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Background: 6-mercaptopurine has been a standard component of long-term continuing treatment for childhood lymphoblastic leukaemia, whereas 6-thioguanine has been mainly used for intensification courses. Since preliminary data have shown that 6-thioguanine is more effective than 6-mercaptopurine, we compared the efficacy and toxicity of the two drugs for childhood lymphoblastic leukaemia. Methods: Consecutive children with lymphoblastic leukaemia diagnosed in the UK and Ireland between April, 1997, and June, 2002, were randomly assigned either 6-thioguanine (750 patients) or 6-mercaptopurine (748 patients) during interim maintenance and continuing therapy. All patients received 6-thioguanine during intensification courses. We analysed event-free and overall survival on an intention-to-treat basis. We obtained toxicity data using an adverse-event reporting system, with follow-up questionnaires to seek detailed information for specific toxicities. This trial is registered with the International Standard Randomised Controlled Number 26727615 with the name ALL97. Findings: After a median follow up of 6 years, there was no difference in event-free or overall survival between the two treatment groups. Although 6-thioguanine conferred a significantly lower risk of isolated CNS relapse than did 6-mercaptopurine (odds ratio [OR] 0.53, 95% CI 0.30-0.92, p=0.02), the benefit was offset by an increased risk of death in remission (2.22, 1.20-4.14, p=0.01), mainly due to infections during continuing therapy. Additionally, 95 patients developed veno-occlusive disease of the liver. Of these, 82 were randomly assigned 6-thioguanine, representing 11% of all 6-thioguanine recipients. On long-term follow-up, about 5% of 6-thioguanine recipients have evidence of non-cirrhotic portal hypertension due to periportal liver fibrosis or nodular regenerative hyperplasia. Interpretation: Compared with 6-mercaptopurine, 6-thioguanine causes excess toxicity without an overall benefit. 6-mercaptopurine should remain the thiopurine of choice for continuing therapy of childhood lymphoblastic leukaemia.

AimsThe response to 6‐mercaptopurine (6MP) is highly variable. Its antileukaemic effect can be related to drug derived 6‐thioguanine nucleotides (TGNs). The inherited level of thiopurine methyltransferase (TPMT) activity may be a major factor in the clinical response to 6MP because TPMT forms methylmercaptopurine metabolites (MeMPs) at the expense of TGNs. The aim of this study was to explore the clinical importance of TPMT phenotype.

Summary. We have examined the factors influencing prognosis in over 4000 children with acute lymphoblastic leukaemia (ALL) aged 1–14 who have been treated on consecutive MRC UKALL trials from 1972 to 1990. During this time the results of treatment have improved steadily but are consistently superior in girls when compared with boys; the 5‐year event‐free survival in girls improving from 51% to 71% and in boys from 31% to 57%. These results were independent of age and presenting leucocyte count. Boys not only had a testicular relapse rate of 10% but an excess of bone marrow relapse, particularly evident after 2 years from diagnosis. Other prognostic factors included organomegaly and the morphology of leukaemic blast cells; immuno‐phenotype of the leukaemia, however, had no independent significance after allowance for age, sex and leucocyte count.

In the M.R.C. UKALL VIII study, 35 of 199 (17.6%) patients on protocol developed interstitial pneumonitis within the first 44 weeks of treatment (median 10 weeks). These episodes were carefully documented, and serological investigation for Pneumocystis carinii infection was performed in 33 cases. These 35 patients were divided into two groups, one having either histological or serological evidence of P. carinii infection and the other without such evidence. We saw no false positive serological tests in the nine cases with histological confirmation, but the P. carinii immunofluorescence test was of most help in retrospectively confirming P. carinii infection. High dose trimethoprim-sulphamethoxazole was successful in treating 26 of 33 cases (78%). We did note an improvement in survival in children given steroids and we suggest they may have a role in anti-pneumonitis therapy.

