Standard-risk Acute Myeloid Leukemia Research Articles

Umbrella Trial in Myeloid Malignancies: The Myelomatch National Clinical Trials Network Precision Medicine Initiative

Introduction: To accelerate myeloid cancer therapeutics, the National Clinical Trials Network (NCTN) is launching the National Cancer Institute (NCI) Myeloid Malignancies Molecular Analysis for Therapy Choice (myeloMATCH) precision medicine clinical trial. Sponsored by NCI and conducted across the entire NCTN system with initial trials to be led by SWOG, Alliance, ECOG-ACRIN, and CCTG, the initiative leverages public-private partnerships with pharmaceutical industry and biotech companies to create an efficient regulatory model incorporating cross-company novel-novel combinations in trials for acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS). The goal is to create a portfolio of rationally designed treatment substudies onto which patients sequentially enroll over their treatment journey. As patients transition to higher tiers with increasingly lower remaining tumor burden, the focus will be to target residual disease more precisely. Methods: Newly diagnosed patients are enrolled onto the Master Screening and Reassessment Protocol (MSRP) for baseline clinical and laboratory evaluation. Specimens are sent to the Molecular Diagnostics Network (MDNet) with a 72-hour turn-around for patient assignment to an initial treatment substudy via an integrated informatics system developed by the NCI Precision Medicine Analysis and Coordination Center (PMACC). Assignments will be based on algorithms adjusted for prevalence of co-mutations to enhance accrual of rare molecular subsets to specific targeted treatment trials. As shown in figure 1, there are 4 tiers and 5 clinical baskets. Tier-1 is for initial therapy grouped by MDS, younger AML, and older AML. These are typically randomized phase 2 studies testing sensitivity to novel drug combinations with measurable residual disease (MRD) assessment conducted centrally by MDNet. Subsequent therapy occurs in higher tiers. These assignments are made by MDNet/PMACC based on prior treatment substudy outcome. Flow cytometry and duplex sequencing will be employed in Tier-4 clinical trials that will target residual disease. Statistical designs will evaluate the clinical utility of the assays and biomarkers to determine if targeting residual disease confers clinical benefit. Planned activation is quarter 4 of 2022 with the MSPRP, 2-young adult tier-1 studies and 1 tier-2 study. These are testing combinations of azacitidine, venetoclax, CPX351, 7+3 for ELN defined high risk AML, standard risk AML, and in tier-2 the ability to "erase” residual disease after tier-1 treatment. Additional studies in development include agents for mutant TP53, KIT, FLT-3, NPM1, IDH 1/2, higher and lower-risk MDS and a study for reduced intensity transplant and maintenance to include efforts for diverse populations. Launch for these studies is planned for mid 2023. Tier-4 studies to target KIT, IDH, FLT3 and others are in discussion. Discussion: MyeloMATCH is the largest focused investment of infrastructure and researchers ever coordinated by the US Network Groups and NCI that follows patients from diagnosis thorough all treatment in a single disease area. The charge is to rapidly advance therapeutics in myeloid malignancies. MyeloMATCH is designed to efficiently screen and assign patients to precision treatment trials of promising therapeutic combinations. By using early endpoints to identify large activity signals, myeloMATCH will generate data with promising findings for definitive study. The clinical and laboratory data can be interrogated across the initiative to generate hypotheses for additional focused testing. Participants receiving their treatment journey through myeloMATCH will contribute to a unique clinically annotated database with specimens for "omics” serially collected from pre-treatment and through follow up. This will provide a national resource for understanding drug sensitivity and resistance, as well as clonal evolution. In this manner, myeloMATCH aims to reduce the time and investment in failed phase 3 studies, and instead aims to provide high-quality randomized trial data that will enhance selection of phase 3 priorities. We believe this new paradigm for the collaborative conduct of clinical trials may provide meaningful advances for patients with AML and MDS, mentor investigators, and accelerate drug development. Updates and specific treatment-trial designs will be discussed at the meeting. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Stratification of Intermediate-Risk Acute Myeloid Leukemia According to the Expression Level of WT1 mRNA at Diagnosis

IntroductionDespite advances in the genetic analysis of acute myeloid leukemia (AML), Wilms' tumor oncogene 1 (WT1) mRNA remains an important pan-marker of AML. A log reduction in WT1 mRNA expression after chemotherapy is a predictor of prognosis, and WT1 mRNA can be used as a marker of minimal residual disease or relapse. In the 1990s, a high expression level of WT1 mRNA at diagnosis was considered a poor prognostic factor. However, recent analyses have found that WT1 mRNA expression varies with AML type. Since the prognosis is affected by cytogenetic abnormalities and therapeutic intensity, we re-evaluated the clinical significance of WT1 mRNA expression at diagnosis in the context of the European Leukemia Net (ELN) risk classification and treatment.MethodWe retrospectively analyzed 216 patients at five institutions between 2011 and 2020. The expression level of WT1 mRNA was measured for all patients at diagnosis using bone marrow (BM) samples from 191 patients and peripheral blood (PB) samples from 25. WT-1 mRNA expression was measured using real-time quantitative polymerase chain reaction and the measured values were converted in normal logarithm. The median age at diagnosis was 62 (range: 23-93) years. The cytogenetic risk of ELN was classified as favorable (n = 41), intermediate (n = 123), and adverse risk (n = 41). Selected therapeutics were standard chemotherapy (n = 182, 84.3%, including CAG regimen; cytarabine, aclarubicin and G-CSF), hypomethylating agents or low-dose cytarabine (n = 19, 8.8%), and best supportive care (n = 15, 6.9%). Also, 143 patients (66.2%) received one or more courses of standard consolidation chemotherapy and 61 (28.3%) underwent allogeneic hematopoietic stem cell transplantation (allo-HCT). The median observation period was 518 [1-3418] days (Table 1). The overall survival (OS) and event-free survival (EFS) rates were assessed. Relapse or death was defined as an event, and OS was evaluated on the date of death.ResultThe median expression level of WT1 mRNA was 4.68 (range: 1.0-5.72) [log copies/µg RNA, units are omitted thereafter] in BM and 3.66 (range: 1.34-5.20) in PB (Table 1). Favorable-risk AML had the highest WT1 mRNA expression level in BM (4.95, p = 0.0054), whereas there was no difference between the expression levels in intermediate- and adverse-risk AML in BM (4.63 and 4.47, p = 0.711, Fig. 1A). WT1 mRNA expression in PB were 4.04, 3.84, and 3.05 for favorable-, intermediate-, and adverse-risk AML, respectively, and were higher for those with a favorable-risk AML (vs. adverse risk, p = 0.048, Fig. 1B). When WT1 mRNA expression level in BM was compared between the two groups, i.e., <4.0 (BM-WT1 low) vs. ≥4.0 (BM-WT1 high), 2-year EFS rates were 26.8% and 49.7% (p = 0.0035) and 2-year OS rates were 44.9% and 61.5% (p = 0.00053), respectively. When WT1 mRNA expression level in PB was compared between the two groups, i.e., <3.0 (PB-WT1 low) vs. ≥3.0 (PB-WT1 high), 2-year EFS rates were 0% and 51.5% (p = 0.023) and 2-year OS rates were 0% and 73.1% (p = 0.01), respectively. PB- or BM-WT1 low showed a worse prognosis at diagnosis.Since AML prognosis is affected by ELN risk and selected therapeutics, we evaluated 109 intermediate-risk patients who had received at least one course of standard chemotherapy. Fifty of them (45.9%) underwent subsequent allo-HCT. The 2-year EFS rates of PB or BM-WT1 low (n = 25, PB = 2 and BM = 23) vs. PB or BM-WT1 high (n = 84, PB = 23 and BM = 61) were 23.9% and 50.4%, respectively (p = 0.023, Fig. 2A) and the 2-year OS rates were 51.2% and 64.6%, respectively (p = 0.03, Fig. 2B). PB or BM-WT1 low of intermediate-risk AML had a poor prognosis even with standard chemotherapy. Multivariate analysis adjusted for ages, the reduction rate of WT1 mRNA after induction chemotherapy, duration of receiving consolidation chemotherapy, allo-HCT, and PB or BM-WT1 low at diagnosis were independent poor prognosis factors for EFS in intermediate-risk AML (HR: 3.32, 95%CI: 1.29-8.53, p = 0.013). The OS rate of PB or BM-WT1 low also worsened (HR: 2.33, 95%CI: 0.88-6.18, p = 0.089) (Table 2). Whereas, the outcome of adverse-risk AML was not affected by PB or BM-WT1 low/high.ConclusionThe expression level of WT1 mRNA at diagnosis was negatively correlated with prognosis. Favorable-risk AML had higher expressions of WT1 mRNA. PB or BM-WT1 low in intermediate-risk AML was associated with a poor prognosis. In standard-risk AML, WT1 mRNA may be a useful pan-marker to predict prognosis at diagnosis. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Replacing the Anthracycline By Gemtuzumab Ozogamicin in Older Patients with De Novo Standard-Risk Acute Myeloid Leukemia Treated Intensively - Results of the Randomized ALFA1401-Mylofrance 4 Study

