ALBERT

Description: Background. Asn depletion in the serum during ASNase treatment (Tx) causes the death of lymphoblasts that lack ability to synthesize Asn. The Asn serum levels during treatment depend on ASNase activity, the rate of clearance of the enzyme and the input rate of Asn into circulation from the liver and other organs (Imax). We developed a population pharmacokinetic & pharmacodynamic (PK-PD) model in NONMEM to estimate the input rate in the standard risk ALL patients (pts) treated on CCG-1962. Methods. We compared the Asn concentrations during induction determined by HPLC amino acid assay in 24 pts who subsequently relapsed and in all event-free (CCR) pts from CCG-1962 SR ALL study. A best fit population PK-PD model between ASNase and Asn concentrations was developed for each patient to estimate the maximum input rate, Imax, and its additive error correction factor. Six year event-free survival was correlated with Imax or additive error with Kaplan-Meier analyses. Results. Patients received either 2500 IU/m2 PEG-ASNase once or 6000 IU/m2 native ASNase x 9, intramuscularly, during induction. The deaminated Asn serum levels were superimposable in patients randomized to native or PEG-ASNase arm. The mean and SDEV for Imax and the additive error are shown in Table 1 for each enzyme preparation for patients in CCR and for those who later developed BM, CNS, or testicular relapses. Imax was significantly greater in the relapsed than in the CCR patients, in the PEG-ASNase (p=0.002) and in the native ASNase (p=0.005) arms. Table 1. Imax and additive error of Asn in relapsed and in continuous complete remission (CCR) patients - CCG-1962 CCG-1962 Arms PEG-ASNase arm, n=56 PEG-ASNase arm, n=56 Native ASNase arm, n=57 Native ASNase arm, n=57 * nmoles/ml/min, ** All values are mean ± SDEV PD Parameters Imax* Additive error* Imax* Additive error* CCR patients 9.93 E -6 ± 2.4 E -5** 1.51 E -6 ± 2.86 E -6 2.92 E -5 ± 1.52 E -4 2.52 E -6 ± 6.61 E -6 N 46 46 43 43 Relapsed 8.9 E -3 ± 6.0 E -3 2.87 E -6 ± 3.1 E -6 4.51 E -3 ± 4.94 E -3 1.05 E -5 ± 1.76 E -5 N 10 10 14 14 p, unequal variance t-test P=0.0021 p=0.25 p=0.0047 p=0.118 Table 2 shows the summary of the Kaplan-Meier analyses in all patients. Both the Imax and additive factor significantly correlate with relapse. Table 2. Summary of Kaplan - Meier analyses Patients PEG-ASNase arm, n=56 PEG-ASNase arm, n=56 Native E.Coli-ASNase arm, n=57 Native E.Coli-ASNase arm, n=57 Parameter and cutoff Imax 〈 0.001 Imax 〉 0.001 Imax 〈 0.001 Imax 〉 0.001 Events, Observed/Exp 2/8.59 8/1.41 3/6.14 11/7.86 Total 45 11 26 31 Log-rank p value 1.64 E -6 0.046 Parameter and cutoff Add rate 〈 1E -6 Add rate 〉 1E -6 Add rate 〈 1E -6 Add rate 〉 1E -6 Events, Observed/Exp 1/5.63 9/4.37 5/10.36 9/3.64 Total 28 28 39 18 Log-rank P value 0.002 0.0024 Conclusions. The Asn levels after Tx are similar in both arms in pts with events and CCR. Nevertheless this PD model strongly suggests that the Imax of input of Asn in the circulation is faster in relapsed than in CCR patients. Furthermore, Imax and its additive rate are correlated with 6 year event free survival of SR ALL pts, independent of the ASNase formulation used in their Tx.

