Cancer Chemotherapy and Pharmacology

, Volume 66, Issue 3, pp 485–491 | Cite as

DNA incorporation of 6-thioguanine nucleotides during maintenance therapy of childhood acute lymphoblastic leukaemia and non-Hodgkin lymphoma

  • Rikke L. Hedeland
  • Kristian Hvidt
  • Jacob Nersting
  • Susanne Rosthøj
  • Kim Dalhoff
  • Birgitte Lausen
  • Kjeld SchmiegelowEmail author
Original Article



To explore the DNA incorporation of 6-thioguanine nucleotide levels (DNA-6TGN) during 6-mercaptopurine (6MP) therapy of childhood acute lymphoblastic leukaemia (ALL) and non-Hodgkin lymphoma (NHL) and its relation to erythrocyte levels of their metabolites: 6-thioguanine-nucleotides (E-6TGN), methylated metabolites (E-MeMP), Methotrexate polyglutamates (E-MTX), and to thiopurine methyltransferase activity (TPMT).


We studied these metabolites in 229 blood samples from 18 children with ALL (N = 16) or NHL (N = 2) on 6MP/Methotrexate maintenance therapy.


DNA-6TGN levels were significantly correlated to E-6TGN (r p = 0.66, p = 0.003) with a trend to reach a plateau at high E-6TGN levels. To explore the relative DNA incorporation of 6TGN in relation to cytosol 6TGN levels, a DNA-6TGN index was calculated as DNA-6TGN/E-6TGN. The DNA-6TGN index was inversely correlated to E-6TGN (r p = −0.58, p = 0.012), which implies that with increasing levels of E-6TGN relatively less 6TGN are incorporated into DNA. E-MeMP levels were correlated to the DNA-TGN index (r p = 0.60, p = 0.008), indicating that high levels of MeMP result in enhanced DNA-6TGN incorporation, possibly due to inhibition of purine de novo synthesis, mediated by some of the methylated 6MP metabolites.


DNA-6TGN may prove to be a more relevant pharmacokinetic parameter for monitoring 6MP treatment intensity than the previously used erythrocyte 6MP metabolites levels. Prospective clinical trials are needed to evaluate the usefulness of DNA-6TGN for individual dose adjustments.


Acute lymphoblastic leukaemia Non-Hodgkin lymphoma Maintenance therapy Pharmacokinetics Paediatric cancer Paediatric pharmacology 



This study has received financial support from H:S Research Fund 2000 (Grant no. 120), the Danish Cancer Society (Grant no. 99144069132), the Danish Childhood Cancer Foundation, The University Hospital Rigshospitalet, Novo Nordic Foundation, Home Secretary Research Grant for Individualised Therapy, and Danish Research Council for Health and Disease (Grant no. 271-0615). Kjeld Schmiegelow holds the Childhood Cancer Foundation Research Professorship in Pediatric Oncology. A special thanks to Jens Bo Thomsen, who has performed the DNA-6TGN analysis and data registration.

