Abstract: Abstract 80 Background: Mutations in the nucleophosmin-1 gene (NPM1) are the most common genetic abnormalities in acute myeloid leukemia (AML) and define a provisional AML entity in the current WHO classification. In a retrospective biomarker study within a randomized trial of older patients with AML, we demonstrated that patients with mutated NPM1 and absence of a FLT3 internal tandem duplication (ITD) benefit from all-trans retinoic acid (ATRA) as adjunct to conventional chemotherapy (Schlenk et al. Haematologica 2009;94:54–69). Aims: To evaluate the impact of ATRA in combination with conventional chemotherapy on outcome, and to assess the NPM1 mutational status as predictive marker for response to this therapy in younger adult patients with AML entered in the prospective randomized controlled treatment trial AMLSG 07-04 (ClinicalTrials.gov Identifier: NCT00151242). Methods: Patients (18 to 60 years of age) were accrued between August 2004 and August 2009. They were randomized up-front for open-label treatment with ATRA. Induction therapy consisted of two cycles of ICE (idarubicin 12mg/m2, day 1,3,5 [in induction II reduced to d 1, 3]; cytarabine 100mg/m2 continuous i.v., day 1 to 7; etoposide 100mg/m2, day 1–3). For consolidation therapy, patients with high-risk AML, defined either by high-risk cytogenetics or induction failure, were assigned to receive allogeneic hematopoietic stem cell transplantation (HSCT) from a matched related (MRD) or unrelated donor (MUD). Starting from November 2006, AML with FLT3-ITD was also categorized as high-risk. All other patients were assigned to 3 cycles of high-dose cytarabine (HiDAC; 18g/m2 per cycle). In all but patients with core-binding-factor AML an allogeneic HSCT was intended when a MRD was available. During induction cycles, ATRA was given in a dosage of 45mg/m2 from day 6 to 8, and 15mg/m2 from day 9 to 21; and during HiDAC cycles in a dosage of 15mg/m2 from day 6 to 28. The primary end points of the study were event-free survival (EFS) and rate of complete remission (CR) after induction therapy; secondary end points were, relapse-free (RFS) and overall survival (OS). For survival analyses, patients receiving an allogeneic HSCT in first CR were censored at the date of transplantation. Forty patients were treated either with or without ATRA despite being randomized in the opposite treatment arm; predictive marker analyses were performed on a per protocol basis excluding those patients. Results: A total of 1112 patients were randomized, 562 (per protocol 542) in the standard arm, and 550 (per protocol 530) in the investigational arm with ATRA. Median follow-up was 3.3 years. NPM1 mutational status was assessed in 1018 patients (92%) and a mutation was identified in 289 (28%) patients. Pretreatment patient characteristics at diagnosis were well balanced between the standard and the ATRA-arm of the study, except for higher white blood counts (WBC) in the standard arm (median, 16.1/nl vs. 8.9/nl, p=0.001). The CR-rate was significantly increased in NPM1-mutated AML by ATRA (OR, 2.20; p=0.05), independent of the FLT3-ITD status (OR, 0.66; p=0.33); there was no effect of ATRA in NPM1-wild-type AML (OR, 1.00; p=0.99). Multivariable analyses on EFS revealed a significant risk reduction in NPM1-mutated AML by ATRA (hazard ratio [HR], 0.65; p=0.02), whereas there was no effect of ATRA in NPM1-wild-type AML (HR, 0.99; p=0.95). Other significant factors in NPM1-mutated AML were IDH1R132 mutation (HR, 1.72; p=0.04), IDH2R140 mutation (HR,1.73; p=0.03), FLT3-ITD (HR, 1.55; p=0.04), log-transformed WBC (HR, 1.47; p=0.03), and in NPM1-wild-type AML IDH2R172 mutation (HR,1.82; p=0.03), FLT3-ITD (HR, 1.56; p=0.002), logarithm of WBC (HR, 1.20; p=0.02), male gender (HR, 1.34; p=0.003), cytogenetic risk (p 〈 0.001; HR (high-risk vs intermediate risk), 2.18; HR (low-risk vs intermediate risk) 0.31). ATRA had no influence on the cumulative incidence of relapse. OS of patients treated with ATRA (n=549) was significantly better (p=0.02) compared with that of patients not treated with ATRA (n=562). Rates and severity of toxicities were similar in both treatment arms. Conclusions: ATRA in addition to conventional chemotherapy significantly improved response to induction therapy and EFS in NPM1 mutated-AML, as well as OS in the whole cohort of younger adult patients with AML. NPM1 mutation was confirmed as a predictive factor for response to this combination therapy. Disclosures: Schlenk: Roche GmbH: Research Funding; Amgen GmbH: Research Funding; Pfizer GmbH: Research Funding. Krauter:Novartis: Consultancy, Honoraria. Kuendgen:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees. Fiedler:Pfizer: Research Funding.

