Genetics of ancestry-specific risk for relapse in acute lymphoblastic leukemia.

The causes of individual relapses in children with acute lymphoblastic leukemia (ALL) remain incompletely understood. We evaluated the contribution of germline genetic factors to relapse in 2225 children treated on Children's Oncology Group trial AALL0232. We identified 302 germline single-nucleotide polymorphisms (SNPs) associated with relapse after adjusting for treatment and ancestry and 715 additional SNPs associated with relapse in an ancestry-specific manner. We tested for replication of these relapse-associated SNPs in external data sets of antileukemic drug pharmacokinetics and pharmacodynamics and an independent clinical cohort. 224 SNPs were associated with rapid drug clearance or drug resistance, and 32 were replicated in the independent cohort. The adverse risk associated with black and Hispanic ancestries was attenuated by addition of the 4 SNPs most strongly associated with relapse in these populations (for blacks: model without SNPs hazard ratio (HR)=2.32, P=2.27 × 10-4, model with SNPs HR=1.07, P=0.79; for Hispanics: model without SNPs HR=1.7, P=8.23 × 10-5, model with SNPs HR=1.31, P=0.065). Relapse SNPs associated with asparaginase resistance or allergy were overrepresented among SNPs associated with relapse in the more asparaginase intensive treatment arm (20/54 in Capizzi-methorexate arm vs 8/54 in high-dose methotrexate arm, P=0.015). Inherited genetic variation contributes to race-specific and treatment-specific relapse risk.

Unexpected acute neurologic toxicity in the treatment of children with acute lymphoblastic leukemia.

BACKGROUND: Our current protocol for treatment of childhood acute lymphoblastic leukemia (ALL) was designed to assess the efficacy of methotrexate (MTX) plus L-asparaginase and of etopisode (VP-16) plus cytarabine (ARA-C) during intensive consolidation and continuation therapies and to determine the feasibility of intensifying MTX therapy by the use of divided oral doses of MTX. The protocol was associated with unexpected acute neurotoxicity. There are few reports of such toxic effects during therapy for ALL. PURPOSE: This report describes these toxic effects and outlines our successful approach to the problem. METHODS: The standard four-drug induction regimen consisted of vincristine, L-asparaginase, daunorubicin, and prednisone. In consolidation therapy, oral MTX was given in divided doses (dMTX) of 25 mg/m2 every 6 hours four times daily in four weekly courses concomitant with weekly triple intrathecal therapy--MTX, ARA-C, and hydrocortisone--plus one dose of leucovorin 24 hours after triple intrathecal therapy. Consolidation treatment ended with three daily doses of intravenous VP-16 and ARA-C. The first 16 months of continuation therapy included 6-week cycles of dMTX and L-asparaginase, both given every other week for 5 weeks, with 6-mercaptopurine nightly, and then two doses of VP-16 plus ARA-C and one dose of triple intrathecal therapy. RESULTS: Twenty-five of the 138 patients evaluated had acute neurotoxicity. Ten of the first 72 experienced a seizure or episode of transient neurological deficit 9-11 days following the administration of intravenous ARA-C, VP-16, and triple intrathecal therapy. Despite discontinuation of intrathecal ARA-C, which eliminated simultaneous intravenous and intrathecal treatment with ARA-C, acute neurotoxicity was observed in six previously unaffected patients and six of 42 patients treated after the elimination of intrathecal ARA-C. Therefore, as a second amendment, oral leucovorin was given 24 and 36 hours after dMTX and intrathecal MTX in continuation therapy. No acute neurotoxicity has been seen in 24 patients subsequently entered in the study. CONCLUSION: These findings suggest that folate replacement due to administration of leucovorin modulated MTX toxicity and/or modified an interaction among VP-16, ARA-C, intrathecal therapy, and the central nervous system.

Common region of ALL-1 gene disrupted in epipodophyllotoxin-related secondary acute myeloid leukemia.