The effect of folic acid supplements on 6-mercaptopurine remission maintenance therapy in lymphoblastic leukaemia (ALL) was investigated in a retrospective longitudinal study of 10 children. Red cell concentrations of 6-thioguanine nucleotide, a cytotoxic metabolite of 6-mercaptopurine, were measured and the peripheral neutrophil count was used as an index of myelosuppression. During the control period of the study there were significant correlations between 6-mercaptopurine dose and 6-thioguanine nucleotide concentration (rs = 0.59, P less than 0.0005) and between 6-thioguanine nucleotide concentration and the peripheral neutrophil count at 14 days (rs = 0.58, P less than 0.0005). These relationships were absent when the same children were subsequently taking folate supplements. Also when taking folate supplements the children tolerated significantly more 6-mercaptopurine (P less than 0.005) for a significantly longer time (P less than 0.005) before neutropenia developed. There was no significant difference in red cell 6-thioguanine nucleotide concentration in the absence and presence of folate supplements. These findings suggest that folate supplements may interfere with remission maintenance therapy in ALL.

Pretreatment marrow blast cells were studied in 38 boys and 27 girls (aged 1-14) with acute lymphoblastic leukaemia by flow cytometry after staining with propidium iodide.The percentage of blast cells in the S phase of the cell cycle ranged from 1% to 40% (median 6%). A correlation was found between the percentage of cells in S and the morphological classification of the French American British Cooperative Group (FAB), presence of T or B cell markers, haemoglobin concentration, blast size, bone pain, platelet count, and an inverse correlation with coarse granule and block staining with Periodic-acid-Schiff (PAS).63 of the 65 children attained complete remission. During the first 24 months of follow up there were fewer relapses (P = 0·054), and deaths (P = 0·004) in those children with 6% or fewer blasts in S phase. The difference was most marked in the first 12 months with 4 relapses out of 33 in the group with 6% or fewer cells in S compared with 13/30 in the group with > 6% cells in S.In order to investigate the prognostic significance of the pretreatment proliferative studies in greater detail, remission duration was correlated with 17 presenting features. Each feature was correlated individually and then the simultaneous effect of all the features was assessed by stepwise multiple regression.Only 3 features of the disease at diagnosis were individually correlated with duration of remission. These were% cells in S (P < 0·001), log white cell blood count (WBC) (P < 0·01) and the presence of T- or B-cell surface markers (P < 0·05). However, the multiple regression analysis showed that cell markers were not an independent prognostic feature, whereas the percentage cells in S and log WBC were independently and significantly correlated with duration of first remission (P < 0·001 in each case).

Early studies indicated that low immunoglobulin (Ig) concentrations at diagnosis predict a worse outlook for children with acute lymphoblastic leukemia (ALL). We re-examined this suggestion in the light of immunophenotypic disease subtyping and more modern therapy. The concentrations of Ig, IgA, and IgM at diagnosis of ALL in 199 children were reviewed. One hundred forty-one (71%) had normal values and 58 (29%) had at least one abnormal value. No obvious pattern of abnormality relating to disease immunophenotype emerged. Disease-free survival (DFS) was not significantly different when simply comparing children with normal and abnormal values. Those with a white blood cell count lower than 20 × 109/L (n = 106) included 22 with at least one low Ig whose 5-year DFS was inferior to the "normal" group (56% vs 83%; P < 0.005). The 140 children with CD10 + B precursor ALL included 31 with one or more low Ig concentrations who also showed an inferior 5-year DFS to the remainder (55% vs 77%; P < 0.02). Minor abnormalities in Ig concentrations are seen in approximately one third of children with ALL regardless of immunologic subtype. Low values may relate to poor outcome in patients with the disease at otherwise good risk.

The outcome in childhood leukaemia has shown steady improvement over the last decade and efforts are now concentrated on the stratification of patients by risk factors which may avoid overtreatment of good risk patients and limit dose escalation strategies, including those with bone marrow transplantation, to the higher risk patients. In ALL, risk stratification is based on the presenting white cell count, sex, age and cytogenetics of the tumour cells. Even in acute myeloid leukaemia, the outcome with chemotherapy alone is now sufficient to limit elective allogeneic bone marrow transplantation to those who do not have cytogenetically favourable disease. In non-Hodgkins lymphoma, a dramatic improvement in overall survival from 50% to in excess of 80% has been achieved by an escalation in dose and dose intensity of chemotherapy. With this improvement, the prognostic influence of clinical staging has become less clear and recent efforts have concentrated on determining which groups of patients would be cured by less intensive treatment. As for ALL, there is concern about the potential late sequelae in these highly curable children. There remain groups of unusual tumour types, such as anaplastic large cell and peripheral T cell lymphoma, where there remains much to be learned about the pathogenesis and clinical behaviour. The optimum treatment strategy for these subgroups remains to be clarified.