▪IntroductionBased on the results of the ALFA-0701 trial (Castaigne et al. Lancet 2012), the addition of fractionated doses of the anti-CD33 antibody drug conjugate gemtuzumab ozogamicin (GO, Mylotarg®) to conventional chemotherapy has been approved in 2017 for frontline treatment of adults with CD33-positive acute myeloid leukemia (AML). Prolonged event-free survival (EFS) was observed in patients with AML of favorable or intermediate risk, while not in those with adverse cytogenetics. Nevertheless, more frequent added toxicities could make the addition of GO a questionable option in patients over 60-65 years of age. In this ALFA-1401/Mylofrance 4 trial (NCT02473146), we investigated if the replacement of the anthracycline by GO might also improve EFS in older patients.MethodsBetween January 2016 and March 2019, 225 patients were randomized 2:1 to receive an experimental GO-cytarabine combination (154 patients) or a standard anthracycline-cytarabine treatment (71 patients). Patients aged 65 to 80 years old (later extended to patients aged 60-64 years old), with previously untreated de novo AML of favorable or intermediate cytogenetics were eligible for the trial. Standard treatment arm consisted in a 7+3 using idarubicin at 12 mg/m 2/d on day 1 to 3 and cytarabine 200 mg/m 2/d on day 1 to 7. Experimental arm (GO arm) consisted of two doses of GO 3 mg/m 2/d on day 1 and 4 and cytarabine 200 mg/m 2/d on day 1 to 7. Post-remission therapy comprised two courses of intermediate-dose cytarabine (IDAC) at 1.5 g/m 2/12h on day 1, 3 and 5. In the GO arm, a third dose of 3mg/m 2/d GO was administered on day 1 of the first IDAC course. The first IDAC course could serve as second induction in patients not responding to the first one. The decision to perform allogeneic hematopoietic stem cell transplantation (allo-HSCT) in first remission was left to the discretion of the physician. The efficacy analysis was conducted in the modified intent-to-treat (mITT) population excluding patients who did not meet the cytogenetic eligibility criteria. The primary study endpoint was EFS. Secondary endpoints were response rate defined by complete remission (CR), CR with incomplete platelets recovery (CRp) and CR with incomplete hematological recovery (CRi), early mortality, relapse incidence, overall survival (OS) and safety.ResultsAmong the 225 randomized patients, 214 (71 standard arm, 143 GO arm) were included in the mITT population. There were 126 men and 88 women. Median age was 70 years (61-80). Cytogenetics was of favorable and intermediate risk in 14 and 200 patients, respectively. One hundred and eighty-one patients reached CR/CRp/CRi, 64 (90%) in standard arm and 117 (82%) in GO arm (p=0.17). Median follow-up time was 38 months. At 2 years, estimated EFS was 38% [95% CI, 28-51] in the standard arm vs. 29% [22-37] in the GO arm (Hazard Ratio (HR), 1.37 [0.98-1.93]; p= 0.067) (Figure 1A). Overall, 118 patients relapsed, 36 (51%) in standard arm and 82 (57%) in GO arm. At 2 years, estimated cumulative incidence of relapse was 48% [36-61] in standard arm vs. 61% [52-70] in GO arm (csHR, 1.32 [0.90-1.93]; p= 0.078). Overall, 122 patients died, 38 (54%) in standard arm, 84 (59%) in GO arm. Sixty-day mortality was 4% in standard arm vs. 10% in GO arm (p=0.13). At 2 years, estimated OS was 65% [55-77] in standard arm vs. 52% [45-61] in GO arm (HR, 1.27 [0.86-1.87]; p= 0.23). In subgroup analysis for EFS (Figure 1B), we found a significant interaction with gender, GO having a detrimental effect in women which persisted after adjustment on known prognostic factors. A total of 33 patients received allo-HSCT in first remission, 19 (30%) in standard arm and 14 (12%) in GO arm (p=0.006). When censoring these patients at transplant time, HR of GO on EFS was 1.27 ([0.88-1.83]; p=0.19). Regarding safety, 76% and 80% of patients had at least one grade 3 to 5 adverse event (p=0.81), including infection in 30% vs. 21% and bleeding in 7% vs. 29% in standard arm and GO arm respectively. Serious adverse events were reported in 34% of patients in standard arm vs. 49% in GO arm (p=0.031). Sinusoidal obstruction syndrome occurred in 2 patients in the GO arm.ConclusionFrontline use of GO instead of idarubicin, when combined to cytarabine, does not benefit older patients with de novo standard-risk AML. At the reduced dose schedule used in this study, GO remains associated with significant toxicities while non-significant higher relapse incidence, shorter EFS and shorter OS were observed. [Display omitted] DisclosuresLambert: ASTELLAS: Consultancy; CELGENE/BMS: Consultancy. Pautas: ABBVIE: Consultancy; PFIZER: Consultancy. Gastaud: PFIZER: Consultancy; CELGENE/BMS: Consultancy; ABBVIE: Consultancy; GSK: Consultancy. Barbieux: ASTRA-ZENECCA: Consultancy. Rousselot: Incyte, Pfizer: Consultancy, Research Funding. Castaigne: PFIZER: Consultancy. Dombret: Amgen: Honoraria, Research Funding; Incyte: Honoraria, Research Funding; Jazz Pharmaceuticals: Honoraria, Research Funding; Novartis: Research Funding; Pfizer: Honoraria, Research Funding; Servier: Research Funding; Abbvie: Honoraria; Daiichi Sankyo: Honoraria; BMS-Celgene: Honoraria.

Flow Cytometric Minimal Residual Disease As Risk Stratification Tool in Younger Adults with NPM1 Wild Type Standard Risk Acute Myeloid Leukemia

Response to induction therapy is a powerful prognostic indicator in acute myeloid leukemia (AML). Measurable/minimal residual disease (MRD) assays are being increasingly incorporated into response assessment; however, their value as a surrogate for long-term outcome remains unclear because of assay limitations and the effect of post remission therapies. This is particularly the case for the molecularly heterogeneous NPM1 wild type standard risk patients for whom treatment decisions, including predicting the benefit of allogeneic stem cell transplantation (SCT), are particularly challenging. We investigated the effect on outcome of MRD status compared to other response criteria after each cycle of induction for NPM1 wild type (wt) standard risk patients and other risk groups in a cohort of younger AML patients.Patients and MethodsAs part of the UK NCRI AML17 trial (ISRCTN55675535) for patients with AML or high risk MDS up to the age of 60, prospective flow cytometric MRD (MFC-MRD) monitoring was performed after each course of induction. Any level of MRD detected was considered MRD+ (sensitivity thresholds: ~0.02% by tracking diagnostic leukemic aberrant phenotypes/LAIP, ~0.05-0.1% by “different-from normal” blast LAIP). Clinicians were not informed of MFC-MRD results. Following cycle 1 of induction with daunorubicin/ara-C based therapy, patients were allocated a risk group by a validated score (comprising cytogenetics, WBC, age, secondary disease, blast response to cycle 1 and mutation status). Standard risk patients including ITD wt/ NPM1 wt received a second daunorubicin/ara-C induction with some proceeding to SCT after 1 or 2 consolidation cycles (high dose ara-C).Results2450 non APML patients, median age 51, randomised 4/09 to 12/14, contributed data. For the total cohort, 5 year overall survival (OS) excluding early deaths was 52% for those achieving CR/CRi vs 31% for those not in CR (p<.001).Post cycle 1 (C1) responses were categorised as resistant disease (RD), partial remission (PR) and CR/CRi by clinicians, with CR/CRi subdivided by MFC-MRD assay into MRD+ and MRD-. RD vs PR vs MRD+ vs MRD- frequencies were 15% vs 14% vs 40% vs 31% for all patients with response data (n=1443), 5% vs 7% vs 45% vs 43% for good / standard risk (n=785), 10% vs 8% vs 47% vs 35% NPM1 wt standard risk (n=320).Post C1 response was prognostic for survival: OS for RD vs PR vs MRD+ vs MRD- were 25% vs 36% vs 43% vs 63% all patients; 33% vs 49% vs 51% vs 70% good/standard risk and 28% vs 38% vs 47% vs 59% NPM1 wt standard risk. (p<.0001 throughout). For NPM1 wt standard risk as well as good/standard risk patients adjusted analyses confirmed significant OS differences between C1 RD vs PR or MRD+ but not PR vs MRD+. Results were similar when censored at SCT.Post C1 best response was CRi for 39/446 MRD- vs 49/575 MRD+ overall (11/111 vs 8/149 for NPM1 wt standard risk). C1 CRi was associated with reduced OS overall (39% vs 53% CR, p=.002) and in MRD+ (19% vs 45% CR, p=.001) but not for MRD- patients. OS for NPM1 wt standard risk MRD+ was 25% CRi vs 43% CR (p=0.4). No significant difference was detected for 5 year cumulative incidence of relapse (CIR).The effect on outcomes of MRD status post cycle 2 (C2) for the 806 patients in CR/CRi with MFC-MRD data appeared greater (relapse HR 1.88(1.50- 2.36), p<0.001; OS HR 1.77(1.41-2.22) p<0.001)than C1 (relapse HR 1.70(1.40- 2.06), p<0.001; OS HR 1.50(1.23-1.84) p<0.001). MRD+ levels of ≥0.1% predicted highest CIR in NPM1 wt standard risk patients at C1 (CIR 71%) and C2 (CIR 89%) but any MRD+ at C2 reduced OS (32% vs 63% MRD-, p=0.003) and OS censored at any SCT (35% vs 88%). The effect of CR vs CRi was smaller post C2 although outcomes were still worst in CRi/MRD+ve patients.Of 204 NPM1 wt standard risk patients with C2 MRD data 83 had an allograft (44 in CR1: 29 MRD-, 15 MRD+). In Mantel-Byar analyses, although numbers were small, there was a suggestion that transplant could be considered in MRD +ve (HR 0.72 (0.31-1.69)) but not MRD -ve patients (HR 1.68 (0.75-3.85); p-value for interaction p=0.16).ConclusionMRD status by MFC refines response criteria at induction time-points to differentiate NPM1 wild type standard risk patients with poor outcome and helps define a group of patients who may benefit from SCT in CR1. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Bone Marrow Stroma Protects FLT3 Acute Myeloid Leukemia (AML) through CYP3A4-Mediated Drug Metabolization of FLT3 Tyrosine Kinase Inhibitors (TKIs)