Abstract: The addition of valproic acid to intensive induction therapy in combination with all-trans retinoic acid did not result in an improvement of clinical outcome. Valproic acid-related hematologic toxicity and higher death rates were observed when valproic acid and idarubicin were given in parallel.

Abstract: High CD33 expression in acute myeloid leukemia (AML) with mutated NPM1 provides a rationale for the evaluation of gemtuzumab ozogamicin (GO) in this AML entity. We conducted a randomized trial to evaluate GO in combination with intensive induction and consolidation therapy in NPM1-mutated AML. PATIENTS AND METHODS Between May 2010 and September 2017, patients ≥ 18 years old and considered eligible for intensive therapy were randomly assigned up front for induction therapy with idarubicin, cytarabine, etoposide, and all- trans-retinoic acid with or without GO. The early ( P = .02) primary end point of event-free survival (EFS) was evaluated 6 months after completion of patient recruitment. RESULTS Five hundred eighty-eight patients were randomly assigned (standard arm, n = 296; GO arm, n = 292). EFS in the GO arm was not significantly different compared with that in the standard arm (hazard ratio, 0.83; 95% CI, 0.65 to 1.04; P = .10). The early death rate during induction therapy was 10.3% in the GO arm and 5.7% in the standard arm ( P = .05). Causes of death in both arms were mainly infections. The cumulative incidence of relapse (CIR) in patients achieving a complete remission (CR) or CR with incomplete hematologic recovery (CRi) was significantly reduced in the GO arm compared with the standard arm ( P = .005), with no difference in the cumulative incidence of death ( P = .80). Subgroup analysis revealed a significant beneficial effect of GO in female, younger (≤ 70 years), and FLT3 internal tandem duplication–negative patients with respect to EFS and CIR. CONCLUSION The trial did not meet its early primary end point of EFS, mainly as a result of a higher early death rate in the GO arm. However, in patients achieving CR/CRi after induction therapy, significantly fewer relapses occurred in the GO compared with the standard arm.