Description: Background: Residual disease or rapidity of response to induction therapy is among the most powerful predictors of outcome in pediatric Acute Lymphoblastic Leukemia (ALL). Various methods to determine response during induction have been in use in clinical investigation. We hypothesize that drug sensitivity at the cellular level predicts rapidity of response to induction therapy in ALL. We recently developed a high resolution flow cytometry based cytotoxicity assay for in vitro cellular drug response profiling for pediatric ALL. This method has a turnaround time of 48 hours and the ability to measure drug effect specific to leukemic cells regardless of number of admixed normal cells. We report preliminary data that correlate results of this drug sensitivity assay with rapidity of response to induction therapy among patients with ALL. Methods: We performed in vitro tests, applying a multiparameter flow cytometric drug cytotoxicity assay, on bone marrow (BM) samples of 23 patients with newly diagnosed standard (n = 10), high (n = 11), and very high (n = 2) risk ALL, as defined by the Children’s Oncology Group (COG) and NCI risk classification. Fourteen patients were rapid early responders (RER) and 9 were slow early responders (SER) by COG criteria at day 15 and 29. Cryopreserved cells from BM samples were thawed and determined to have adequate viability by trypan blue dye exclusion. Drugs were tested at three different concentrations, each in triplicate. Concentrations tested were based on an empirically derived cut-off concentration (EDCC) adopted from published in vitro studies, chosen to produce a large scatter of survival index values among samples. Leukemic blasts were specifically identified by surface markers, CD 45, CD 19 and CD 10 or CD 3, while cytotoxicity was measured with Annexin V based apoptosis. Leukemic cell survival index (LCSI = Average Replicate /Average Control x 100) was determined at 48 hours after in vitro exposure to individual standard induction agents for pediatric ALL: vincristine, asparaginase, dexamethasone, prednisone and daunomycin. LCSI differences between RER and SER were compared using Wilcoxon rank sum test for each drug and concentration. The mixed effects model was used to evaluate the overall difference of LCSI between RER and SER over the three concentrations (referred to as “averaged concentrations”). Results: For dexamethasone, a significantly lower LCSI was seen in the RER compared with the SER cohort at individual and averaged concentrations: RER mean LCSI = 40.2%, SER mean LCSI = 70.1% (p = 0.01, mixed effects model). A trend toward a lower mean LCSI in the RER compared with the SER group was noted for asparaginase and vincristine at individual and averaged concentrations (p 〈 0.1). Mean LCSI was not different between the RER and SER groups for daunomycin and prednisone at individual or averaged concentrations. Conclusions: This in vitro drug sensitivity assay provides a response profile for dexamethasone that correlates with in vivo response to induction therapy. Research is ongoing with more patient samples in order to achieve a greater statistical power to detect a smaller difference for all drugs tested. Further research will also correlate clinical response with LCSI using drug combinations in vitro. Results of these studies will determine the potential value of this assay for early risk stratification and modification of therapy in de novo or relapsed ALL.

Description: Introduction: Overall, children with new-onset SR (age 1-9 years (yr), WBC

Description: The event-free survival (EFS) of children with SR-ALL is now 〉 80%. Nonetheless, ALL relapse remains a major problem in pediatric cancer, and optimal therapy for these children remains to be defined. We examined post relapse outcomes for children initially treated on the Children’s Cancer Group CCG-1952 study and compared stem cell transplant (SCT) with chemotherapy (CT) as salvage treatment in second remission (CR2). Between 5-96 and 1-00, 2176 eligible patients with SR-ALL (WBC 〈 50,000/mcl; age ≥ 1, 〈 10 years) were enrolled on CCG-1952, 321 of whom experienced a relapse after initially achieving M1 remission. Among the relapses, 196 are bone marrow (BM) ± extramedullary (EM) relapses (61%) and 125 are isolated EM (iEM: CNS, testicular, or ocular) (39%) relapses. The mean time (range) from first remission to BM relapse, iCNS relapse, and itesticular relapse is 34 ± 1 (2 to 79) month (mo.), 22 ± 1 (1 to 70) mo., and 37 ± 3 (4 to 74) mo., respectively. The 3-year EFS and overall survival (OS) after relapse for all patients are 41% and 53%. Three-year EFS and OS after BM ± EM relapse are 32% and 40% and after iEM relapse are 55% and 71%. Patients with early BM relapse (CR1 〈 36 months) or early iEM relapse (CR1 〈 18 months) are 2.3 times (p=0.002) and 2.8 times (p=0.01) more likely to suffer a subsequent adverse event than patients with later relapse by Cox regression analysis after adjustment for age, pre-relapse treatment, day 7/14 BM status, and type of treatment in CR2. The 3-year OS for early and late relapses are 31% vs 59% (p=0.001) for BM relapse and 50% vs 87% (p=0.01) for iEM relapse. We compared 73 SCT and 215 CT patients, excluding 33 patients with adverse events prior to the median time to SCT (130 days, range 56 to 1148 days) using Kaplan-Meier life table analysis and the log rank test. The cohorts are similar with respect to age (p=0.57), duration of CR1 (p=0.96), and initial Day 14 BM status (p=0.69). Thirty-seven percent (n=62) of patients with BM relapse (33 early; 29 late) and 9% (n=11) of patients with iEM relapse (6 early, 5 late) received SCT. Thus far, outcomes are similar between cohorts for patients with any BM relapse (OS, p=0.45; EFS, p=0.70), early BM relapse (OS, p=0.95; EFS, p=0.66), and iEM relapse (OS, p=0.44; EFS, p=0.96). Among patients with later BM relapse, a trend favors CT (OS, p=0.08; EFS, p= 0.14). In summary, duration of CR1 remains the most significant predictor of outcome after either BM or iEM relapse in this SR-ALL cohort. Prognosis after early BM relapse remains poor with either SCT or CT.

