Koichiro Ikuta

Six Months of Maintenance Chemotherapy After Intensified Treatment for Acute Lymphoblastic Leukemia of Childhood

PURPOSE We postulated that intensification of chemotherapy immediately after remission induction might reduce the leukemic cell burden sufficiently to allow an abbreviated period of antimetabolite therapy. PATIENTS AND METHODS Three hundred forty-seven children (ages 1 to 15 years) with previously untreated acute lymphoblastic leukemia (ALL) were enrolled onto the Tokyo L92-13 study, which excluded patients with mature B-cell ALL and patients less than 1 year old. One hundred twenty-four patients were classified as standard risk, 122 as high risk, and 101 as extremely high risk, according to age, peripheral-blood leukocyte count, selected genetic abnormalities, and immunophenotype. All subjects received four drugs for remission induction, followed by a risk-directed multidrug intensification phase and therapy for presymptomatic leukemia in the CNS. Maintenance chemotherapy with oral mercaptopurine and methotrexate was administered for 6 months, with all treatment stopped by 1 year after diagnosis. RESULTS The mean (+/- SD) event-free survival (EFS) and overall survival rates for all patients were 59.5% +/- 3.4% and 81.5% +/- 2.2%, respectively, at 5. 5 years after diagnosis. EFS rates by risk category were similar (60. 2% +/- 6.0% for standard risk, 57.7% +/- 5.6% for high risk, and 62. 5% +/- 5.7% for extremely high risk), whereas overall survival rates differed significantly (91.2% +/- 2.7%, 80.0% +/- 4.1%, and 72.1% +/- 4.5%, respectively, P <.0001 by the log-rank test). There were 107 relapses. Eighty-five (79.4%) of these 107 patients achieved second complete remissions, with subsequent EFS rates of 61.5% +/- 7. 9% (standard risk), 42.6% +/- 8.1% (high risk), and 9.6% +/- 6.4% (extremely high risk). Of the five risk factors analyzed, only the response to prednisolone monotherapy among extremely high-risk patients proved important. CONCLUSION Early treatment intensification did not compensate for a truncated phase of maintenance chemotherapy in children with standard- or high-risk ALL. However, 6 months of antimetabolite treatment seemed adequate for extremely high-risk patients who were good responders to prednisolone and received intensified chemotherapy that included high-dose cytarabine early in the clinical course.

Significance of the complete clearance of peripheral blasts after 7 days of prednisolone treatment in children with acute lymphoblastic leukemia: the Tokyo Children’s Cancer Study Group Study L99-15

Early treatment response is one of the most useful prognostic indicators in childhood acute lymphoblastic leukemia. This study adds novel information that patients whose peripheral blood blasts disappeared after 7 days of prednisolone monotherapy had an excellent prognosis, that is, a 4-year event-free survival of 90%. See related perspective article on page 1124. Background Treatment response has become one of the most important prognostic factors in childhood acute lymphoblastic leukemia. We evaluated the significance of the complete clearance of peripheral leukemic blasts on survival in children with acute lymphoblastic leukemia. Design and Methods Seven hundred and fifty-four children diagnosed with acute lymphoblastic leukemia, consecutively enrolled from 1999 to 2003 in the TCCSG L99-15 study, were eligible for analysis. Patients were stratified into three risk groups based on presenting features, such as age and the leukocyte count before starting the treatment, followed by reclassification into three categories 7 days after prednisolone monotherapy based on the peripheral blast count; 0/μL (Day8NoBlasts), 1-999/μL and ≥ 1,000/μL. Results After 7 days of prednisolone monotherapy, 249 patients (33%) were classified as Day8NoBlasts, 392 patients (52%) had blast counts of 1-999/μL, and 113 patients (15%) had blast counts ≥ 1,000/μL. The event-free survival for all patients was 79.6±1.6 (SE)% at 4 years, whereas that for patients with Day8NoBlasts was 90.4±2.0% (n=249) and the event-free survival for the other patients was 74.2±2.2% (n=504) (log rank p<0.001). The event-free survival for Day8NoBlasts patients with B-lineage acute lymphoblastic leukemia and T-cell acute lymphoblastic leukemia was 89.8±2.1% (n=226) and 95.7±4.3% (n=23), respectively. In a multivariate analysis, age at diagnosis, the initial white blood cell count, immunophenotype, and gender did not remain as independent risk factors for treatment failure, whereas Day8NoBlasts and marked hyperdiploidy (more than 50 chromosomes) became statistically significant. Conclusions Children with Day8NoBlasts constituted one third of all the cases with childhood acute lymphoblastic leukemia with an excellent outcome, and should be candidates for curative management with less intensive treatment.