Translocations at chromosomal band 11q23 characterize most de novo acute lymphoblastic leukemias (ALL) of infants, acute myeloid leukemias (AML) of infants and young children, and secondary AMLs following epipodophyllotoxin exposure. The chromosomal breakpoints at 11q23 have been cloned from isolated cases of de novo ALL and AML. Using an 859-base pair BamHI fragment of human ALL-1 complementary DNA that recognizes the genomic breakpoint region for de novo ALL and AML, we investigated two cases of secondary AML that followed etoposide-treated primary B-lineage ALL. In the first case, the translocation occurred between chromosomes 9 and 11 and the breakpoint at 11q23 localized to the same 9-kilobase region of the ALL-1 gene that is disrupted in most of the de novo leukemias. In the second case the translocation was between chromosomes 11 and 19. The breakpoint occurred outside of the ALL-1 breakpoint cluster region.

Frequency and diversity of p53 mutations in childhood rhabdomyosarcoma.

The p53 gene was examined in primary or metastatic tumors from six patients with rhabdomyosarcoma (RMS) and in five RMS cell lines by screening methods including single-strand conformation polymorphism analysis, the RNase protection assay, sequencing of complementary DNA subclones, and Southern blotting. Six original tumors were of embryonal histology, four alveolar, and one mixed. p53 mutations were identified in four of the six tumors or cell lines derived from tumors with embryonal histology and in one of the four with alveolar histology. Consistent with p53 allele loss, each mutation was found in the homo- or hemizygous state. One tumor showed a G to C transversion at p53 codon 213 (arginine to proline), and another showed deletion of the entire gene. The p53 mutations in cell lines included a codon 248 C to T transition (arginine to tryptophan) in RD and a codon 280 A to T transversion (arginine to serine) in RH30. The cell line CTR contained a 4-base pair deletion at codons 219/220 in exon 6 with resultant frame shift and premature termination in exon 7. These data support the role of diverse types of p53 mutations in the pathogenesis and/or progression of a significant proportion of cases of childhood RMS.

Fractionated cyclophosphamide and etoposide for children with advanced or refractory solid tumors: a phase II window study.

PURPOSE: Cyclophosphamide (CPA) has a broad spectrum of activity against solid tumors. Hepatic self-induction of the active metabolite 4-hydroxycyclophosphamide occurs after repeated administration. We evaluated the clinical efficacy of a window regimen that administers fractionated CPA in conjunction with etoposide (VP16) in children with advanced or refractory solid tumors. PATIENTS AND METHODS: Seventeen children with advanced (n = 12) or refractory (n = 5) solid tumors were entered onto this phase II window study. The treatment regimen consisted of intravenous (IV) CPA 500 mg/m(2)/d and IV VP16 100 mg/m(2)/d. Both drugs were administered daily by short infusions for 5 consecutive days. RESULTS: A total of 34 courses were administered, with a median of two courses per patient. The median interval between chemotherapy courses was 21 days (range, 17 to 35 days). Thirty-three courses were assessable for toxicity, and all patients were assessable for response. No life-threatening toxicities were observed. The incidence of grade 3 or 4 neutropenia was 94% and of fever and neutropenia 38%. Fever and neutropenia occurred after 12 of 26 courses without recombinant human granulocyte colony-stimulating factor (rhG-CSF) and after one of eight courses with rhG-CSF (P =. 09). Grade 3 or 4 thrombocytopenia occurred after 10 courses (29%). There were no positive blood cultures. One heavily pretreated patient developed a localized perirectal abscess that required drainage. There were 10 patients (59%) with partial responses, four (23.5%) with stable disease, and three with progressive disease. CONCLUSION: Fractionated IV CPA and VP16 over 5 days can be safely administered in children with advanced or refractory solid tumors and has notable antineoplastic activity.

Aerosolized pentamidine for the prevention of Pneumocystis carinii pneumonia in children with cancer intolerant or allergic to trimethoprim/sulfamethoxazole.

PURPOSE: Trimethoprim/sulfamethoxazole (TMP/SMX) is the drug of choice for Pneumocystis carinii pneumonia (PCP) prophylaxis in immunocompromised patients. In children with malignancy, TMP/SMX is well tolerated, but adverse reactions that necessitate discontinuation can occur. We evaluated the safety and efficacy of aerosolized pentamidine (AP) as an alternative prophylaxis modality in children with malignancy who are intolerant of or allergic to TMP/SMX. PATIENTS AND METHODS: AP (200 mg/m2 every 4 weeks) was administered to 60 children with malignancy receiving chemotherapy who had experienced severe adverse reactions to TMP/SMX. Seven hundred twenty doses of AP have been administered during a 3 1/2-year period (21,600 patient-days), with 30 patients treated for > or = 12 months (range, 12 to 25). RESULTS: Adverse reactions occurred during 79 (10%) of the 720 treatments and included bronchospasm in 23, cough in 40, vomiting in 10, and nausea in six. Only two patients had severe bronchospasm. AP was discontinued due to toxicity in three patients (5%). None of the patients (upper 95% confidence limit, 0.049) have developed PCP. CONCLUSION: AP appears to be well tolerated and effective in the prevention of PCP in children with malignancy.