Twelve boys and 10 girls on similar long term remission maintenance treatment for lymphoblastic leukaemia had 79 random assays of their red cell 6 thioguanine nucleotide ( 6TGN ) concentrations performed as an index of cytotoxic activity generated by oral 6-mercaptopurine ( 6MP ). Correlation between the dose of 6MP and 6TGN was statistically significant in the girls (r = 0.58, P less than 0.001) but not in the boys (r = 0.15). Additionally, as a group the boys tolerated more 6MP (P less than 0.05), despite similar prescribing criteria, but this did not result in a higher mean 6TGN concentration or increased myelotoxicity. It appears that girls develop 6MP cytotoxicity at lower doses and more predictably than boys. If so, this may be relevant to the as yet unexplained but marked sex difference in prognosis apparent in some studies.

Of 100 children and adolescents with lymphoblastic leukaemia (ALL) seen over a 6-year period, 25 developed clinically evident infiltration of the central nervous system (CNS), despite early treatment with cranial radiotherapy and intrathecal methotrexate. Nine of these 25 had the features of T ALL, though there were only 17 such patients overall. Not only did those with T ALL get CNS disease more frequently, but they did so much sooner after diagnosis (P less than 0.001) and more commonly had associated facial palsies (P less than 0.05). The tendency to develop CNS infiltration appeared to be significantly related to the possession of T-cell markers (P less than 0.02), but not to the diagnostic white cell count (P = 0.37). These findings suggest that current CNS prophylactic therapy is ineffective in most patients with T ALL.

Fifty consecutive episodes of fever in neutropenic children with malignant disease or aplastic anaemia were randomized to treatment with either ceftazidime alone or a combination of azlocillin and tobramycin, pending the results of bacteriological investigation. More than 90% of organisms isolated from these episodes were sensitive to ceftazidime, which appears to be a non-toxic alternative to aminoglycosides in such circumstances.

Blast cell chromosome abnormalities at presentation in childhood acute lymphoblastic leukemia (ALL) are common, and dijferent patterns are known to be related to outcome. In contrast, the frequency and importance of further changes at the time of relapse remain unclear. Blast cell karyotype evolution was therefore studied in a group of children with recurrent disease. Of 134 consecutive children diagnosed between 1982 and 1992, 31 had a marrow relapse, and 24 had complete cytogenetic studies at both diagnosis and the time of recurrence. Fourteen (58%) of the 24 showed additional chromosomal abnormalities at relapse, 5 (21%) retained abnormalities identical to those seen at diagnosis, and 5 (21 %) remained cytogenetically normal. The 14 with additional changes had shorter first remissions and showed shorter survival after relapse compared with the others. These findings indicate that emergence of cytogenetically recognizable subclones during the progression of childhood ALL could be a marker of more resistant disease.

The clinical usefulness of the measurement of red cell zinc protoporphyrin (ZPP), an indicator of iron-deficient erythropoiesis, was assessed in a group of UK children undergoing investigation for red cell microcytosis. Of 213 children studied, 136 had increased ZPP values. Of these, 86 also had reduced iron stores as indicated by serum ferritin concentration. The 50 children with increased ZPP and normal ferritin values could be divided into two main groups. One group comprised 28 children who had evidence of coexistent infection or inflammatory disease. The other included 21 children who had β-thalassemia trait (n = 19) or disease (n = 2). Among the 77 children with normal ZPP values, 22 had reduced serum ferritin concentrations and 45 did not, nor did they have evidence of β-thalassemia. Microcytosis in some of these children could have been due to α-thalassemia trait. Measurement of ZPP is a simple, quick, and relatively cheap method of confirming the presence of iron-deficient erythropoiesis even when inflammation makes serum ferritin measurements unreliable. It is not as sensitive as the ferritin assay to the early stages of iron deficiency, and its specificity is reduced by the occurrence of raised values in most children with β-thalassemia trait. Where there is microcytosis, normal values, together with normal hemoglobin A2 and serum ferritin concentrations, are likely to indicate α-thalassemia trait.