Myelodysplastic syndrome in childhood: a retrospective study of 189 patients in Japan.

We report a retrospective analysis of children with myelodysplastic syndrome (MDS) diagnosed between 1990 and 1997 in Japan. In total, 189 patients were enrolled: 122 cases of primary MDS (26 RA, 18 RAEB, 25 RAEBt, 53 CMML/JMML), 24 cases with constitutional predisposition to MDS, and 43 cases of therapy-related MDS (t-MDS). The frequency of pediatric MDS was estimated to be 7.7% of all leukemias. Cytogenetic abnormalities were observed in 41% of primary MDS and 90% of t-MDS cases. The 4-year survival rate, estimated by Kaplan–Meier analysis, for primary RA was 78.9%, while other types of MDS and JMML had rates lower than 40%, and t-MDS showed an even more unfavorable prognosis. In primary MDS, the survival rate of patients with cytogenetic abnormalities was significantly lower. Among prognostic variables by IPSS, only the cytogenetic pattern was useful for predicting outcome in childhood MDS. There was no apparent advantage to chemotherapy for RA, and the survival rate in patients with primary RA, JMML, or t-MDS receiving stem cell transplantation was significantly higher. More precise designs of our diagnostic and classification systems, as well as therapeutic trials in large-scale prospective studies, are necessary for further improvements in MDS outcome.

Clinical significance of early T‐cell precursor acute lymphoblastic leukaemia: results of the Tokyo Children’s Cancer Study Group Study L99‐15

Early T‐cell precursor acute lymphoblastic leukaemia (ETP‐ALL) is a recently identified subtype of T‐ALL with distinctive gene expression and cell marker profiles, poor response to chemotherapy and a very high risk of relapse. We determined the reliability of restricted panel of cell markers to identify EPT‐ALL using a previously classified cohort. Then, we applied the cell marker profile that best discriminated ETP‐ALL to a cohort of 91 patients with T‐ALL enrolled in the Tokyo Children’s Cancer Study Group L99‐15 study, which included allogeneic stem cell transplantation (allo‐SCT) for patients with poor prednisone response. Five of the 91 patients (5·5%) met the ETP‐ALL criteria. There were no significant differences in presenting clinical features between these and the remaining 86 patients. Response to early remission induction therapy was inferior in ETP‐ALL as compared with T‐ALL. The ETP‐ALL subgroup showed a significantly poorer event‐free survival (4‐year rate; 40%) than the T‐ALL subgroup (70%, P = 0·014). Of note, three of four relapsed ETP‐ALL patients survived after allo‐SCT, indicating that allo‐SCT can be effective for this drug‐resistant subtype of T‐ALL.

Long-term follow-up of childhood acute lymphoblastic leukemia in Tokyo Children's Cancer Study Group 1981-1995.