Serum aminotransferase elevation during and following treatment of childhood acute lymphoblastic leukemia.

PURPOSE: The clinical significance of methotrexate (MTX)-induced hepatic toxicity in children with acute lymphoblastic leukemia (ALL) is poorly defined. Therefore, we conducted a study to determine whether intensive MTX therapy could be safely delivered despite isolated serum ALT elevations in children with ALL. PATIENTS AND METHODS: A total of 243 children with B-precursor ALL were treated with extended pulses of oral divided-dose MTX (dMTX). Serum ALT levels were measured approximately every 7 weeks during therapy, as well as after its cessation. By protocol design, treatment was continued without modification in the presence of ALT elevations if there was no other evidence of liver dysfunction. RESULTS: Of 239 assessable patients, 159 (66.5%) had an ALT level > or = 180 IU/L during therapy and 28 patients (17.6%) had one or more values > or = 720 IU/L. After the completion of therapy, only 17 of 104 assessable patients have had one or more elevated ALT value. Eight of these 17 patients (47%) are hepatitis C virus (HCV)-seropositive. The remaining nine children had subsequent normal or near normal ALT values, and none have clinical evidence of liver disease. CONCLUSION: Our data show that MTX can be safely delivered without dose modification in patients with isolated ALT elevations and that continued therapy does not lead to clinically apparent liver disease. ALT elevations are not a reliable predictor of the presence or extent of hepatic injury, and persistently increased ALT values following the completion of ALL therapy are rare in the absence of HCV infection. Continued MTX therapy allows for increased dose-intensity and may improve outcome in children with ALL.

Secondary acute myeloid leukemia in children with acute lymphoblastic leukemia treated with etoposide.

PURPOSE: To describe the occurrence of secondary acute myeloid leukemia (AML) in children with acute lymphoblastic leukemia (ALL) treated with etoposide (VP-16). PATIENTS AND METHODS: Two hundred five consecutive children with early B-lineage ALL were treated according to the Dallas/Fort Worth (DFW) protocol between January 1986 and July 1, 1991. Therapy included a four-drug induction followed by consolidation and continuation phases of nightly oral mercaptopurine (6-MP) and repetitive courses of divided-dose oral methotrexate (dMTX) and asparaginase (L-asp). Three doses of VP-16 and cytarabine (Ara-C) were given during consolidation and later, during continuation, two doses were given 3 to 4 days apart, every 9 weeks. Intrathecal (IT) chemotherapy was given throughout the treatment period. RESULTS: Two hundred three of the 205 patients entered remission. Only eight of these 203 children have had a bone marrow relapse (ALL). However, 10 other children have developed secondary AML 23 to 68 months following the diagnosis of ALL. Overall event-free survival (EFS) at 4 years is 79.3% +/- 5.1%, with a risk of secondary AML at 4 years of 5.9% +/- 3.2%. CONCLUSION: This experience provides strong evidence for a link between epipodophyllotoxin therapy and secondary AML since none of these children received alkylating agent therapy or irradiation. This serious complication raises concern as to the appropriate use of epipodophyllotoxins in the treatment of childhood ALL.

Treatment of CNS relapse in children with acute lymphoblastic leukemia: A Pediatric Oncology Group study.