One hundred and nineteen unselected and similarly treated patients with Ph1-positive chronic granulocytic leukaemia (CGL) had the precise nature of their chromosome rearrangements producing the Ph1 studied to determine whether this had any clinical relevance. Eighteen (15%) did not have the usual 9/22 translocation and these, by life-table analysis, had a significantly shorter benign phase of their disease than the others (P less than 0.01). It further appeared that possession of a non-standard Ph1 was related to age, in that whereas only 24 patients were over 60 at diagnosis, 9 (33%) had a non-9/22 translocation (P less than 0.01). As the duration of the benign phase seemed to be shorter in those over 60 irrespective of Ph1 type (P less than 0.01), the questions arose whether non-standard PhI chromosomes were simply occurring in older patients or whether they were affecting prognosis independently. Their independent effect was suggested by the 11 patients under 60 with a non-9/22 Ph1 who still had a significantly shorter benign phase than the 84 of similar age with a standard Ph1 (P less than 0.01). It is concluded that the myeloid karyotype can provide prognostic as well as diagnostic information in patients with CGL.

6-Mercaptopurine (6MP) pharmacokinetics and red blood cell 6-thioguanine nucleotide (TGN) concentrations were studied in 19 children receiving remission maintenance treatment for lymphoblastic leukemia. There was a high interpatient variation in all the pharmacokinetic parameters measured. The pharmacokinetic parameters measured in two children who subsequently had relapses were within the 95% confidence limits of the 17 other children. There was no difference in 6MP pharmacokinetic parameters with respect to neutropenia either after or before the study. The children who developed neutropenia 10 to 19 days after study had significantly higher TGN concentrations (U = 8; P < 0.001) and had spent a longer time receiving reduced 6MP dosage in the 12 weeks before the study (U = 19.5; P < 0.025). TGN concentrations are a better index of a child's ability to form active cytotoxic metabolites than 6MP dose or plasma concentrations.

Aims The response to 6-mercaptopurine (6MP) is highly variable. Its antileukaemic effect can be related to drug derived 6-thioguanine nucleotides (TGNs). The inherited level of thiopurine methyltransferase (TPMT) activity may be a major factor in the clinical response to 6MP because TPMT forms methylmercaptopurine metabolites (MeMPs) at the expense of TGNs. The aim of this study was to explore the clinical importance of TPMT phenotype.

We studied the influence of patient, leukaemia and treatment characteristics on the kinetics of Minimal Residual Disease (MRD) clearance in children with lymphoblastic leukaemia treated using an intensive risk stratified approach. UK MRC protocol ALL97 (1997–1999), and its amended version ALL 97/99 (1999–2002), compared the efficacy and toxicity of dexamethasone (DEX) with prednisolone (PRED), and 6-thioguanine (TG) with 6-Mercaptopurine (MP) in a randomised fashion. The trial produced a 5 year event-free survival (EFS) of 80%, with better systemic and Central Nervous System outcomes in DEX compared with PRED recipients but no difference between TG and MP recipients. Several changes to the risk stratification and treatment regimens during the period of the trial provided an opportunity to determine their impact on MRD clearance. We compared this with clearance in those treated on the successor trial ALL 2003 (more intensive induction containing DEX and Pegylated Asparaginase). The variables investigated for their potential influence on MRD status at the end of induction (EOI) were: NCI Risk; Asparaginase intensity (Erwinia Asparaginase [ERW] in ALL 97 and early part of ALL97/99 vs native or Pegylated E. Coli Asparaginase [E. Coli] in later part of ALL 97/99 and ALL2003); DEX vs PRED; and marrow response at day 8/15 of induction (Slow Early Response [SER] >25% blasts vs Rapid Early Response [RER] ⩽ 25% blasts). MRD was assessed using either a semi-quantitative sequence-specific PCR (ALL97) or Real-Time Quantitative PCR (ALL99 and ALL 2003) of antigen receptor gene re-arrangements at EOI. MRD status was defined as NEG if no MRD was detected by two markers sensitive to 10−4; POS if > 10−4, and Positive Outside Quantitative Range (POQR) if positive < 10−4. Results were available from retrospective testing in 66 ALL97 and 76 ALL97/99 patients, and 204 ALL2003 patients monitored prospectively. There was no significant difference in the proportions of patients MRD NEG, POS or POQR in steroid or NCI sub-groups. Significantly more ERW Asparaginase recipients were MRD POS compared with E.Coli (p< 0.0005). Although SER was predictive of slow MRD clearance, 80/235 (34%) RERs were MRD POS and 9/27(33%) SERs were MRD NEG. Relapse risk at 4 years is 8% in NEG v 28% in POS group in ALL97, 0% in NEG/POQR, versus 15% in POS group in ALL97/99 (p=.001) 2 of the ALL2003 patients have relapsed; one was POS and the other POQR. In the context of intensive risk stratified treatment of childhood ALL, type of Asparaginase influenced rapidity of MRD clearance, whilst NCI risk and type of steroid had no significant influence. Patients with detectable MRD < 10−4 at EOI have the same low risk of relapse as those who have undetectable MRD.