Description: Abstract 2634 Poster Board II-610 Background: Philadelphia chromosome positive Acute Lymphoblastic Leukemia (Ph+ALL) occurs in 2–5% of pediatric ALL and is associated with a poor prognosis. COG AALL0031 treated children with an intensified chemotherapy backbone plus imatinib. All subjects received imatinib at 340mg/m^2 daily. Exposure to imatinib progressively increased in each of five cohorts. Patients had a total imatinib exposure (before maintenance) of 42 days in cohort 1, 63 days in cohort 2, 84 days in cohort 3, 126 days in cohort 4, and 280 days in cohort 5. All groups received an additional 336 days of imatinib exposure in maintenance cycles 1 through 12 for approximately 2 years (with imatinb given on 21 day cycles for maintenance cycles 1 – 4, and a two-week on/two-week off schedule for maintenance cycles 5 - 12). Early results of this trial show encouraging outcome with a 3-year event free survival of 80±11% (95% CI 64 – 90%) for patients in cohort 5. In studies of adults with Ph+ALL treated with imatinib many patients recurred with imatinib resistant BCR-Abl mutations. To date, there are no data on the occurrence of BCR-Abl mutations in pediatric Ph+ALL. Patients and Methods: We performed nested PCR to identify BCR-Abl point mutations in nine samples obtained at bone marrow (BM) relapse from Ph+ALL subjects on AALL0031. Results: (Table 1) Three samples from cohort 1 that had no exposure to imatinib prior to relapse showed wild-type sequence. There were 5 of 6 samples that also showed wild-type sequence. One sample was from cohort 2 and 3 samples were from cohort 3. Each subject relapsed 1 to 2 years after diagnosis while receiving varying amounts of imatinib with continued intensive therapy. One subject recurred after stem cell transplant in first remission. One sample from cohort 4 recurred after the completion of chemotherapy and imatinib. One subject from cohort 5 carried the histidine 396 to proline (H396P) mutation. This mutation, which increases the imatinib IC50 by 10-fold, has been previously described to occur in adults with CML and Ph+ALL treated with imatinib. The subject from cohort 5 recurred 1 year after diagnosis on therapy with imatinib. Conclusions: Only 1 resistant mutation in BCR-Abl has been identified among nine children with Ph+ALL treated on AALL0031. Therefore, unlike results in the adults, resistant mutations do not appear to drive early recurrence in Ph+ALL. Further studies will be needed to identify whether BCR-Abl mutations are identified in subjects who develop a late relapse after treatment with AALL0031 or subsequent treatment studies. Disclosures: Druker: OHSU patent #843 - Mutate ABL Kinase Domains: Patents & Royalties; MolecularMD: Equity Ownership; Roche: Consultancy; Cylene Pharmaceuticals: Consultancy; Calistoga Pharmaceuticals: Consultancy; Avalon Pharmaceuticals: Consultancy; Ambit Biosciences: Consultancy; Millipore via Dana-Farber Cancer Institute: Patents & Royalties; Novartis, ARIAD, Bristol-Myers Squibb: Research Funding. Schultz:novartis: Membership on an entity's Board of Directors or advisory committees.