PURPOSE: To assess the efficacy and toxicity of chemotherapy and cranial radiation for the treatment of children with acute lymphoblastic leukemia (ALL) following first isolated CNS relapse. PATIENTS AND METHODS: One hundred twenty children were treated on Pediatric Oncology Group (POG) protocol 8304. All children had received prophylactic CNS therapy during their initial treatment. The treatment protocol included a four-drug reinduction and six weekly doses of triple intrathecal therapy (TIT). Cranial radiation, 24 Gy, was followed by monthly TIT. Systemic consolidation and maintenance therapy included 6-week cycles of mercaptopurine/methotrexate (6MP/MTX) and vincristine/cyclophosphamide (VCR/CTX), with randomization to intervening pulses of prednisone/doxorubicin (PDN/DOX) or teniposide (VM26)/cytarabine (Ara-C) for a total of 88 weeks. RESULTS: All 120 patients achieved a second complete remission. There have been 61 protocol failures. Thirty-five patients had a bone marrow relapse, four with simultaneous CNS involvement and one with concurrent testicular leukemia. Thirteen patients had a second isolated CNS relapse, 10 a testicular relapse, and two relapsed in other sites. One patient died in remission. Overall event-free survival (EFS) at 4 years was 46% +/- 7%. The toxicity associated with this protocol was minimal except for leukoencephalopathy, which occurred in 20 (17%) patients. The treatment comparison between VM26/Ara-C or PDN/DOX pulses showed a trend toward superior EFS (P = .12) in favor of VM-26/Ara-C. CONCLUSION: To date, this represents the largest series of patients with ALL treated uniformly for an isolated CNS relapse. Since marrow relapse remains the primary site of failure, future protocols must intensify systemic therapy.

Phase I trial of intrathecal liposomal cytarabine in children with neoplastic meningitis.

PURPOSE: We performed a phase I trial of intrathecal (IT) liposomal cytarabine (DepoCyt; Enzon Pharmaceuticals, Piscataway, NJ and SkyePharma Inc, San Diego, CA) to determine the maximum-tolerated dose, the dose-limiting toxicities, and the plasma and CSF pharmacokinetics of IT lipsomal cytarabine in children >/= 3 years of age with advanced meningeal malignancies. PATIENTS AND METHODS: Eighteen assessable patients received IT liposomal cytarabine through either an indwelling ventricular access device or via lumbar puncture. Liposomal cytarabine was given once every 2 weeks during induction, once every 4 weeks during consolidation, and once every 8 weeks during the maintenance phase of treatment. The initial dose was 25 mg, with subsequent escalations to 35 and 50 mg. CSF pharmacokinetic samples were obtained in a subset of patients. RESULTS: Arachnoiditis, characterized by fever, headache, nausea, vomiting, and back pain was noted in the first two patients at the 25 mg dose level. Therefore, subsequent patients were treated with dexamethasone, beginning the day of liposomal cytarabine administration and continuing for 5 days. Headache (grade 3) was dose limiting in two of eight patients enrolled at the 50 mg dose level. Eight of the 14 patients assessable for response demonstrated evidence of benefit manifest as prolonged disease stabilization or response. CONCLUSION: The maximum-tolerated dose and recommended phase II dose of liposomal cytarabine in patients between the ages of 3 and 21 years is 35 mg, administered with dexamethasone (0.15 mg/kg/dose, twice a day for 5 days). A phase II trial of IT liposomal cytarabine in children with CNS leukemia in second or higher relapse is in development.

Glutathione S-transferase genotypes in children who develop treatment-related acute myeloid malignancies.

Epipodophyllotoxin-associated secondary myeloid leukemia is a devastating complication of acute lymphoblastic leukemia (ALL) therapy. The risk factors for treatment-related myeloid leukemia remain incompletely defined. Genetic deficiencies in glutathione S-transferase (GST) activities have been linked to higher frequencies of a number of human malignancies. Our objective was to determine whether the null genotype for GSTM1, GSTT1, or both, was more frequent in children with ALL who developed treatment-related myeloid malignancies as compared to those who did not. A PCR technique was used to assay for the null genotype for GSTM1 and GSTT1 in 302 children with ALL, 57 of whom also subsequently developed treatment-related acute myeloid leukemia or myelodysplastic syndrome. Among children with ALL who did not develop treatment-related myeloid malignancies, the frequencies of GSTM1 and GSTT1 wild-type, GSTM1 null-GSTT1 wild-type, GSTM1 wild-type-GSTT1 null, and GSTM1 and GSTT1 null genotypes were 40%, 42%, 9% and 9%, respectively. The corresponding frequencies for patients who developed acute myeloid malignancies were 42%, 32%, 11% and 16%, respectively (P = 0.26). A statistically significant increase in the frequency of the GST null genotype was observed in male patients who developed myeloid malignancies as compared to male ALL control patients (P = 0.036), but was not observed in female patients (P = 0.51). Moreover, a logistic regression analysis of possible predictors for myeloid malignancies, controlling for gender and race, did not reveal an association of GSTM1 or GSTT1 null genotypes (P = 0.62 and 0.11, respectively) with treatment-related malignancies. Our data suggest that GSTM1 and GSTT1 null genotypes may not predispose to epipodophyllotoxin-associated myeloid malignancies.