Abstract: Background: Mutations of the NPM1 gene are one of the most frequent genetic aberrations in adult AML. AML with mutated NPM1 is categorized as a disease entity according the WHO-2016 classification and clinically associated with female sex, high white blood cells at diagnosis, normal karyotype and high CD33 antigen expression. We recently showed that patients with NPM1-mutated AML benefit from all-trans retinoic acid (ATRA) as adjunct to intensive induction therapy (Ann Hematol. 2016; 95:1931-1942; Haematologica. 2009;94:54-60). Based on the regular high CD33 expression in AML with mutated NPM1 we hypothesized that gemtuzumab ozogamicin (GO) added to intensive therapy with ATRA may further improve clinical outcome in AML with mutated NPM1. Aim: To evaluate GO in combination with intensive induction and consolidation therapy and ATRA in NPM1 mutated AML within the randomized AMLSG 09-09 trial (NCT00893399) Methods: Between May 2010 and September 2017, patients ≥18 years of age and considered eligible for intensive therapy were randomized up-front for open-label treatment with GO. Induction therapy consisted of two cycles of A-ICE (idarubicin 12mg/m² iv, day 1,3,5 [in induction II and for patients 〉 60 years reduced to d 1, 3]; cytarabine 100mg/m² continuous iv, day 1 to 7; etoposide 100mg/m² iv, day 1-3 [in induction II and for patients 〉 60 years reduced to d 1, 3]; ATRA 45 mg/m²/day po on days 6-8 and 15mg/m² days 9-21, +/- GO 3mg/m² iv day 1). Consolidation therapy consisted of 3 cycles of high-dose cytarabine (HiDAC; 3g/m² [reduced to 1g/m² in patients 〉 60 years] bid, days 1-3; Pegfilgrastim 6mg sc, day 10; ATRA 15 mg/m²/day po, days 4-21; +/- GO 3mg/m² on day 1 [first consolidation only] ). The primary endpoints of the study were event-free survival (EFS) as early endpoint tested 6 months and overall survival (OS) tested 4 years after study completion with sequential testing according the fallback procedure described by Wiens (Statistics 2003;2:211-215). This report focusses on the early EFS endpoint. Further secondary endpoints were response to induction therapy, cumulative incidence of relapse (CIR) and cumulative incidence of death (CID). Results: In total 588 patients were evaluable for analysis (n=296, standard-arm; n=292 GO-arm). Median age was 58.7 years (range, 18.4-82.3 years), ECOG performance status was 0 in 34.1% and 1 in 55.1%, and FLT3-ITD was present in 16.8% of the patients, with baseline characteristics well balanced between the two arms. After first induction therapy death rates were significantly higher in the GO-arm (7.5%) (p=0.02) compared to the standard-arm (3.4%); in both study-arms causes of death were mainly infections. Following induction therapy complete remission (CR) and CR with incomplete count recovery (CRi) were 88.5% and 85.3% (p=0.28), refractory disease (RD) 6.1% and 5.1% (p=0.72), death 5.4% and 9.6% (p=0.06) in the standard- and GO-arm, respectively. Due to prolonged thrombocytopenia after second induction therapy in the GO-arm, the protocol was amended in that GO was omitted in second induction and first consolidation cycles, if prolonged cytopenias were observed during first induction therapy. The study treatment was completed in 197 and 171 patients (p=0.11), allogeneic hematopoietic cell transplantation in first CR was performed in 18 and 21 patients (p=0.51) in the standard- and GO-arm, respectively. Median follow-up was 2.6 years (95%-CI, 2.4-3.1 years). Two- and 4-year EFS were 53% (95%-CI, 48-60%) and 58% (95%-CI, 52%-64%), and 44% (95%-CI, 38-52%) and 52% (95%-CI, 46%-59%) in the standard- and GO-arm, respectively. According to the pre-specified significance level of 0.025, EFS in the GO-arm was not different to that in the standard-arm (p=0.21). In patients achieving CR/CRi after induction therapy, CIR was significantly reduced in the GO-arm compared to the standard-arm (p=0.018), whereas no difference in CID was noted between both arms (p=0.89). Conclusion: The addition of GO to intensive induction therapy with ICE plus ATRA was associated with a higher death rate. In patients achieving a CR/CRi after induction therapy significantly less relapses occurred in the GO- compared to the standard-arm. Disclosures Schlenk: Pfizer: Research Funding, Speakers Bureau. Paschka:Astex: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees, Travel support; Otsuka: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Sunesis: Membership on an entity's