Description: Children's Cancer Group-1991 selected 2 components from the Children's Cancer Group studies shown to be effective in high-risk acute lymphoblastic leukemia and examined them in children with National Cancer Institute standard-risk acute B-precursor lymphoblastic leukemia. These were (1) vincristine and escalating IV methotrexate (MTX) without leucovorin rescue during the interim maintenance (IM) phases and (2) addition of a second delayed intensification (DI) phase. Eligible patients (n = 2078) were randomly assigned to regimens containing either oral (PO) MTX, PO mercaptopurine, dexamethasone, and vincristine or IV MTX during IM phases, and regimens with either single DI or double DI. Five-year event-free survival (EFS) and overall survival for patients on the PO MTX arms were 88.7% ± 1.4% and 96% ± 0.9% versus 92.6% ± 1.2% and 96.5% ± 0.8% for those on the IV MTX arms (P = .009, P = .66). Five-year EFS and overall survival for patients who received single DI were 90.9% ± 1.3% and 97.1% ± 0.8% versus 90.5% ± 1.3% and 95.4% ± 3.8% for those who received double DI (P = .71, P = .12). No advantage was found for a second DI; however, replacement of PO MTX, PO mercaptopurine, vincristine, and dexamethasone during IM with vincristine and escalating IV MTX improved EFS.

Description: The Children's Cancer Group 1952 (CCG-1952) clinical trial studied the substitution of oral 6-thioguanine (TG) for 6-mercaptopurine (MP) and triple intrathecal therapy (ITT) for intrathecal methotrexate (IT-MTX) in the treatment of standard-risk acute lymphoblastic leukemia. After remission induction, 2027 patients were randomized to receive MP (n = 1010) or TG (n = 1017) and IT-MTX (n = 1018) or ITT (n = 1009). The results of the thiopurine comparison are as follows. The estimated 7-year event-free survival (EFS) for subjects randomized to TG was 84.1% (± 1.8%) and to MP was 79.0% (± 2.1%; P = .004 log rank), although overall survival was 91.9% (± 1.4%) and 91.2% (± 1.5%), respectively (P = .6 log rank). The TG starting dose was reduced from 60 to 50 mg/m2 per day after recognition of hepatic veno-occlusive disease (VOD). A total of 257 patients on TG (25%) developed VOD or disproportionate thrombocytopenia and switched to MP. Once portal hypertension occurred, all subjects on TG were changed to MP. The benefit of randomization to TG over MP, as measured by EFS, was evident primarily in boys who began TG at 60 mg/m2 (relative hazard rate [RHR] 0.65, P = .002). The toxicities of TG preclude its protracted use as given in this study. This study is registered at http://clinicaltrials.gov as NCT00002744.