The objectives were as follows: Firstly, to estimate the overall probability of event-free survival (EFS) and isolated CNS relapse in the studies for children with acute lymphoblastic leukemia (ALL) during the 1980s and 1990s. Secondly, to report the EFS according to presenting features and lineage. Thirdly, to evaluate the treatment results re-classified by the risks of NCI criteria. Four consecutive protocol studies were performed in the Tokyo Childrens Cancer Study Group: L81–10 protocol (1981–1984, 189 patients), L84–11 (1984–1989, 484 patents), L89–12 (1989–1992, 418 patients) and L92–13 (1992–1995, 347 patients). Overall EFS at 5 years in each protocol was 56.5 ± 3.8(1 s.e.)%, 71.0 ± 2.1%, 67.8 ± 2.3%, and 63.4 ± 2.7%, respectively. The cumulative isolated CNS relapse rate at 5 years was 8.1 ± 2.1%, 3.5 ± 0.9%, 3.6 ± 1.0%, 1.0 ± 0.6. The EFS in SR/HR (standard risk/high risk) according to the NCI criteria in B-precursor ALL at 5 years was 61.9 ± 4.3%/41.4 ± 7.4% (lineage was not confirmed.), 72.5 ± 2.6%/63.4 ± 5.0%, 77.4 ± 2.7%/56.3 ± 4.7%, and 67.8 ± 3.4%/56.7 ± 5.4% in each protocol. Also EFSs according to NCI SR/HR at 5 years of T-ALL in protocols L84–11, L89–12 and L92–13 were 55.6 ± 16.6%/60.9 ± 10.1%, 72.7 ± 13.4%/51.6 ± 9.1%, and 77.1 ± 14.4%/53.6/10.1%, respectively. The truncation of maintenance therapy to 6 months resulted in a decreased EFS in L92–13, particularly due to an increase of bone marrow relapse after cessation of therapy in SR and HR. The NCI risk criteria work properly even in the patients treated by different intensities, so that it makes the comparison possible among the patients in various groups. The overall EFSs in childhood ALL improved in 1980s, but it seemed stable or decreased in 1990s. The short maintenance therapy resulted in poor outcome in SR on the L92–13 protocol. Many of these late relapsers were effectively rescued and overall survival remained at a high level. The proportion of patients who received cranial irradiation reduced without any increase of the CNS events.

Cord blood transplantation from HLA‐mismatched unrelated donors as a treatment for children with haematological malignancies

Factors influencing the outcome for 39 children with haematological malignancy who were subjected to a cord blood transplantation (CBT) from genotypically HLA‐mismatched unrelated donors were analysed. This retrospective study included 21 children with acute lymphoblastic leukaemia, 15 with acute myelogenous leukaemia and one each with chronic myelogenous leukaemia, refractory anaemia with myelodysplastic syndrome (MDS) and juvenile myelomonocytic leukaemia (JMML). Those subjected to CBT during the first or second complete remission (CR) and MDS without blasts were assigned to the standard‐risk (SR) group (n = 16). Patients in third or subsequent remission, relapse or partial remission with refractory leukaemia at the time of CBT were considered to be in advanced phase, and placed in the high‐risk (HR) group (n = 11). JMML and the second CR after a relapse (n = 8), or bone marrow failure after a rejection (n = 3), following haematopoietic stem cell transplantation (HSCT) in the first CR were included in the high‐risk group. Kaplan–Meier estimates for neutrophil and platelet recovery were 83·7 ± 12·2 at d 60 and 55·4 ± 16·6% at d 100 respectively. The incidence of grades II–VI acute graft‐versus‐host disease was 58·5 ± 16·8%. The Kaplan–Meier estimate for 3‐year event‐free survival (EFS) was 49·2 ± 16·6. From multivariate analysis, the most important factor influencing EFS was disease status at CBT: SR patients had a 3‐year EFS of 75·0 ± 21·6%, compared with 29·6 ± 20·6% for those with HR disease (P = 0·013, RR 4·746, 95% CI 1·382–16·298). These data confirm that HLA‐mismatched, unrelated CBT is a feasible procedure to cure a significant proportion of children with leukaemia, especially if conducted in a favourable phase of the disease.

N-(4-Hydroxyphenyl)retinamide (4-HPR) induces leukemia cell death via generation of reactive oxygen species.