Board of Directors or advisory committees; Bristol-Meyers Squibb: Other: Travel support, Speakers Bureau; Jazz: Speakers Bureau; Amgen: Other: Travel support; Janssen: Other: Travel support; Novartis: Membership on an entity's Board of Directors or advisory committees, Other: Travel support, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Other: Travel support, Speakers Bureau; Agios: Membership on an entity's Board of Directors or advisory committees; Takeda: Other: Travel support. Fiedler:Amgen: Other: support for meetíng attendance; Gilead: Other: support for meeting attendance; Pfizer: Research Funding; Amgen: Research Funding; Amgen: Patents & Royalties; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees; ARIAD/Incyte: Membership on an entity's Board of Directors or advisory committees, support for meeting attendance; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; GSO: Other: support for meeting attendance; Teva: Other: support for meeting attendance; JAZZ Pharmaceuticals: Other: support for meeting attendance; Daiichi Sankyo: Other: support for meeting attendance. Lübbert:Cheplapharm: Other: Study drug; Celgene: Other: Travel Support; Janssen: Honoraria, Research Funding; TEVA: Other: Study drug. Götze:Novartis: Honoraria; Takeda: Honoraria, Other: Travel aid ASH 2017; JAZZ Pharmaceuticals: Honoraria; Celgene: Honoraria, Research Funding. Schleicher:Novartis: Membership on an entity's Board of Directors or advisory committees, Other: Investigator; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Ipsen: Membership on an entity's Board of Directors or advisory committees; Eissai: Other: Investigator; Astra Zeneca: Other: Investigator; Pfizer: Speakers Bureau; Janssen: Speakers Bureau; Celgene: Speakers Bureau. Greil:Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; MSD: Honoraria, Research Funding; Janssen: Other: TRAVEL, ACCOMMODATIONS, EXPENSES; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees; BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Honoraria, Research Funding; Astra Zeneca: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sandoz: Honoraria, Research Funding; Amgen: Honoraria, Other: TRAVEL, ACCOMMODATIONS, EXPENSES, Research Funding; Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Heuser:Novartis: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding; Astellas: Research Funding; BergenBio: Research Funding; Karyopharm: Research Funding; Daiichi Sankyo: Research Funding; Sunesis: Research Funding; Tetralogic: Research Funding; Bayer Pharma AG: Consultancy, Research Funding; StemLine Therapeutics: Consultancy; Janssen: Consultancy. Ganser:Novartis: Membership on an entity's Board of Directors or advisory committees. Döhner:Agios: Consultancy, Honoraria; Pfizer: Research Funding; Sunesis: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Astex Pharmaceuticals: Consultancy, Honoraria; Astellas: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria; Seattle Genetics: Consultancy, Honoraria; AROG Pharmaceuticals: Research Funding; Agios: Consultancy, Honoraria; Bristol Myers Squibb: Research Funding; Celator: Consultancy, Honoraria; Astellas: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Astex Pharmaceuticals: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; AROG Pharmaceuticals: Research Funding; Jazz: Consultancy, Honoraria; Jazz: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Pfizer: Research Funding; Sunesis: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Research Funding; Seattle Genetics: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding; Celator: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding.

Abstract: Background Cyto- and molecular-genetic abnormalities evaluated at initial diagnosis are the most powerful prognostic and in part also predictive markers in acute myeloid leukemia (AML) with regard to achievement of complete remission (CR) and survival. Nonetheless, after relapse the prognostic impact of clinical characteristics and genetic abnormalities assessed at initial diagnosis with respect to achievement of subsequent CR and survival are less clear. Aims To evaluate the probability of CR achievement and survival in relapsed AML patients in correlation to clinical characteristics and genetic abnormalities assessed at initial diagnosis as well as treatment strategy. Methods The study includes intensively treated adults with newly diagnosed AML enrolled in 5 prospective AMLSG treatment trials