Description: Children with T-ALL have generally had a poorer prognosis than patients with precursor-B ALL. Patients with T-ALL are more likely than those with B-precursor ALL to be 〉 9 years and present with WBC 〉 50,000/ul, bulky lymphadenopathy and mediastinal mass. Since 1996, the former CCG has stratified protocol eligibility for patients with ALL based on NCI defined Standard Risk (SR = age between 1-9.99 years; WBC 〈 50,000/ul) or High Risk (HR) groups, regardless of immunophenotype. In contrast, the former POG has treated all T-cell patients on protocols separate from those for precursor-B ALL. This report compares the outcomes among children with T-cell ALL treated on recently completed or ongoing CCG and POG phase III trials for ALL. CCG-1952 enrolled 2176 eligible children with SR ALL between 1996 and 2000; 106 (5%) had T-cell immunophenotype. Treatment was a standard BFM regimen with prednisone and 2 phases of delayed intensification (DI) with a 2x2 randomization of IT methotrexate (MTX) v ITT and MP v TG. From June 2000 to March 2004, CCG-1991 has enrolled 1794 SR ALL patients: 80 (4%) had T-ALL. Treatment included a similar BFM backbone with substitution of dexamethasone and a 2x2 randomization between escalating IV v oral MTX and 1 v 2 DIs. POG-9404 (opened 1996; closed 2001) was developed exclusively for T-cell disease and enrolled 363 patients with T-ALL; 84 (23%) fit the NCI SR group criteria. On the latter protocol, patients received treatment similar to that developed by the Dana-Farber Leukemia Consortium for high risk B-precursor ALL, with randomization to ± 4 cycles of high dose MTX (5 Gm/m2) and leucovorin. All patients on 9404 received 1800 cGy prophylactic cranial radiation while CCG patients did not. Outcome for T-ALL on CCG-1952 is substantially worse than for B-precursor ALL, with 5 year event-free survival (EFS) of 73% compared to 82% (p = 0.007). Interim analysis of CCG-1991 also shows a significantly worse outcome for T-ALL compared to B-precursor ALL: 3y estimated EFS 78% v 90%, p = 0.0002. In contrast, estimated 5y EFS for patients with SR T-ALL on POG-9404 is 88% (90% on the superior high dose MTX regimen). Comparison of the SR v HR T-ALL patients treated on POG 9404 shows a significant advantage for the SR group (5y EFS of 90% v 75%, p 〈 0.004). This is in contrast to comparison of T-ALL patients on CCG-1952 (SR) v the concurrent CCG-1961 HR study where T-ALL patients have similar outcome (5y EFS 73% v 72%, p = 0.77). These data suggest that patients with T-ALL and SR features have better EFS when treated with more intensified chemotherapy regimens. Because early EFS for T-cell patients treated on CCG-1991 is worse than on POG 9404, the former study was closed to further accrual of patients with T-ALL. The COG ALL Committee is developing a study for exclusive enrollment of patients with T-ALL using intensive therapy based on the current COG HR ALL regimen, regardless of SR or HR features.

Description: The EFS/OS for SR (age 1-9.99 yrs and initial white blood cell count

Description: ALPS is a heritable disorder of apoptosis resulting in accumulation of autoreactive lymphocytes, leading to childhood onset of marked lymphadenopathy, hepatosplenomegaly with multilineage cytopenias due to splenic sequestration and/or autoimmunity. Affected individuals have characteristic increases (≥1%) in circulating (TCRαβ+ CD4-CD8-) double negative T lymphocytes. ALPS Type Ia, Ib, II and IV are associated with genetic mutations in Fas, FasL, Caspases and NRAS respectively, while patients without identified mutations are classified as Type III or ALPS phenotype. Of the estimated 350 ALPS patients reported worldwide, 241 individuals with ALPS, belonging to 166 families are enrolled in studies at the NIH Clinical Center. Nearly 50% of them with refractory cytopenias associated with hypersplenism have undergone splenectomy leading to associated long term risk of pneumococcal sepsis. Currently published reports include occasional cases of rituximab used for refractory ITP and AIHA with variable response in patients classified as ALPS. We summarize experience of rituximab use during the last 7yrs in 11 ALPS patients (8 males and 3 females), including 2 adults aged 45, 46yrs and 9 children with a median age of 11yrs (range 1–14yrs) at commencement of this therapy for refractory grade 4 immune thrombocytopenia (platelet count 12 months follow up; 2 of them (aged 14 and 15yrs) relapsed at +18 and +22 months after completion of therapy: only one of them responded to a second course of rituximab. Thrombocytopenia in the second patient is currently under control with MMF. One out of the 4 non-responders underwent splenectomy, while the other 3 patients, including the 2 asplenics, required prednisone pulses (5–30 mg/kg) followed by MMF and weekly vincristine for 6 wks (in 1 patient) as a steroid sparing measure. Noted toxicities include prolonged neutropenia in 1 patient, profound and prolonged hypogammaglobulinemia in 4 patients requiring replacement IVIG, and total absence of antibody response to polysaccharide vaccines lasting up to 4years after rituximab infusions in 1 patient. Rituximab therapy relieved thrombocytopenia for 12–18 months in 7 out of 11 ALPS patients. However associated toxicities, including quantitative and qualitative IgG deficiencies that are additional infection risk burden, especially in asplenic individuals, warrant further investigation and long term follow up of ALPS patients with cytopenias.