Conflict of interest statement



  1. 1.
    Schmiegelow K, Gustafsson G (2005) Acute lymphoblastic leukemia. In: Caron M (ed) Cancer in children. Oxford University Press, Oxford, pp 138–170Google Scholar
  2. 2.
    Schmiegelow K, Pulczynska MK (1990) Maintenance chemotherapy for childhood acute lymphoblastic leukemia: should dosage be guided by white blood cell counts? Am J Pediatr Hematol Oncol 12:462–467CrossRefPubMedGoogle Scholar
  3. 3.
    Arico M, Baruchel A, Bertrand Y, Biondi A, Conter V, Eden T, Gadner H, Gaynon P, Horibe K, Hunger SP, Janka-Schaub G, Masera G, Nachman J, Pieters R, Schrappe M, Schmiegelow K, Valsecchi MG, Pui CH (2005) The seventh international childhood acute lymphoblastic leukemia workshop report: Palermo, Italy, January 29–30, 2005. Leukemia 19:1145–1152CrossRefPubMedGoogle Scholar
  4. 4.
    Uribe-Luna S, Quintana-Hau JD, Maldonado-Rodriguez R, Espinosa-Lara M, Beattie KL, Farquhar D, Nelson JA (1997) Mutagenic consequences of the incorporation of 6-thioguanine into DNA. Biochem Pharmacol 54:419–424CrossRefPubMedGoogle Scholar
  5. 5.
    Bokkerink JP, Stet EH, De Abreu RA, Damen FJ, Hulscher TW, Bakker MA, van Baal JA (1993) 6-Mercaptopurine: cytotoxicity and biochemical pharmacology in human malignant T-lymphoblasts. Biochem Pharmacol 45:1455–1463CrossRefPubMedGoogle Scholar
  6. 6.
    Schmiegelow K, Forestier E, Kristinsson J, Soderhall S, Vettenranta K, Weinshilboum R, Wesenberg F (2009) Thiopurine methyltransferase activity is related to the risk of relapse of childhood acute lymphoblastic leukemia: results from the NOPHO ALL-92 study. Leukemia 23:557–564CrossRefPubMedGoogle Scholar
  7. 7.
    Wang L, Weinshilboum R (2006) Thiopurine S-methyltransferase pharmacogenetics: insights, challenges and future directions. Oncogene 25:1629–1638CrossRefPubMedGoogle Scholar
  8. 8.
    Lilleyman JS, Lennard L (1994) Mercaptopurine metabolism and risk of relapse in childhood lymphoblastic leukaemia. Lancet 343:1188–1190CrossRefPubMedGoogle Scholar
  9. 9.
    Erb N, Harms DO, Janka-Schaub G (1998) Pharmacokinetics and metabolism of thiopurines in children with acute lymphoblastic leukemia receiving 6-thioguanine versus 6-mercaptopurine. Cancer Chemother Pharmacol 42:266–272CrossRefPubMedGoogle Scholar
  10. 10.
    Relling MV, Hancock ML, Boyett JM, Pui CH, Evans WE (1999) Prognostic importance of 6-mercaptopurine dose intensity in acute lymphoblastic leukemia. Blood 93:2817–2823PubMedGoogle Scholar
  11. 11.
    Schmiegelow K, Schroder H, Gustafsson G, Kristinsson J, Glomstein A, Salmi T, Wranne L (1995) Risk of relapse in childhood acute lymphoblastic leukemia is related to RBC methotrexate and mercaptopurine metabolites during maintenance chemotherapy. Nordic Society for Pediatric Hematology and Oncology. J Clin Oncol 13:345–351PubMedGoogle Scholar
  12. 12.
    Lennard L (2001) Therapeutic drug monitoring of cytotoxic drugs. Br J Clin Pharmacol 52:75–87CrossRefGoogle Scholar
  13. 13.
    Gustafsson G, Schmiegelow K, Forestier E, Clausen N, Glomstein A, Jonmundsson G, Mellander L, Makipernaa A, Nygaard R, Saarinen-Pihkala UM (2000) Improving outcome through two decades in childhood ALL in the Nordic countries: the impact of high-dose methotrexate in the reduction of CNS irradiation. Nordic Society of Pediatric Haematology and Oncology (NOPHO). Leukemia 14:2267–2275CrossRefPubMedGoogle Scholar
  14. 14.
    Miller SA, Dykes DD, Polesky HF (1988) A simple salting out procedure for extracting DNA from human nucleated cells. Nucleic Acids Res 16:1215CrossRefPubMedGoogle Scholar
  15. 15.
    Bruunshuus I, Schmiegelow K (1989) Analysis of 6-mercaptopurine, 6-thioguanine nucleotides, and 6-thiouric acid in biological fluids by high-performance liquid chromatography. Scand J Clin Lab Invest 49:779–784CrossRefPubMedGoogle Scholar
  16. 16.
    Erdmann GR, Steury JC, Carleton BC, Stafford RJ, Bostrom BC, Canafax DM (1991) Reversed-phase high-performance liquid chromatographic approach to determine total lymphocyte concentrations of 6-thioguanine, methylmercaptopurine and methylthioguanine in humans. J Chromatogr 571:149–156CrossRefPubMedGoogle Scholar
  17. 17.
    Kamen BA, Takach PL, Vatev R, Caston JD (1976) A rapid, radiochemical-ligand binding assay for methotrexate. Anal Biochem 70:54–63CrossRefPubMedGoogle Scholar