IntroductionFms-like tyrosine kinase 3 internal tandem duplications (FLT3/ITD) are among the most common mutations found in AML patients. Interestingly, although initial remission rates of FLT3/ITD AML are similar to standard-risk AMLs, these AML have a high relapse rate and poor overall survival. Moreover, FLT3 TKIs have been much more successful in eliminating peripheral FLT/ITD blasts, than eradicating marrow disease and generating durable responses clinically. Studies have addressed several mechanisms underlying the FLT3 TKI resistance, including a protective role for the bone marrow (BM) microenvironment. However, the mechanisms underlying this protection remain unknown. We previously showed that human bone marrow stromal cells (BMSCs) highly express most cytochrome P450 (CYP) enzymes, and these enzymes are a major means of chemoprotection within the BM niche (Alonso et al. Oncotarget 2015). Among them, CYP3A4 is a key enzyme responsible for hepatic inactivation of many chemotherapy agents, including many FLT3 TKIs used clinically. Therefore, we hypothesized that BM stromal CYP3A4 contributes to BM microenvironment mediated FLT3 TKI resistance by FLT3/ITD AML.Methods and ResultsTo test our hypothesis in vitro, FLT3/ITD AML cell lines (MV4-11 and Molm14) were co-cultured with FLT3 TKIs with or without BMSCs (F/STRO), and their clonogenic activity was assessed. We found that 3 different clinically active FLT3 TKIs (sorafenib, quizartinib, and gilteritinib), while active against the FLT3/ITD AML in liquid culture, had little activity when co-cultured with BMSCs (Figure 1A). Moreover, when FLT3/ITD AMLs were co-cultured with BMSCs with short hairpin (sh) RNA knockdown of CYP3A4 (shCYP3A4), the activity of the FLT3 TKIs was restored (Figure 1A). To test the hypothesis in vivo, we investigated the growth of FLT3/ITD AML tumors containing control or shCYP3A4 human primary BMSCs upon FLT3 TKI treatments in a xenograft mouse model. We observed that the tumors with shCYP3A4 BMSCs were significantly more sensitive to FLT3 TKI treatments compared to those with control BMSCs.To further examine the role of CYP3A4 in BMSCs, we examined the inhibitory effects of FLT3 TKIs in the absence of stromal cellular contact with FLT3/ITD AML cells. FLT3 TKI-containing media was incubated with and without control BMSCs and shCYP3A4 BMSCs, and then incubated with FLT3/ITD AML cells. Western blot performed on the FLT3/ITD AML cells showed that FLT3 TKI media conditioned with BMSCs blocked the inhibition of FLT3 phosphorylation, and this was reversed by CYP3A4 knockdown in the BMSCs. These results demonstrated that stromal CYP3A4 protected against FLT3 TKIs even in the absence of stromal cell contact.We next studied if combining a clinically active CYP3A4 inhibitor with FLT3 TKIs would increase the FLT3/ITD AML cells' sensitivities to FLT3 TKIs in the presence of BMSCs. Clarithromycin, a clinically active CYP3A4 inhibitor, significantly reversed the protective effects of BMSCs to 3 different FLT3 TKIs (Figure 1B).ConclusionsThese data confirm the protective effects of stroma for FLT3/ITD AML against FLT3 TKIs. Further, we found for the first time that CYP3A4 expressed in BMSCs contributes to BMSC mediated FLT3/ITD AML protection against FLT3 TKIs both in vitro and in vivo mouse model. Our results also demonstrated that combining FLT3 TKIs with clinically available CYP3A4 inhibitors may become a promising strategy to overcome drug resistance in FLT3/ITD AML treatment. [Display omitted] DisclosuresLevis:Novartis: Consultancy, Honoraria, Research Funding; Daiichi Sankyo, Inc.: Consultancy, Honoraria; Astellas Pharma Us: Consultancy, Research Funding; Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Limited: Research Funding; FujiFilm: Research Funding.

Added prognostic value of secondary AML-like gene mutations in ELN intermediate-risk older AML: ALFA-1200 study results

AbstractIn this study, we aimed to refine prognostication of older with acute myeloid leukemia (AML) after intensive chemotherapy. Five hundred and nine patients aged 60 years or older (median age, 68 years) were prospectively enrolled in the intensive Acute Leukemia French Association (ALFA)-1200 trial between 2012 and 2016, and 471 patient samples were submitted to multigene analysis. Mutations in any of 8 genes frequently altered in myelodysplastic syndromes (MDS), including ASXL1, SRSF2, STAG2, BCOR, U2AF1, EZH2, SF3B1, and ZRSR2, defined a secondary AML (sAML)-like disease, as reported. Of the samples analyzed, 48% included sAML-like gene mutations. These mutations were associated with a shorter event-free survival, both overall (hazard ratio, 1.46; 95% confidence interval, 1.19-1.79; P < .001) and within the European LeukemiaNet (ELN)-2017 intermediate-risk subgroup (hazard ratio, 1.52; 95% confidence interval, 1.01-2.28; P = .044), which excludes ASXL1-mutated cases by definition. We therefore included patients with intermediate-risk AML carrying sAML-like mutations in a single high-risk patients group together with adverse-risk patients with AML, whereas other intermediate-risk patients were included in a standard-risk group together with favorable-risk patients (high-risk/standard-risk patient ratio, 1.00). Using this 2-class risk assessment, we observed that transplantation prolonged overall survival from remission in patients with high-risk AML only, not in patients with standard-risk AML. Routine analysis of sAML-like gene mutations may thus improve the definition of high-risk older patients with AML, and better identify the half of older patients who clearly derive survival benefit from allogeneic transplantation in first remission. This trial was registered at www.clinicaltrials.gov as #NCT01966497.

Pediatric LSC3 (pLSC3) Score Derived from DNMT3B-CD34-GPR56 As a Prognostic Tool to Predict AML Patient Outcome: Results from Two Independent Pediatric AML Cohorts

Introduction: Resistance and relapse remain major obstacles in the treatment of acute myeloid leukemia (AML). Pre-existence and persistence of drug resistant leukemia stem cells (LSCs) is considered one of the major causes of relapse. A previous study (Ng et al., 2016) reported a prognostic signature of 17 genes (LSC17 score) differentially expressed in LSC+ compared to LSC- cell fractions that predicted outcome in patients with AML thereby classifying patients into high and low risk groups. The goal of this study is to determine the validity of LSC17 score in pediatric AML patients and to enhance its clinical utility by exploring a new score with limited number of stem cell genes.Methods: 150 pediatric patients with AML enrolled in the multicenter AML02 clinical trial (ClinicalTrials.gov Identifier: NCT00136084) with Affymetrix U133A microarray gene expression data and clinical data were included in the study. Since only 14 of the 17 genes were represented on the Affymetrix U133A gene chip we tested the validity of the LSC14 score using the previously defined equation (Ng et al, 2016) with multiple clinical endpoints such as minimum residual disease (MRD), event free survival (EFS) and overall survival (OS).To reduce the model complexity, we applied a penalized regression algorithm called the least absolute shrinkage and selection operator (LASSO) implemented in the glmnet R-package using event free survival (EFS) as an outcome variable. Score of the new equation, which included three genes, was designated as pediatric-LSC3 (pLSC3). pLSC3 was tested in the AML02 cohort for association of high or low pLSC3 (based on the median value) with clinical endpoints mentioned above. pLSC3 score equation was validated using publically available gene-expression data from 117 pediatric relapse enriched AML patient cohort enrolled in Children's Oncology Group (COG) protocol (TARGET database). COX-proportional hazard models and Log rank test were used for survival data analysis.Results: AML02 cohort: Patients with high LSC14 scores (greater than median), had significantly worse MRD (p<0.0001), EFS (HR = 3.72, P <0.00001) and OS (HR = 4.85, P <0.00001) compared to patients with low LSC14 scores. After applying LASSO regression to simplify the score equation, only three genes (DNMT3B, CD34 and GPR56) remained significant to the model fit of the EFS data thus we created a pLSC3 with coefficients as described in the equation: pLSC3_SCORE = (DNMT3B*0.0431) + (CD34*0.00076) + (GPR56*0.0326). Patients were classified as high or low pLSC3 and patients with high pLSC3 scores had significantly worse EFS (HR=3.595, P < 0.0001; Figure 1A) and OS (HR= 4.53, P<0.0001) and higher MRD after induction 1 and induction II, respectively (P<0.00001 and p=0.0001 respectively; Figure 1C). These results were further validated in an independent cohort of patients from TARGET database, where higher pLSC3 score was associated with worse EFS, OS and MRD (EFS: HR=1.64, P=0.0248; Figure 1B, OS: HR = 1.77, P = 0.0349 and MRD p=0.0002, Figure 1D). Consistent results were also observed with high pLSC3 predictive of significantly worse outcome within standard risk group patients within both AML02 and COG cohorts (AML02-EFS: HR = 2.97, P = 0.0153, COG-EFS: HR = 2.22, P = 0.0096; Figure 1E and F respectively).In a multivariate COX regression model, pLSC3 score groups was the only significant covariate (table 1). It explained 13.1% of variability in EFS and 11.6% of variability in OS, while other prognostic factors such as risk groups, FLT3 status, treatment arm and age collectively explained 15.1 and 12.1 % of variability.Discussion: In summary, our results show validity of a previously defined LSC14 score in a pediatric AML population from the multicenter AML02 clinical trial. To enhance the clinical utility, score equation was further simplified and the final score (pLSC3) was derived from three genes: DNMT3B, which encodes for DNA methyltransferase; CD34, an important cell surface marker for early-undifferentiated LSCs; and GPR56, a G protein coupled receptor of significance in AML. Given that there is need to refine classification of a highly heterogeneous group of patients with standard risk AML, we show that differentiating standard risk patients based on pLSC3 score should be considered in the future. We show the relevance of pLSC3 in two independent cohorts, opening up opportunities to improve treatment outcomes of pediatric patients with AML. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Epigenetic heterogeneity affects the risk of relapse in children with t(8;21)RUNX1-RUNX1T1-rearranged AML.

