Ecto-5'-nucleotidase (ecto-5'NT) catalyzes the extracellular dephosphorylation of nucleotides like IMP. Cytoplasmic 5'NT (cyto-5'NT) and non-specific (e.g. acid- and alkaline) phosphatases (AP) regulate the intracellular degradation of nucleotides. High NT and AP activities might cause a resistance to the thiopurines 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG). We studied the relation between these enzymes and immunophenotype, drug resistance and prognosis in 77 children with acute lymphoblastic leukemia (ALL). Enzyme activities were assessed radiochemically; in vitro drug resistance was measured with the MTT assay. AP activities were higher in T-ALL and B-ALL than in precursor B-ALL. Cyto-5'NT activity was very low in all phenotypes and accounted for a significant proportion of total IMPase activity only in the very immature CD10- c mu- precursor B-ALL. CD10+ ALL cases with high ecto-5'NT activities showed a trend (p = 0.065) for a lower probability of continuous complete remission than those with a low activity. Ecto-5'NT activity was not related to in vitro drug resistance to 6-TG. A weak correlation was found between in vitro 6-TG resistance and cyto-5'NT and AP activities. We conclude that high ecto-5'NT activities do not cause a resistance to 6-thiopurines in childhood ALL. Some patients have high cyto-5'NT and AP activities associated with 6-thiopurine resistance.