A case for the use of aminopterin in treatment of patients with leukemia based on metabolic studies of blasts in vitro.

Clinical and laboratory investigations during the past four decades have resulted in numerous schedules, doses, and routes of delivery for methotrexate (MTX). It remains as an important drug for the treatment of children with acute lymphoblastic leukemia (ALL). Aminopterin (AMT) was the initial antifolate showing promise as an anticancer drug. It is more potent than MTX and also is known to be accumulated more efficiently than MTX in model systems. Because Whitehead et al. (Blood, 76: 44-49, 1990) have shown that MTX accumulation by blasts at diagnosis is of prognostic significance in children with ALL, we reasoned that if accumulation of a "stoichiometric inhibitor" of dihydrofolate reductase by leukemic blasts was of prognostic importance, then whether it was AMT or MTX may be relevant only with respect to the absolute dose. To compare MTX and AMT metabolism, we incubated lymphoblasts with 1 microM radiolabeled drug in vitro. MTX and AMT accumulation by ALL cells (n = 24) was 0.7 +/- 0.7 and 1.47 +/- 0.9 pmol/10(6) cells, respectively. Based on the data of Whitehead et al., this predicts pharmacological success in 59 and 84% of the MTX and AMT groups, respectively. Moreover, 5 of 10 patients considered poor risks based on MTX accumulation would be "cures" based on AMT uptake. Even at only 0.1 microM AMT, a concentration at which there is little accumulation of MTX, 5 of 11 patients studied would be "pharmacological cures" based on AMT uptake. Accumulation of AMT by blasts from 11 patients with T-cell-lineage ALL and 5 patients with acute myelogenous leukemia was also found to be twice the uptake of MTX. These data allow the suggestion that AMT, despite increased potential for toxicity, may be useful in children who are identified as poor risks with respect to MTX uptake.

Methotrexate (MTX) concentration in tumors following low-dose MTX.

Methotrexate (MTX) is a folate analog competitive with reduced folates for cellular transport and metabolism. Since the normal plasma folate concentration is only 10(-8) M, we tested the possibility that there may be a saturable uptake of MTX by proliferating tumor tissue at plasma MTX concentrations of only 10(-7) to 10(-6) M. Patients with advanced malignancies, refractory to accepted therapy, were given low-dose oral MTX (30-60 mg/m2 total dose in four to eight divided doses). Tumor tissue was biopsied 18-24 h after the last oral dose of MTX. The concentrations of MTX and its polyglutamated derivatives were measured in these samples. Forty-eight percent of the drug in the tumor samples was present as a polyglutamated derivative.

Evidence for negative feedback of extracellular methotrexate on blasts of acute lymphoblastic leukemia in vitro.

STUDY OBJECTIVE: To explore the value of high-dose methotrexate (MTX). SUBJECTS: Blast cells from 15 patients with acute lymphoblastic leukemia. INTERVENTIONS: We compared uptake and polyglutamation of [3H]-MTX by freshly isolated leukemic blasts in vitro after 24-hour exposure to 1, 10, and 50 microM [3H]-MTX. MEASUREMENTS AND MAIN RESULTS: Mean MTX uptake (pmol/10(6) cells) was 0.78 +/- 0.19, 2.3 +/- 0.54, and 5.9 +/- 1.9, respectively, and mean polyglutamation was 82%, 66%, and 46%. Consequently, mean MTX polyglutamates were 0.68 +/- 0.18, 1.5 +/- 0.47, and 2.2 +/- 0.67 pmol/10(6) cells. Three of 15 patient samples had no detectable polyglutamation of MTX at 50 microM but MTX polyglutamates were detectable at 1 microM. Two of these three had a decrease in MTX polyglutamates at 10 versus 1 microM. In eight precursor B cell samples there was a significant difference in median MTX polyglutamates at 1 versus 10 microM but not 10 versus 50 microM. CONCLUSION: Increasing extracellular MTX concentrations may be counterproductive for some patients with acute lymphoblastic leukemia. If MTX polyglutamates are important for efficacy, optimal delivery of MTX may have to be determined by individual metabolism rather than by targeting a specific drug concentration.