The somatic translocation t(8;21)(q22;q22)/RUNX1-RUNX1T1 is one of the most frequent rearrangements found in children with standard-risk acute myeloid leukemia (AML). Despite the favorable prognostic role of this aberration, we recently observed a higher than expected frequency of relapse. Here, we employed an integrated high-throughput approach aimed at identifying new biological features predicting relapse among 34 t(8;21)-rearranged patients. We found that the DNA methylation status of patients who suffered from relapse was peculiarly different from that of children maintaining complete remission. The epigenetic signature, made up of 337 differentially methylated regions, was then integrated with gene and protein expression profiles, leading to a network, where cell-to-cell adhesion and cell-motility pathways were found to be aberrantly activated in relapsed patients. We identified most of these factors as RUNX1-RUNX1T1 targets, with Ras Homolog Family Member (RHOB) overexpression being the core of this network. We documented how RHOB re-organized the actin cytoskeleton through its downstream ROCK-LIMK-COFILIN axis: this increases blast adhesion by stress fiber formation, and reduces mitochondrial apoptotic cell death after chemotherapy treatment. Altogether, our data show an epigenetic heterogeneity within t(8;21)-rearranged AML patients at diagnosis able to influence the program of the chimeric transcript, promoting blast re-emergence and progression to relapse.

Comparison of Reduced-Intensity Conditioning with Fludarabine/Busulfan and Fludarabine/Melphalan in Patients 50 Years or Older

Introduction: Reduced-intensity conditioning (RIC) regimens have been increasingly used in allogeneic hematopoietic stem cell transplantation (allo-HCT), because they can reduce non-relapse mortality in comparison with standard myeloablative conditioning regimens and allow previously ineligible patients such as older patients and patients with comorbidities to receive allo-HCT. However, the optimal RIC regimen in allo-HCT remains unclear. Therefore, we conducted a nationwide retrospective study to compare the transplant outcomes between fludarabine/busulfan (FB) and fludarabine/melphalan (FM) regimens which are two most widely used RIC regimens for allo-HCT.Methods: We retrospectively analyzed 2008 patients aged 50 years or older with acute myeloid leukemia (AML) (n=1219), acute lymphoblastic leukemia (ALL) (n=209), or myelodysplastic syndrome (MDS) (n=580) who underwent allo-HCT using fludarabine/busulfan (FB) or fludarabine/melphalan (FM) between 2001 and 2014. The patients who underwent cord blood transplantation were excluded in this study, but those who received low dose of total body irradiation in addition to the FB or FM regimen were included. Acute leukemia in the first or second remission and MDS excluding refractory anemia with excess blasts or leukemic transformation were defined as standard-risk diseases. Clinical data for these patients were obtained from the Transplant Registry Unified Management Program (TRUMP), which includes clinical data of HCT performed in Japan. We compared the outcomes between FB2 (busulfan at 8 mg/kg po or 6.4 mg/kg iv, n=738), FB4 (busulfan at 16 mg/kg po or 12.8 mg/kg iv, n=749), and FM140 (melphalan at 140 mg/m2, n=521).Results: The median ages were 60 (50-74), 60 (50-75), 59 (50-71) years in the FB2, FB4, and FM140 groups, respectively. The cumulative incidences of grade II-IV and III-IV acute GVHD were 30.7% (95% confidence interval (CI), 27.3-34.1%) and 10.1% (95% CI, 8.0-12.4%), 36.8% (95% CI, 33.3-40.2%) and 12.5% (95% CI 10.2-15.0%), and 39.3% (95% CI, 35.0-43.5%) and 15.2% (95% CI, 12.2-18.4%) in the FB2 group, the FB4 group, and FM140 group, respectively (p=0.0061 and p=0.050). The incidence of relapse at 3 years was higher in the FB2 group (38.2%; 95% confidence interval (CI), 34.5- 41.9%) than the FB4 group (31.5%; 95% CI, 28.0- 35.0%) or the FM140 groups (28.3%; 95% CI, 24.2- 32.4%, p<0.001, Figure 1A). Conversely, the incidence of non-relapse mortality was lower in the FB2 group (19.7%; 95% CI, 16.7- 22.9%) than the FB4 group (27.4%; 95% CI, 23.9- 30.9%) or the FM140 group (26.2%; 95% CI, 22.3- 30.3%, p=0.0015, Figure 1B). Overall survival (OS) at 5 years in patients with high-risk AML and MDS in the FM140 group (35.3%; 95% CI, 26.2-44.6% and 57.0%; 95% CI 45.2-67.2%) was superior to that in the FB2 group (15.2%; 95% CI 9.3-22.3% and 38.1%; 95% CI 28.8-47.2%) or FB4 group (20.8%; 95% CI, 14.5- 27.8% and 33.8%; 95% CI 23.9-43.9%) (p=0.0028 and p=0.0065, Figure 2A, B), although OS in patients with standard-risk AML or MDS did not differ among the 3 groups. On the other hand, no statistically significant differences were found in OS in patients with standard- and high-risk ALL in the FB2, FB4, and FM140 groups (OS at 5 years in standard-risk ALL patients: 54.9%; 95% CI 40.2-67.4%, 33.6%; 95% CI 10.9-58.4%, and 49.2%; 95% CI 34.8-62.1%; p=0.32, and OS at 3 years in high-risk ALL patients: 20.8%; 95% CI 0.9-59.5%, 0%; 95% CI NA-NA, and 11.3%; 95% CI 0.7-38.3%; p=0.54, respectively). In multivariate analysis (Table 1), the non-relapse mortality rate was significantly higher in the FB4 and FM140 groups than that in the FB2 group (hazard ratio (HR) 1.49; 95% CI 1.17-1.90; p=0.0012 and HR 1.46; 95% CI, 1.14-1.87; p=0.0029). On the other hand, the relapse rate was significantly lower in the FB4 and FM140 groups than that in the FB2 group (HR 0.69; 95% CI, 0.56-0.86; p<0.001 and HR 0.54; 95% CI, 0.43-0.68; p<0.001), and overall survival (OS) was better in the FM140 group than in the FB2 group (HR 0.81; 95% CI, 0.68-0.97; p=0.021), although there were no significant differences between the FB4 and FB2 groups (HR 1.08; 95% CI, 0.91-1.27; p=0.39).Conclusion: FM140 was associated with better OS in patients with high-risk AML and MDS than FB2 because of lower relapse rate and FB4 was not superior to FB2 in OS, although OS in patients with standard-risk disease did not differ among the 3 groups. A large prospective study is warranted to confirm these findings. [Display omitted] [Display omitted] [Display omitted] DisclosuresKako:Otsuka Pharmaceutical: Honoraria. Kanda:Otsuka Pharmaceutical: Honoraria, Research Funding.

