and thio-GMP as a substrate. Inhibition by thio-GMP also seems to occur in patients treated with 6-mercaptopurine (6 MP); subsequently, this may lead to toxicity in these patients.
AB - Thiopurines are used for treatment of several diseases. Cytotoxicity is caused by the derived compounds 6-thioguanine nucleotides (TGNs) and methyl-6-thioinosine monophosphate (methylthio-IMP). The 6-thiopurine mononucleotides 6-thio-IMP (thio-IMP), 6-thio-GMP (thio-GMP) and methylthio-IMP can be catabolized by purine 5′-nucleotidase. It has been shown that the various 5′-nucleotidases are key enzymes for (6-thio)-purine metabolism. We aimed to investigate whether the overall 5′-nucleotidase (5′NT) activity is correlated with the efficacy and toxicity of 6-thiopurine nucleotides. Substrate affinity of 5′NT for IMP, GMP, AMP, thio-IMP, thio-GMP and methylthio-IMP was studied in human lymphocytes. For each of the substrates, the pH for optimal overall enzyme activity has been determined at a pH range between 6 and 10. At the optimal pH, assays were performed to establish Km and Vmax values. Optimal pH values for the various substrates were between 7 and 8.5. Km values ranged from 33 to 109 μM, Vmax ranged from 3.99 to 19.5 nmol/106 peripheral mononuclear cells (pMNC) h, and Vmax/Km ratios ranged from 105 to 250. The results did not show a distinct preference of 5′NT activity for any of the tested thiopurine nucleotides. The enzyme kinetic studies furthermore revealed substrate inhibition by thio-IMP and thio-GMP as a substrate. Inhibition by thio-GMP also seems to occur in patients treated with 6-mercaptopurine (6 MP); subsequently, this may lead to toxicity in these patients.
KW - 5′-Nucleotidase
KW - 6-Thiopurines
KW - Acute lymphoblastic leukemia
KW - Substrate specificity
UR - https://www.scopus.com/pages/publications/25844503464
U2 - 10.1016/j.cccn.2005.05.006
DO - 10.1016/j.cccn.2005.05.006
M3 - Article
C2 - 15990089
AN - SCOPUS:25844503464
SN - 0009-8981
VL - 361
SP - 95
EP - 103
JO - Clinica Chimica Acta
JF - Clinica Chimica Acta
IS - 1
ER -