Factors influencing survival after relapse from acute lymphoblastic leukemia: a Children's Oncology Group study.

Despite great progress in curing childhood acute lymphoblastic leukemia (ALL), survival after relapse remains poor. We analyzed survival after relapse among 9585 pediatric patients enrolled on Children's Oncology Group clinical trials between 1988 and 2002. A total of 1961 patients (20.5%) experienced relapse at any site. The primary end point was survival. Patients were subcategorized by the site of relapse and timing of relapse from initial diagnosis. Time to relapse remains the strongest predictor of survival. Patients experiencing early relapse less than 18 months from initial diagnosis had a particularly poor outcome with a 5-year survival estimate of 21.0+/-1.8%. Standard risk patients who relapsed had improved survival compared with their higher risk counterparts; differences in survival for the two risk groups was most pronounced for patients relapsing after 18 months. Adjusting for both time and relapse site, multivariate analysis showed that age (10+ years) and the presence of central nervous system disease at diagnosis, male gender, and T-cell disease were significant predictors of inferior post-relapse survival. It can be noted that there was no difference in survival rates for relapsed patients in earlier vs later era trials. New therapeutic strategies are urgently needed for children with relapsed ALL and efforts should focus on discovering the biological pathways that mediate drug resistance.

"Above all do no harm:" horizons in pediatric oncology.

The articles selected in this review highlight some advances in the use of the "old standards" methotrexate and 6-mercaptopurine for the treatment of children with acute lymphoblastic leukemia, especially the laboratory studies, which may allow a "pharmacologic phenotyping" to identify children most likely to relapse. In addition, we review the use of "newer" drugs (epipodophyllotoxins), which may be effective but are also associated with high morbidity (ie, secondary acute myeloid leukemia). As a second issue, "dose intensity" and the use of high-dose chemotherapy followed by bone marrow transplantation will be challenged, especially in light of peripheral stem cell harvest and the use of growth factors.

Epstein-Barr virus lymphoproliferative disorder in children with leukemia: case report and review of the literature.

PURPOSE: The purpose of this study is to report a case of Epstein-Barr virus-related lymphoproliferative disorder (EBV-related LPD) in a child with leukemia and present a review of literature on the subject. PATIENTS AND METHODS: A 6-year-old boy with acute lymphoblastic leukemia (ALL) undergoing maintenance chemotherapy presented with fever, abdominal pain, and vomiting. Imaging studies revealed multiple mass lesions in the liver, kidneys, and lungs. Liver biopsy was obtained. Immunocytochemistry for T and B lymphocytes and in situ hybridization for evidence of latent EBV infection was performed. RESULTS: Hepatic portal tracts were infiltrated with lymphocytes, monocytes, eosinophils, and large atypical mononuclear cells. Both T and B lymphocytes were present, but the large atypical cells were of B origin and were positive for latent EBV infection. Chemotherapy was discontinued, and the patient was treated with intravenous gammaglobulin, interferon, and acyclovir. He had an excellent response and has been free of disease for 19 months. CONCLUSION: EBV-related LPD can complicate the course of patients with ALL in remission. The clinical findings and diagnosis in patients with ALL are similar to other immunocompromised patients. Withdrawal of chemotherapy and treatment with immune modulators should be strongly considered.

Adjuvant chemotherapy for treatment of unresectable and metastatic angiomatoid malignant fibrous histiocytoma.

Angiomatoid malignant fibrous histiocytoma (AMFH) is a low grade soft tissue sarcoma usually treated with surgery alone. Only one adult patient has been treated with systemic chemotherapy. The authors report a case of unresectable, metastatic AMFH treated initially with vincristine, doxorubicin, dactinomycin, and cyclophosphamide. A complete response at the metastatic site and a marked reduction in the size of the primary tumor allowed complete surgical excision 7 months after treatment was initiated. The patient remains disease free 19 months after being diagnosed. It was concluded that systemic chemotherapy may be effective in patients with AMFH.