Using Chargemaster Data to Understand Childhood Leukemia Costs

Introduction: Recent changes in health care financing have increased the out-of-pocket costs borne by families. The potential "financial toxicity" can pose challenges for families beyond those encountered in the treatment of cancer. Several families of children diagnosed with leukemia at our center have recently requested information on the expected costs of the planned treatment. There is little published information on the patient's cost (that is, the health system patient charges) for childhood cancer treatment, and costs can be expected to vary between patients and institutions. To better understand the cost of leukemia treatment at our center, the charges accumulated over the course of therapy for a cohort of children with acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) were analyzed.Methods: A sample of children who successfully completed treatment at our institution in 2014 using the current Children's Oncology Group (COG) treatment protocols for standard risk ALL (AALL0932) and AML (AAML0531) was chosen. All inpatient and outpatient charges for the entire course of therapy for each child were used to determine total charges for treatment and types of charges (room and board, medications, laboratory, radiology, professional fees). Using our health system's chargemaster data, the expected charges for ALL therapy based upon protocol requirements was estimated and compared to actual charges. We analyzed high frequency and high cost items to identify potential drivers of charges to suggest areas for potential cost reduction.Results: Charges for 11 patients with ALL and 7 with AML were assessed. The average charges for ALL therapy were $311,000 (range $182,000-634,000) and for AML were $1.05 million (range $733,000-$1,346,000). For ALL, room and board, medications, and laboratory accounted for 37%, 16%, and 19% of charges respectively. For AML, room and board medications and laboratory accounted 66%, 14%, and 14% of charges respectively. (Figure 1) Low risk ALL treatment had higher overall charges driven by increased inpatient chemotherapy. Expected charges for low risk ALL treatment were $245,000 compared to $165,000 for average risk standard therapy. Actual charges however did not differ greatly between low risk and average risk ALL subtypes. (Figure 2) The highest charges for items in ALL therapy were for intravenous immune globulin (IVIG), asparaginase, and inpatient stays (intensive care and oncology ward). For AML, the most expensive items were inpatient stays (intensive care and oncology ward), blood products, and complete blood counts (CBC).Conclusion: AML therapy is more expensive than ALL therapy, driven primarily by the prolonged hospitalizations awaiting count recovery after each cycle. ALL treatment charges were highly variable depending upon experienced toxicities and arm of therapy. Low risk therapy for ALL has the potential to be the most expensive ALL therapy due to six inpatient courses of intermediate dose methotrexate. Estimates for ALL therapy charges did not capture medical toxicity expense or other supportive care related charges. Many drivers of cost cannot be altered without affecting quality of care, but increasing outpatient therapy has the potential to provide the largest reduction in charges. The use of IVIG in some ALL patients and the frequency of CBCs in AML patients highlighted areas where clinical interventions could be implemented without obvious decrements to care quality and with potential cost savings. Understanding data on patient charges can provide an expected range of charges for families and may inform areas to target for cost savings. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Assessment of Minimal Residual Disease in Standard-Risk AML

BackgroundDespite the molecular heterogeneity of standard-risk acute myeloid leukemia (AML), treatment decisions are based on a limited number of molecular genetic markers and morphology-based assessment of remission. Sensitive detection of a leukemia-specific marker (e.g., a mutation in the gene encoding nucleophosmin [NPM1]) could improve prognostication by identifying submicroscopic disease during remission.MethodsWe used a reverse-transcriptase quantitative polymerase-chain-reaction assay to detect minimal residual disease in 2569 samples obtained from 346 patients with NPM1-mutated AML who had undergone intensive treatment in the National Cancer Research Institute AML17 trial. We used a custom 51-gene panel to perform targeted sequencing of 223 samples obtained at the time of diagnosis and 49 samples obtained at the time of relapse. Mutations associated with preleukemic clones were tracked by means of digital polymerase chain reaction.ResultsMolecular profiling highlighted the complexity of NPM1-mutated AML, with segregation of patients into more than 150 subgroups, thus precluding reliable outcome prediction. The determination of minimal-residual-disease status was more informative. Persistence of NPM1-mutated transcripts in blood was present in 15% of the patients after the second chemotherapy cycle and was associated with a greater risk of relapse after 3 years of follow-up than was an absence of such transcripts (82% vs. 30%; hazard ratio, 4.80; 95% confidence interval [CI], 2.95 to 7.80; P<0.001) and a lower rate of survival (24% vs. 75%; hazard ratio for death, 4.38; 95% CI, 2.57 to 7.47; P<0.001). The presence of minimal residual disease was the only independent prognostic factor for death in multivariate analysis (hazard ratio, 4.84; 95% CI, 2.57 to 9.15; P<0.001). These results were validated in an independent cohort. On sequential monitoring of minimal residual disease, relapse was reliably predicted by a rising level of NPM1-mutated transcripts. Although mutations associated with preleukemic clones remained detectable during ongoing remission after chemotherapy, NPM1 mutations were detected in 69 of 70 patients at the time of relapse and provided a better marker of disease status.ConclusionsThe presence of minimal residual disease, as determined by quantitation of NPM1-mutated transcripts, provided powerful prognostic information independent of other risk factors. (Funded by Bloodwise and the National Institute for Health Research; Current Controlled Trials number, ISRCTN55675535.)

Two or More Chemotherapy Consolidation Courses, Followed By Autologous Bone Marrow Transplantation, and MRD Negativity, Give Long Term Overall Survival in Acute Myeloid Leukemia Patients

Introduction. Autologous Bone Marrow Transplantation (Auto-BMT) is currently rarely used in the treatment of Acute Myeloid Leukemia (AML). However, it may represent a good therapeutic option in a specific subset of patients, mainly in consolidation of both low risk (LR) and MRD negative AML without an available HLA matched donor.Aims. To review our database of AML patients who received Auto-BMT from 2005 to 2014 and who were referred to Bologna Institution, in order to assess the efficacy of the procedure in terms of Overall Survival (OS) and Disease Free Survival (DFS).Patients and methods: From 2005 to 2014, 98 AML patients underwent Auto-BMT in several Italian Institutions. 89/98 patients are evaluable for survival and outcome data. The 89 patients considered (42 female, 47 male), had a median age of 49 years (range 15-70). Cytogenetics was performed in all patients by conventional karyotype (22 patients were also analyzed by Single Nucleotide Polymorphisms Array); molecular analysis (FLT3 TKD and ITD, and NPM1 mutational analysis) was available for 51/89 patients. Molecular monitoring by specific fusion transcripts (CBF-MYH11 and AML1-ETO) was performed in CBF positive leukemias (inv(16) and t(8;21)) at the time of diagnosis, after induction, consolidation courses, and every 3 months in the first 2 years of follow-up. Based on this data, and according to ELN guidelines, a risk stratification identified 41 patients with a LR AML (t(8:21), inv(16) or NPM1+/FLT3- with normal karyotype), 4 patients with a high risk (HR) AML (complex karyotype or FLT3 ITD mutated or inv(3) or t(6;9)) and 44 patients with a standard risk (SR) AML (normal karyotype, other alterations).Results. All the patients received an induction chemotherapy treatment, as follows: a "3+7-like" course in 48 cases, a Fludarabine-based regimen in 20 patients and a Gemtuzumab-ozogamicin (GO)-based regimen in 21. 83/89 (93.3%) patients received a median of 2 consolidation courses of chemotherapy (range 1-4) before proceeding to Auto-BMT, performed in 1st CR. 6/89 (6.7%) patients received Auto-BMT in first relapse.41 patients relapsed after auto-BMT and were treated with a re-induction chemotherapy, or were enrolled in clinical trials. 24 patients reached a 2nd complete remission, and 12 patients underwent an allogeneic BMT in 2nd CR.With a median follow up of 6 years, the median Overall Survival (OS) of the entire population was 64.3 months (range 5.8-294.2 months); the 1 year OS and the 5 years OS were, 97.1%, and 67.9%, respectively. The median Disease Free Survival (DFS) of the 83 patients treated with Auto-BMT in 1st CR was 36 months (range 1.3-293 months). The 1-year DFS and the 5-years DFS were 85% and 56.7%, respectively. Transplant related mortality (TRM, death in 100 days after BMT) was 1.2% for auto-BMT and 6.5% for allogeneic BMT.First, to assess the role of the number of consolidation courses we compared patients who received none or 1 consolidation course with patients who received 2 or more cycles, who showed a better OS (p= 0.0061, Figure 1). There was no statistical difference in terms of OS between young and elderly patients (cut off=65 years).Second, we compared patients who achieved a negative minimal residual disease status before auto-BMT (n=37) with patients who did not (n=9). MRD negativity offered a significantly better outcome in terms of 5-years OS (83.4% and 50% respectively); the median OS of MRD neg was not yet reached; the median OS of MRD pos was 27 months (p= 0.0130) (Figure 2).Conclusions: Auto-BMT offers a chance to achieve long-term DFS and OS if used as a consolidation therapy both in patients with LR and SR AML. The major role could be played in MRD negative patients, offering the best chances to achieve a long-term OS. Auto-BMT can be also a good choice as consolidation therapy for elderly patients, in which allo-BMT could induce high morbidity and mortality rates. The small patients cohort and the retrospective analysis don't allow us to define the best induction therapy to be used before auto-BMT. However, based on our findings we suggest a therapy schedule including two or more consolidation courses in patients who obtain a first CR, and to proceed then to auto-BMT.Acknowledgments: work supported by ELN, AIL, AIRC, Progetto Regione-Università 2010-12 (L.Bolondi), Fondazione del Monte di Bologna e Ravenna, FP7 NGS-PTL project. [Display omitted] [Display omitted] DisclosuresSoverini:Novartis, Briston-Myers Squibb, ARIAD: Consultancy. Rodeghiero:Celgene Corporation: Honoraria, Research Funding. Cavo:Janssen-Cilag, Celgene, Amgen, BMS: Honoraria. Martinelli:AMGEN: Consultancy; Novartis: Consultancy, Speakers Bureau; Ariad: Consultancy; BMS: Consultancy, Speakers Bureau; ROCHE: Consultancy; Pfizer: Consultancy; MSD: Consultancy.