  18. 18.
    Siegel S, Castellan NJ (1988) Non-parametric statistics for the behavioral science. McGraw-Hill, SingaporeGoogle Scholar
  19. 19.
    Cleveland WS, Devlin SJ (1988) Locally weighted regression: an approach to regression analysis by local fitting. J Am Stat Assoc 83:596–610CrossRefGoogle Scholar
  20. 20.
    Peeters M, Koren G, Jakubovicz D, Zipursky A (1988) Physician compliance and relapse rates of acute lymphoblastic leukemia in children. Clin.Pharmacol Ther 43:228–232PubMedGoogle Scholar
  21. 21.
    Lancaster D, Lennard L, Lilleyman JS (1997) Profile of non-compliance in lymphoblastic leukaemia. Arch Dis Child 76:365–366CrossRefPubMedGoogle Scholar
  22. 22.
    Lennard L, Van Loon JA, Weinshilboum RM (1989) Pharmacogenetics of acute azathioprine toxicity: relationship to thiopurine methyltransferase genetic polymorphism. Clin Pharmacol Ther 46:149–154PubMedGoogle Scholar
  23. 23.
    Relling MV, Hancock ML, Rivera GK, Sandlund JT, Ribeiro RC, Krynetski EY, Pui CH, Evans WE (1999) Mercaptopurine therapy intolerance and heterozygosity at the thiopurine S-methyltransferase gene locus. J Natl Cancer Inst 91:2001–2008CrossRefPubMedGoogle Scholar
  24. 24.
    Cuffari C, Dassopoulos T, Turnbough L, Thompson RE, Bayless TM (2004) Thiopurine methyltransferase activity influences clinical response to azathioprine in inflammatory bowel disease. Clin Gastroenterol Hepatol 2:410–417CrossRefPubMedGoogle Scholar
  25. 25.
    Schmiegelow K, Bjork O, Glomstein A, Gustafsson G, Keiding N, Kristinsson J, Makipernaa A, Rosthoj S, Szumlanski C, Sorensen TM, Weinshilboum R (2003) Intensification of mercaptopurine/methotrexate maintenance chemotherapy may increase the risk of relapse for some children with acute lymphoblastic leukemia. J Clin Oncol 21:1332–1339CrossRefPubMedGoogle Scholar
  26. 26.
    Krynetski EY, Tai HL, Yates CR, Fessing MY, Loennechen T, Schuetz JD, Relling MV, Evans WE (1996) Genetic polymorphism of thiopurine S-methyltransferase: clinical importance and molecular mechanisms. Pharmacogenetics 6:279–290CrossRefPubMedGoogle Scholar
  27. 27.
    Andersen JB, Szumlanski C, Weinshilboum RM, Schmiegelow K (1998) Pharmacokinetics, dose adjustments, and 6-mercaptopurine/methotrexate drug interactions in two patients with thiopurine methyltransferase deficiency. Acta Paediatr 87:108–111CrossRefPubMedGoogle Scholar
  28. 28.
    Bo J, Schroder H, Kristinsson J, Madsen B, Szumlanski C, Weinshilboum R, Andersen JB, Schmiegelow K (1999) Possible carcinogenic effect of 6-mercaptopurine on bone marrow stem cells: relation to thiopurine metabolism. Cancer 86:1080–1086CrossRefPubMedGoogle Scholar
  29. 29.
    Schmiegelow K, Al-Modhwahi I, Andersen MK, Behrendtz M, Forestier E, Hasle H, Heyman M, Kristinsson J, Nersting J, Nygaard R, Svendsen AL, Vettenranta K, Weinshilboum R (2009) Methotrexate/6-mercaptopurine maintenance therapy influences the risk of a second malignant neoplasm after childhood acute lymphoblastic leukemia: results from the NOPHO ALL-92 study. Blood 113:6077–6084CrossRefPubMedGoogle Scholar
  30. 30.
    Harms DO, Gobel U, Spaar HJ, Graubner UB, Jorch N, Gutjahr P, Janka-Schaub GE (2003) Thioguanine offers no advantage over mercaptopurine in maintenance treatment of childhood ALL: results of the randomized trial COALL-92. Blood 102:2736–2740CrossRefPubMedGoogle Scholar
  31. 31.
    Vora A, Mitchell CD, Lennard L, Eden TO, Kinsey SE, Lilleyman J, Richards SM (2006) Toxicity and efficacy of 6-thioguanine versus 6-mercaptopurine in childhood lymphoblastic leukaemia: a randomised trial. Lancet 368:1339–1348CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Rikke L. Hedeland
    • 1
  • Kristian Hvidt
    • 1
  • Jacob Nersting
    • 1
  • Susanne Rosthøj
    • 2
  • Kim Dalhoff
    • 3
  • Birgitte Lausen
    • 1
  • Kjeld Schmiegelow
    • 1
    • 4
    Email author
  1. 1.The Section for Pediatric Hematology and Oncology, Pediatric Clinic II, The Juliane Marie Center, The University HospitalRigshospitaletCopenhagenDenmark
  2. 2.Department of BiostatisticsUniversity of CopenhagenCopenhagenDenmark
  3. 3.Department of Clinical PharmacologyBispebjerg HospitalCopenhagenDenmark
  4. 4.The Faculty of Medicine, Institute of Gynecology, Obstetrics and PediatricsUniversity of CopenhagenCopenhagenDenmark

Personalised recommendations