The UK MRC ALL97 trial included a randomised comparison of the efficacy and toxicity of dexamethasone and thioguanine (TG, experimental arms) with prednisolone and mercaptopurine (MP). The cytotoxic effects of MP and TG are mediated, in part, by thioguanine nucleotide (TGN) metabolites. TGNs are formed directly from TG whereas MP produces intermediate metabolites which are substrates for the enzyme thiopurine methyltransferase (TPMT). TPMT is under the control of a common genetic polymorphism. Both drugs are good substrates for TPMT. Both drugs undergo oxidative metabolism, but TG requires prior deamination. Consecutive children with ALL, diagnosed in the United Kingdom and Ireland between April 1997 and June 2002, were randomised to receive either TG or MP in maintenance courses. All patients received TG in intensification courses. Toxicity data was collected by an adverse event reporting system with follow-up questionnaires to seek detailed information on specific toxicities. Red blood cell (RBC) TGN concentrations and TPMT activities were measured by standard techniques in specific sub-groups. Of 1498 randomised patients, 750 were allocated TG and 748 MP. The trial produced a 5 year event-free survival (EFS) of 80%, with no difference in EFS or overall survival between thiopurine arms. However, TG was associated with an acute hepatitis syndrome with features of veno-occlusive disease (VOD) in 95 patients. Eighty-two of these were in the randomised TG arm (11% of TG recipients), the remaining were patients allocated MP who developed VOD during TG intensification courses. On long term follow-up, 43 (5.7%) TG recipients developed persistent splenomegaly and mild to moderate thrombocytopenia due to portal hypertension. TPMT activities were measured in 73 of the 95 VOD children at a standard protocol dose of thiopurine. For comparison, TPMT activities were measured in children who did not develop VOD or splenomegaly; there was no difference in the range of TPMT activities measured at 40mg/m2 TG (n = 83, median 15.4 units/ml RBCs) compared to 75mg/m2 MP (n = 78, median 15.2 units). Median TPMT activities in these 161 children, taking thiopurines at the protocol standard dose, were 15.2 units (range 5.8 to 23) compared to a median of 13.4 units (range 5.8 to 23) in the 73 VOD children (median difference 1.8 units, 95% CI 0.9 to 2.7, p < 0.001). A similar difference in TPMT activity was also seen when comparing persistent splenomegaly cases with controls. There was no difference in accumulation of TGNs at comparable doses. When used for maintenance therapy, TG causes liver toxicity in a significant minority of children with ALL without improving long term survival. An association with polymorphic variations in TPMT levels, a critical enzyme in thiopurine metabolism, implicates inter-individual variations in metabolism of the drug as a mechanism for liver damage, and points to a possible means of identifying at-risk patients. MP should remain the standard thiopurine for maintenance therapy unless longer follow-up reveals a survival advantage in favour of TG, and the mechanisms of liver toxicity are understood to avoid exposure of at risk patients.