Unmanipulated Haploidentical Hematopoietic Stem Cell Transplantation Might Overcome the Adverse Effect of Minimal Residue Disease Pretransplant in Patients with Standard-Risk Acute Myeloid Leukemia

▪BackgroundsIn patients with acute myeloid leukemia (AML), detection of minimal residual disease (MRD) before allogeneic hematopoietic stem cell transplantation (HSCT) is correlated to the risk of relapse. Persistent intensive chemotherapy might further reduce the load of leukemia pretransplant, but it also brings the risk of treatment mortality. Unmanipulated haploidentical HSCT which was established by Peking University Institute of Hematology would induce a stronger graft-versus-leukemia (GVL) effect in compared with sibling transplantation. Moreover, our previous studies have demonstrated that the prophylactic use of modified donor lymphocyte infusion (DLI) would significantly increase the survival of patients with progressive acute leukemia. However, for those patients with standard-risk AML who has the detectable MRD before transplant, whether they would benefit from further chemotherapy to reduce MRD, or should receive active intervention strategy towards leukemia relapse is still unknown.MethodsThe MRD level was measured immediately before and after transplant in 86 standard-risk AML patients who received unmanipulated haploidentical HSCT at the Peking University Institute of Hematology from January, 2011 to May, 2012. All patients were selected from the clinical trial which was registered at http://www.chictr.org as No. ChiCTR-TRC-11001761 if the following criteria were met: having standard-risk AML, defined as first or second complete remission without t(9;22)(q34; q11), t(15;17), inv(16)(p13q22), t(16;16)(p13; q22) or t(8;21)(q22; q22) cytogenetic abnormalities. The duration of follow-up after HSCT was 980 days (range, 55-1277). MRD were examined by 8-color flow cytometry and real-time quantitative polymerase chain reaction (RQ-PCR). WT1+ was defined as a transcript >0.60% and FCM+ was defined as >0.01% of cells with leukemia-associated aberrant immune phenotypes (LAIPs). Patients had at least one of FCM+/WT1+ before transplant were termed as MRD+. The combinative criteria for positive MRD (MRDco+) were defined as 2 consecutive FCM+ or WT1+ results or both FCM+ and WT1+ in a single sample within 1 year after transplantation.ResultsAmong these 86 AML patients, there were 42 and 44 cases in the 6mg/kg ATG and 10mg/kg ATG group, respectively. The proportion of patients with or without detectable MRD before transplantation was comparable in those two groups with different dose of ATG. During the follow-up, 9 patients underwent hematological relapse at a median time of 157 days (range, 30-726). Finally 18 patients died and 7 patients died of relapse. The number of FCM+/WT1+ patients pretransplant was 8 and 16, respectively. There were 18 patients met the criteria of MRD+ before HSCT. None of the FCM+, WT1+ and MRD+ pretransplant but the FCM+ (P=0.001), WT1+ (P=0.013) and MRDco+ (P<0.001) posttransplant was significantly correlated to relapse during univariate analysis. A multivariate Cox regression analysis that included age, sex, relationship and matched status of blood type between donor and recipient, GVHD and MRD status before and after transplantation was performed. It showed that only MRDco+ posttransplant was an independent risk factor of leukemia relapse (P=0.022, HR=4.653, 95%CI:1.249-17.334). Through Kaplan-Meier analysis, a significant difference in cumulative incidence of relapse between patients with or without posttransplant MRDco+ would be observed (P=0.012). However, it seemed that MRD status before transplant might not affect leukemia relapse after haploidentical HSCT.Conclusion Although detectable MRD before HSCT is a strong prognostic factor, its adverse impact could be diminished and overcome in patients with standard-risk AML receiving our unmanipulated haploidentical HSCT. These kinds of patients might not need to receive further intensive chemotherapy or active intervention around HSCT. DisclosuresNo relevant conflicts of interest to declare.

Autologous Stem Cell Transplantation in Acute Myeloid Leukemia; A Single Centre “Real Life” Study

Background.Autologous stem cell transplant is a consolidation treatment for standard risk acute myeloid leukemia (AML). Traditional preparative regimens, most of all Busulphan (Bu) based, have been widely used, but the relapse risk is high and extra hematologic toxicity is a concern. Even if the combination of Busulphan intravenous formulation in association with cyclophosphamide has shown lower toxicity and suggests favorable safety and efficacy, new drugs associations are continuously explored.Methods.Since April 2004 to January 2013, 39 consecutive patients (20 males, 19 females ) entered this study. Median age at transplant was 51 years (range 21-79). Cytogenetic risk classification at diagnosis was good in 4, intermediate in 29 and poor in 6. At transplant time thirty-six patients were in first complete remission, one in partial remission and two in second complete remissions. Mean number of induction chemotherapy courses received was 2.2.Conditioning consisted of oral Busulfan/Cyclophosphamide in 13, intravenous once-daily infusion Busulphan/Cyclophosphamide in 11, intravenous Busulfan/Fludarabine in 8, intravenous Busulphan /Melphalan in 3, Treosulfan/Cyclophosphamide in 3 and Treosulfan /Melphalan in 1. In all patients aged less than 70 years an extended patient and family HLA typing was performed at diagnosis to eventually plane a salvage allogeneic transplant since the beginning of the therapy course.Cytogenetic risk, status of disease, age, number of chemotherapy courses, conditioning regimens have been analyzed. Primary objective was evaluation of overall survival, disease free survival, transplant related mortality and relapse. Comparison analyses were performed with chi square test. All analyses were intention to treat.Results:Of the 39 patients studied one patient (2.5%) died for intracranial haemorrhage during the period of aplasia. All the other engrafted. Of the latter all but one had sustained long-term engraftment. Fourteen patients (36%) did not relapse; all but one are alive and well in continuous complete remission. The only patient who died in remission had lethal infection after salvage allogeneic transplant performed for persistent cytopenias. Twenty-four patients (61%) relapsedOf the 24 relapsed patients eleven (46%) received a second allogeneic transplant after re-induction chemotherapy (all but one from alternative donors: 8 from a matched unrelated donor, 1 from an aplo-identical family member and one from an unrelated cord blood donor). Of them six have died by transplant related mortality and 5 are alive (4 in complete remission).Overall nineteen over 39 patients (49%) are alive after a median follow up of 60 months (range 18-120); seventeen (44% of the entire cohort) in complete remission and two with persistent disease.As regard the 13 patients who didn’t relapse all were transplanted in first complete remission; cytogenetic risk was good in 2, intermediate in 10 and poor in 1; leukemia chemotherapy induction was standard Daunoblastina- Cytarabine (3+7 schedule) in all; transplant conditioning regimens were oral Busulphan/Cyclophosphamide in 6 and intravenous once-daily dose Busulphan in association with cyclophosphamide or Fludarabine in 7.In our analyses the only risk factor impacting on patients’ outcome was patient age at transplant time. Age impacted overall survival and disease free survival. Median age was 61years (range 21-79 ) in patients who died versus 36 years (range21-68) in patients alive (P = 0.03 ) and 64 years ( 21-79) in patients with relapse versus 36 years (21-60) in patients who didn’t relapse (P = 0.04). No other factor had impact on outcome.Conclusion.This limited, single centre experience, confirms that autologous hematopoietic stem cell transplant is an efficient consolidation treatment for AML in remission. Relapse still remains the major problem even if some selective patients may be rescued by allogeneic transplant after a re-induction strategy. A complete HLA family study and preliminary unrelated research should be early performed in all patients for optimal treatment plan and decision. DisclosuresNo relevant conflicts of interest to declare.

Decision Analysis of Allogeneic Hematopoietic Stem Cell Transplantation Versus Chemotherapy in Cytogenetically Standard-Risk Acute Myeloid Leukemia in First Complete Remission: The Impact of FLT3-ITD Profile