The role of reactive oxygen species (ROS) in the cytotoxicity of N-(4-hydroxyphenyl)retinamide (4-HPR) was studied with use of the B-precursor lymphoblastic leukemia cell line YCUB-2. The increase in intracellular ROS measured with 2′-7′-dichlorodihydrofluorescein diacetate after 3 hours’ incubation was 3.7-fold with 1 μM 4-HPR and 5.8-fold with 5 μM 4-HPR. The rate of apoptosis after 48 hours’ incubation was 9.8% and 56.4% in comparison with untreated cells. Hydroethidine, which is a more specific indicator of superoxide anion radical level, did not effectively detect 4-HPR-induced ROS. The antioxidant 3-methyl-1-phenyl-2-pyrazolin-5-one suppressed 4-HPR-induced ROS production and apoptosis. The cytotoxicity of 4-HPR was analyzed in 4 other leukemia/lymphoma lines (CCRF-HSB2, Molt-4, KG-1, HL-60). We found that the cytotoxicity of 4-HPR correlated with the amount of ROS produced in cell lines, except in HL-60 cells. The intracellular glutathione level varied among the 5 cell lines, the highest levels occurring in Molt-4 and KG-1, which were less sensitive to 4-HPR. Suppression of glutathione by buthionine sulfoximine enhanced the level of 4-HPR-induced ROS production and apoptosis in Molt-4. Our findings suggest that ROS play a significant role in the antileukemia effect of 4-HPR and that the glutathione level in leukemias may be associated the sensitivity of the cells to 4-HPR.

Moyamoya syndrome following childhood acute lymphoblastic leukemia.

Long‐term survivors of childhood acute lymphoblastic leukemia (ALL) sometimes suffer from adverse long‐term sequelae. We analyzed the incidence, clinical course and prognosis of moyamoya syndrome (MoS) following childhood ALL.

Pure red cell aplasia after major ABO‐incompatible bone marrow transplantation: two case reports of treatment with recombinant human erythropoietin

A 34-year-old man with acute myelocytic leukemia (AML: MO) and a 32-year-old woman with AML: M2 developed pure red cell aplasia (PRCA) after receiving a major ABO-incompatible bone marrow transplant (BMT). The first patient responded to recombinant human erythropoietin (rhEPO) therapy, while the second did not. The second patient also received methylprednisolone (m-PSL) but developed reticulocytosis and hemolysis after the administration of m-PSL. Plasmapheresis was then performed and the patient promptly recovered from hemolysis and PRCA. We conclude that close attention must be paid when treating PRCA following major ABO-in-compatible BMT with rhEPO and m-PSL, as there is always the potential for massive hemolysis.

Effects of erythropoietin, IL-3, IL-6 and LIF on a murine megakaryoblastic cell line : growth enhancement and expression of receptor mRNAs

We examined the effects of recombinant human erythropoietin (rhEPO), recombinant murine interleukin 3 (rmIL-3), recombinant human interleukin 6 (rhIL-6), recombinant human interleukin 11 (rhIL-11), recombinant murine leukemia inhibitory factor (rmLIF) and recombinant murine granulocyte-macrophage colony-stimulating factor (rmGM-CSF) on the growth of murine megakaryocytic cell lines. In serum-free methylcellulose culture supplemented with bovine serum albumin (BSA), the addition of rhEPO (0.1-10 U/ml), rmIL-3 (10-500 U/ml), rhIL-6 (100-10,000 U/ml), rmLIF (100-10,000 U/ml), or rmGM-CSF (10-1000 U/ml) enhanced colony growth in L8057Y5 cells, which had been maintained in protein-free culture, mostly in a dose-dependent fashion; rhIL-11 did not have any stimulatory effect at the tested doses (10-1000 U/ml). In addition, colony growth of L8057 cells, which had been maintained in serum-containing culture, was enhanced, but to a lesser extent, by the addition of these cytokines except rhEPO (the cultures were supplemented with 1% fetal bovine serum. Among the cytokines that showed growth-enhancing effects on L8057 cells, the expression of mRNAs encoding receptors for EPO, IL-6 and IL-3 was examined by northern blot analysis or reverse transcription polymerase chain reaction (RT-PCR). In both cell lines, mRNAs for EPO-R, IL-6R, gp130, IL-3R alpha and beta chains were constitutively expressed. The results suggest that L8057 and L8057Y5 cell lines have characteristics of megakaryoblastic cells in their biological responses to cytokines, as well as in the expression of cytokine receptor mRNAs, and that the growth-enhancing effects of these cytokines on the cell lines may be achieved through specific receptors. Our findings show the value of these cell lines for investigating the mechanisms of growth signal transduction in megakaryopoiesis.