BackgroundRecent studies have shown that mutations in internal tandem duplication (ITD) of the FLT3 gene (FLT3-ITD) influence prognosis of cytogenetically normal AML. Our previous decision analysis (Kurosawa et al, Blood 2011) showed that, for patients with cytogenetically standard-risk AML in first complete remission (CR1), allogeneic hematopoietic cell transplantation (HCT) provided better survival with and without a quality-of-life (QOL) adjustment. In this study, we undertook a decision analysis to determine a favored treatment strategy for patients with cytogenetically standard-risk AML in CR1 depending on FLT3-ITD profile.Patients and MethodsInclusion criteria comprised patients aged 16 to 70 years who were diagnosed with intermediate- or unknown-risk AML according to the Southwest Oncology Group cytogenetic classification, and achieved CR1. We collected bone marrow or peripheral blood samples obtained at diagnosis for patients registered, and retrospectively analyzed mutations of FLT3-ITD using DNA extracted from the specimens. We constructed a Markov decision model to reflect outcomes of HCT and chemotherapy in CR1 with three Markov health states; (1) alive after HCT, (2) alive after chemotherapy, or (3) dead. After the decision node of HCT, we added a branch to consider patients who had relapse before receiving HCT in CR1. Cycle length was 3 months and the analyses were performed for 40 cycles, 10 years. Transition probabilities between health states were calculated from the underlying survival rate of the cohort registered. Results were obtained as life expectancy (LE) and QOL-adjusted life expectancy (QALE). We conducted another cross-sectional study to investigate patient-reported QOL from those treated for acute leukemia. Adjusted means of EQ-5D index were adapted from the study as utility estimates that reflect QOL after HCT or chemotherapy (overall mean: post HCT overall, 0.74; post HCT with GVHD, 0.67; post chemotherapy overall, 0.70), and were allowed to change over time (<1 years, 1-2 years, 2-5 years, and 6 years or later from CR1). The analyses were performed using TreeAge Pro software package (TreeAge Software, Williamstown, MA), Stata version 13 (Stata Corp., College Station, TX), and SPSS software (SPSS Inc).ResultsAmong 541 patients registered in the database, mutations in FLT3-ITD were successfully tested in 332 patients (61%). The median age of patients was 52 years and the median follow up was 37 months among survivors. FLT3-ITD mutations were found in 60 patients (18%). Eighty-five patients received allogeneic HCT in CR1 and the other 247 patients were treated with chemotherapy alone during the period of CR1. Patients who received HCT were significantly younger (median age, 41 vs 54 years), more frequently received 2 courses of induction therapy and had dysplasia at diagnosis compared to those who were treated with chemotherapy. In FLT3-positive and FLT3-negative patients, similar proportions of patients received HCT in CR1 (23%, N=14 vs 26%, N=71). In 10-year observation, LE and QALE were 5.9 and 5.1 years, respectively, for total patients. For FLT3-positive patients, allogeneic HCT in CR1 improved LE and QALE by 22 and 17 months, respectively (Table). QALE benefit of HCT was still apparent when we used utility estimates for post HCT with GVHD. Sensitivity analyses across the range of plausible utility estimates (HCT, 0.63-0.83; chemotherapy, 0.64-0.75) did not change the conclusion. For FLT-3 negative patients, HCT benefit was less than 1 year for LE and QALE. Sensitivity analyses showed that chemotherapy would be the preferred strategy when utility estimates for chemotherapy was higher than 0.7. An analysis of FLT3-negative patients stratified by age indicated that HCT in CR1 provided better LE and QALE than chemotherapy alone for patients aged 55 years or older (LE: 77 vs 57; QALE: 54 vs 38 months). In contrast, HCT did not improve LE (74 vs 79 months) or QALE (52 vs 54 months) compared to chemotherapy alone in younger patients with FLT3-negative AML.ConclusionFor patients with FLT3-ITD-positive, cytogenetically standard-risk AML, allogeneic HCT in CR1 offered overall and QOL-adjusted survival benefit. For patients without FLT3-ITD, survival benefit of HCT in CR1 was less apparent. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Prognostic Factors of Reducing Relapse in Patients Undergoing Allogeneic Hematopoietic Cell Transplantation From Related Donor for Acute Myeloid Leukemia: The Japan Society for Hematopoietic Cell Transplantation (JSHCT) AML Working Group

Abstract 3085 Background:A llogeneic hematopoietic cell transplantation (allo-HCT) is a curative treatment option for patients with acute myeloid leukemia (AML), however a major obstacle to success is represented by the relapse of the disease. We retrospectively analyzed prognostic factors to determine whether there are clinical features that predict relapse after transplantation. Patients and Methods:Clinical data were collected from the Transplant Registry Unified Management Program (TRUMP) in Japan. An individual patient data-based analysis was performed on patients with AML who achieved or continued complete remission (CR) after related allo-HCT between 1991 and 2009. A logistic regression model was used to analyze associations between relapse and clinical factors. Patients with CR1 or CR2 were defined as having a standard-risk disease, and patients with ≤CR3 or non-CR were defined as having a high-risk disease. Conditioning regimens included ≤ 8Gy in fractionated doses of total body irradiation, busulfan dose < 9mg/kg, melphalan dose ≤ 140 mg/m2 defined as a reduced-intensity conditioning (RIC) regimen. Results:We analyzed a cohort of 1, 960 patients with AML. The median age at allo-HCT was 42 years (range, 16 to 74 years). Among the 1, 960 patients, 778 patients (38%) relapsed after allo-HCT. The probabilities of survival at 5-year for all patients, standard-risk patients, and high-risk patients were 49%, 61%, and 31%, respectively. In a multivariate analysis, significant factors for no relapse were standard-risk disease at the time of allo-HCT (HR 0.366, 95%CI 0.279–0.480, p<0.0001), presence of chronic GVHD (HR 0.442, 95%CI 0.338–0.579, p <0.0001), and CMV infection (HR 0.452, 95%CI 0.230–0.886, p =0.021). Intensity of conditioning regimen (RIC vs myeloablative), disease subtype of AML, HLA matching (matched donor vs mismatched donor), source of stem cells (peripheral blood vs bone marrow), and presence of acute GVHD did not affect relapse rates. Conclusion:These results indicate that in patients with standard-risk AML at the time of HCT, chronic GVHD or CMV infection after HCT were crucial for reducing relapse. Immune modulation mediated by chronic GVHD or CMV infection may have played a role in preventing disease relapse. The use of donor lymphocyte infusion before relapse may reduce the recurrence for selected patients who do not develop chronic GVHD. Disclosures:No relevant conflicts of interest to declare.

Conventional Cytogenetics, Molecular Profiling, and Flow Cytometric Response Data Allow the Creation of a Two-Tiered Risk-Group System for Risk-Based Therapy Allocation In Childhood AML- a Report From the Children's Oncology Group

AbstractAbstract 761Conventional cytogenetics and morphologic response to induction chemotherapy have historically provided the tools for predicting outcome in patients with acute myeloid leukemia (AML). Several mutations are predictive of clinical outcome and have provided additional tools to help predict outcome in patients with AML without other risk features. Despite these recent advances, our ability to identify specific risk groups has been limited to a subset of patients, and nearly half of the patients with AML are regarded as having standard-risk (SR) disease. We inquired whether adding response by multidimensional flow cytometry (MDF) to data from conventional cytogenetics analysis and presence/absence of genomic alterations of FLT3 (FLT3-ITD), CEBPA, and NPM would provide a more robust risk-stratification system for risk-based therapy allocation. COG AML protocol AAML03P1 collected comprehensive cytogenetics characteristics, mutation profile (FLT3/ITD, CEBPA, and NPM mutation status), and MDF data on most of the 340 eligible patients enrolled on the study. Molecular and cytogenetic data were available for 275 of the 293 (94%) patients with responsive disease at the end of induction I. Disease-free survival (DFS) from the end of induction I was determined based on a combined molecular and cytogenetic risk profile. Risk status was defined based on the presence of t(8;21), inv(16), NPMc, and CEBPA mutations (favorable risk, FR) or the presence of monosomy 7, monosomy 5/del5q, and high allelic ratio FLT3-ITD (high risk, HR). On the basis of this allocation, 88 (32%) patients had FR AML, and 26 (10%) had HR AML. The remaining 161 (59%) patients without specific risk features were considered to have SR AML. DFS at 2 years from the end of induction I was 70%±12% for the FR cohort, 55% ±9% for the SR cohort, and 17%±20% for the HR cohort (p<0.001). The prevalence and prognostic significance of minimal residual disease (MRD) were assessed in the 103 patients with SR disease; 31 (30%) had evidence of MRD by MDF. DFS at 3 years from the end of induction I was significantly worse for those with MRD than for those without it (26%±21% vs. 67%±13%, p=0.01). Corresponding relapse risk in patients with or without MRD was 69%±21% and 30%±13%, respectively (p=0.011). We assessed the clinical impact of MRD in patients with HR or FR disease. Of the 18 patients with HR AML who had MRD data, 8 (44%) had MRD. DFS at 2 years for patients with HR disease with or without MRD was not significantly different (13%±23% vs. 36%±40%; p=0.127). Of the 73 patients with FR AML, MRD was detected in 12 (16%); MRD did not significantly influence DFS at 3 years from the end of induction I in those patients (45%±33% vs. 72%±17%, p=0.138). Thus, although the presence of MRD was significantly associated with worse outcome in patients with SR AML, similar significance could not be demonstrated in the FR or HR cohorts. Clinical outcomes of risk groups were reassessed after combining the MRD data with specific cytogenetic and molecular risk groups, i.e., patients with SR AML and MRD were added to the HR cohort, and those without MRD were added to the FR cohort. In the new risk-stratification system, 57 of 217 (26%) patients were in the HR cohort, and the remaining 160 (74%) patients were in the FR cohort. DFS at 3 years from induction I was 68%±9% for the FR cohort and 20%±16% (p<0.001) for the HR cohort. Cumulative incidence of relapse at 3 years from the end of induction I for those with FR or HR disease was 27%±9% and 71%±17%, respectively (p<0.001). Cytogenetics, molecular genotyping, and post-induction MDF analysis provide a robust means of stratifying all pediatric patients with AML into 2 risk groups with significantly different outcomes. This novel risk-allocation schema will be implemented in the upcoming COG Phase III AML trial. Disclosures:No relevant conflicts of interest to declare.

High Expression of the Very Late Antigen-4 Integrin Independently Predicts Reduced Risk of Relapse and Improved Outcome in Pediatric Acute Myeloid Leukemia: A Report From the Children's Oncology Group

To evaluate the prognostic significance of the integrin cell adhesion molecule very late antigen-4 (VLA-4) in acute myeloid leukemia (AML). We prospectively quantified VLA-4 expression in 216 patients enrolled onto COG-AAML03P1 by flow cytometry and correlated expression levels with disease characteristics and clinical outcome. VLA-4 mean fluorescence intensity (MFI) varied 35-fold (range, 30 to 1,110; median, 219.5). High VLA-4 expression (> median MFI), compared with low expression, was associated with younger age (7.1 v 12.1 years, respectively; P < .001), lower FLT3 internal tandem duplication prevalence (4% v 21%, respectively; P < .001), and higher likelihood of extramedullary disease (16% v 5%, respectively; P = .013). In low- and high-expression groups, rates of remission (89% v 80%, respectively; P = .137) and minimal residual disease (29% v 25%, respectively; P = .700) were similar. Patients with low VLA-4 expression, compared with high expression, had a higher relapse rate (RR; 44% +/- 10% v 24% +/- 9%, respectively; P = .011) and lower disease-free survival (DFS; 48% +/- 11% v 67% +/- 10%, respectively; P = .023) after 3 years. Multivariate analyses showed that low VLA-4 expression was an independent adverse prognostic factor for DFS (hazard ratio [HR] = 1.98; P = .038) and RR (HR = 2.77; P = .009). Subgroup analyses indicated that the prognostic role of VLA-4 expression was most prominent in patients with standard-risk AML, in whom low VLA-4 expression was associated with inferior DFS (34% +/- 16% v 69% +/- 14% for high expression; P = .011) and higher RR (61% +/- 16% v 26% +/- 14% for high expression; P = .009). A similar trend was seen in low-risk but not high-risk patients. High VLA-4 expression is associated with better clinical outcome in pediatric AML and is an independent predictor of relapse that may refine our abilities to stratify patients without identifiable cytogenetic or molecular risk factors.

High Expression of the Very Late Antigen (VLA)-4 (CD49d) Integrin Predicts for Reduced Risk of Relapse and Better Outcome in Pediatric Acute Myeloid Leukemia (AML): A Report From the Children's Oncology Group.

Abstract 1592▪▪This icon denotes an abstract that is clinically relevant.Poster Board I-618 BackgroundPrevious studies highlighted the importance of the cell adhesion molecule, VLA-4, for chemoresistance and minimal residual disease (MRD) in AML, suggesting promise as therapeutic target. By comparison, the prognostic role of VLA-4 in AML remains controversial with retrospective studies implying either adverse or favorable prognosis. Therefore, we prospectively evaluated VLA-4 expression in participants of a recent Children's Oncology Group (COG) AML pilot protocol. MethodsCOG-AAML03P1 enrolled 340 newly diagnosed children (aged 1 month - 21 years) with de novo non-acute promyelocytic AML, excluding those with Down syndrome, and tested the feasibility of combining gemtuzumab ozogamicin (GO) with intensive induction chemotherapy followed by GO-containing intensification therapy or matched related donor stem cell transplantation; 216 patients submitted diagnostic marrow specimens for flow cytometric determination of VLA-4 expression that was then correlated with patient demographics, laboratory characteristics, and clinical outcome. Cytogenetics and molecular prognostic markers were used for risk classification as follows: low risk (mutation in core-binding factor, NPM1, or CEBPa; n=73), high risk (-5/5q-, monosomy 7, or FLT3/ITD with high allelic ratio; n=25), or standard risk (all other patients with cytogenetic/molecular data; n=101); 17 patients had insufficient data for risk classification. ResultsAmong the 216 diagnostic specimens, the mean fluorescence intensity (MFI) of VLA-4 expression varied over 35-fold from a baseline of 30 to 1110 (median, 219.5). Patients with high VLA-expression (>median MFI) were younger (p<0.001), had a lower prevalence of FLT3/ITD (p=0.002). Presence of extramedullary disease (EMD, chloroma or CNS involvement) was significantly higher in patients with high VLA-4 expression (16% vs. 5%, p=0.013), where 17/22 (77%) of patients with EMD had high VLA-4 expression. We initially inquired whether VLA-4 expression as a continuous variable correlated with disease outcome. We demonstrated that over the range of VLA-4 expression levels, every 10-unit increase in VLA-4 MFI corresponded to a 2% decrease in relapse risk (RR; p=0.023) and 2% increase in disease-free survival (DFS) from end of induction I (p=0.015). We subsequently divided the study patients into two cohorts based on median MFI and determined the clinical outcome per median VLA-4 MFI for the entire cohort as well as in specific risk groups (i.e., high risk, low risk, and standard risk). In the evaluation of the entire cohort, patients with high VLA-4 expression had a significantly superior DFS (67% vs 48%, p=0.023) and lower RR (24% vs 44%, p=0.011) compared to those with lower VLA-4 expression. In subgroup analyses, high VLA-4 expression was associated with significantly superior DFS (69% vs 34%, p=0.011) and lower RR (26% vs 61%, p=0.009) in patients with standard-risk AML but not in patients with high or low risk disease. In multivariate Cox regression analyses that included age, cytogenetics, and molecular prognostic factors, low VLA-4 expression remained significantly associated with elevated RR (hazard ratio for low VLA-4: 2.25, p=0.011). Finally, we determined the impact of MRD in the context of VLA-4 expression. The prevalence of MRD was similar for patients with low or high VLA-4 expression (29% vs. 25%, p=0.70). In patients with low VLA-4 expression, those with MRD had a RR of 75% compared to that of 35% for the MRD negative cohort (p=0.005). Corresponding DFS was 19% and 54% for those with and without MRD (p=0.018), a difference mainly attributable to increased RR. In patients with high VLA-4 expression, those with MRD had a RR of 31% vs. 23% for the MRD-negative patients (p=0.28) with a corresponding DFS of 46% and 75% for the MRD positive and negative patients (p=0.014), a difference mainly explained by higher treatment-related mortality in MRD positive patients. ConclusionThis study demonstrates significant heterogeneity of VLA-4 expression in pediatric AML and its association with EMD and clinical outcome. This study further demonstrates that VLA-4 expression is an independent prognostic factor for clinical outcome and can identify the risk status in patients with no identifiable cytogenetic or molecular risk factors. DisclosuresNo relevant conflicts of interest to declare.

Lack of Inhibition of AKT Predicts Clinical Resistance to Therapy with Imatinib + Reinduction Chemotherapy in Relapsed/Refractory c-Kit+ Acute Myeloid Leukemia.

Abstract 2073Poster Board II-50Patients age 18-65 with acute myeloid leukemia (AML) who had persistent leukemia after 3+7 induction with cytarabine and daunorubicin, or relapsed within 24 months of achieving CR, and whose blasts were c-kit (CD117) positive, were enrolled in a phase I/II clinical trial with imatinib on Days 1-10 combined with mitoxantrone 10 mg/m2/day IV Days 4-8, etoposide 100 mg/m2/day IV Days 4-8 and cytarabine 1.5 g/m2 IV q12h × 4 doses on Days 9-10. Imatinib was escalated through 3 dose levels (200, 300, 400 mg daily) in successive 6 patient cohorts. The combination was well-tolerated up to 400 mg/day imatinib, and an additional 15 patients were enrolled at this dose level. Of the 21 patients treated at the 400 mg imatinib dose, 13 (62 %) achieved CR, 7 (33 %) were non-responders, and one died during reinduction. The CR rate was 82% (9/11) among patients with standard risk AML karyotpye, compared to 33% (3/9) for patients with adverse risk karyotype. The CR rate for primary 3+7 non-responders was 6/14 (43%) vs. 7/7 (100%) for relapsed patients. For 14 patients on the Phase II portion of the study, AML blasts from peripheral blood were assayed for pAKT and pERK by flow cytometry, after stimulation with stem cell factor, on Days 1 and 4, pre- and 1 and 2 hours post-imatinib dosing on each day. A heterogeneous response pattern was seen, with some samples exhibiting marked inhibition of pAKT and/or pERK after imatinib therapy, while others demonstrated little change in levels. Of 8 patients achieving CR with re-induction, 6 demonstrated marked (>50%) pAKT inhibition with imatinib therapy, 1 showed no inhibition, while 1 showed intermediate levels of inhibition. In contrast, all 6 non-responders to re-induction demonstrated a lack of pAKT inhibition with imatinib therapy (p<0.01). Of 5 patients with adverse risk karyotype, 4 demonstrated lack of pAKT inhibition with imatinib therapy and none of these responded to re-induction; the remaining patient whose blasts demonstrated pAKT sensitivity to imatinib did achieve CR with re-induction. In contrast pERK inhibition did not correlate with response to therapy. These results indicate that lack of pAKT inhibition in vivo correlates with resistance to re-induction therapy using this regimen. Further studies using agents that are able to inhibit AKT more effectively are warranted to try to overcome drug resistance in this setting. The treatment regimen appears active in relapsed patients, but this requires confirmation in a larger series. Disclosures:Off Label Use: Imatinib for c-kit positive AML.