between 1993 and 2009. Patients with acute promyelocytic leukemia were excluded. All patients received intensive therapy, including allogeneic (allo) and autologous (auto) hematopoietic stem cell transplantation (HSCT) during first line therapy. Results A total of 3218 patients (median age, 54 years; range, 16-85 years) were enrolled in 5 AMLSG treatment trials. Of these, 1307 (41%) patients (16-60 years, n=958; ≥61 years, n=349) experienced relapse, n=194 after alloHSCT, n=75 after autoHSCT and 1038 after chemotherapy. Salvage strategies were as follows: (i) n=907, intensive chemotherapy (INT) followed in n=450 by HSCT (matched related donor [MRD], n=114; matched unrelated donor [MUD] , n=303; cord blood graft [CB], n=3; haplo-identical family donor [HID] , n=18; autoHSCT, n=12); (ii) n=100, direct alloHSCT (MRD, n=31; MUD, n=63; HID, n=4) or n=2 autoHSCT (TPL); (iii) n=29, donor lymphocyte infusions (DLI) in patients after alloHSCT in CR1; (iv) n=60, demethylating agents/low-dose cytarabine (NON-INT); (v) n=24, experimental treatment within phase I/II studies (EXP); (vi) all other patients (n=187) received best supportive care (BSC). After salvage therapy CR rate was 38% and after the different treatment approaches as follows: INT, 37%; TPL, 73%; DLI, 38%; NON-INT, 8%; EXP, 29%. After failure to respond to INT, n=159 additional patients achieved a CR2 after HSCT resulting in an overall CR2 rate of 50%. A logistic regression model revealed CEBPA double-mutant (dm) (OR, 6.42; p=0.0001), core-binding factor (CBF) AML (OR, 2.87; p=0.0002), a direct HSCT strategy (OR, 3.32; p=0.0002), and mutated NPM1 (OR, 1.59; p=0.02) as favorable (only if response after HSCT was included) and FLT3-ITD (OR, 0.66; p=0.04), age (difference of 10 years; OR, 0.82; p=0.003), NON-INT (OR, 0.08; p=0.0001) and in trend a previous alloHSCT in CR1 (OR, 0.65; p=0.08) as unfavorable independent parameters for achievement of CR2. Median follow-up for survival after relapse was 4.3 years and survival after 4 years was 22% (95%-CI, 19-25%). Patients proceeding to alloHSCT after first relapse (n=536; MRD, n=145; MUD, n=366; HID, n=22; CB, n=3) had a 4-year survival of 36% (95%-CI, 32-41%) and those not proceeding to alloHSCT of 8% (95%-CI, 6-11%). In univariable analyses the combined genotype mutated NPM1 in the absence of FLT3-ITD (p=0.66) was not associated with a favorable outcome. A multivariable regression model including alloHSCT as a time-dependent co-variable revealed alloHSCT performed after relapse (HR, 0.34; p 〈 0.0001), CEBPAdm (HR, 0.48; p=0.002), CBF- AML (HR, 0.50; p 〈 0.0003) and DLI in relapsed patients with a previous alloHSCT performed in CR1 (HR, 0.40; p=0.002) as significant favorable factors, whereas FLT3-ITD (HR, 1.35; p=0.005) and in trend NON-INT (OR, 1.40; p=0.06) were unfavorable factors. Due to collinearity of FLT3-ITD with duration of first remission (cut point at 1 yr), the latter was not included into the multivariable models. Of 561 patients achieving CR2, 252 experienced 2nd relapse (REL2) and 114 died in CR2. Most REL2 patients (n=117) received INT whereas n=54 received BSC only. Allo- and autoHSCT were performed in 55 and 3 REL2 patients, respectively. CR3 rate in patients who received treatment was overall 40% including response to HSCT of 58%. Conclusions Patients with relapsed AML have an overall probability of less than 50% to achieve a CR2 and CR3 after intensive salvage chemotherapy; the only exceptions are AML with CEBPAdm and CBF-AML. AlloHSCT either as direct treatment of relapse or as salvage therapy after failure of intensive chemotherapy may overcome chemo-resistance. Disclosures: Schlenk: Celgene: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Chugai: Research Funding; Amgen: Research Funding; Novartis: Research Funding; Ambit: Honoraria. Off Label Use: Pomalidomide in Myelofibrosis. Kindler:Novartis: Membership on an entity’s Board of Directors or advisory committees.

Abstract: Progress in the treatment of acute myeloid leukemia (AML) in older patients (pts) is still limited with poor complete remission (CR) rate and overall survival (OS). This is attributed to a variety of reasons including an inherently poor biology, especially a higher incidence of poor-risk karyotypes, co-morbidities, and an age-related functional impairment. In our randomized AML HD98B trial, the addition of all-trans retinoic acid (ATRA) to intensive chemotherapy resulted in an increased CR rate, event-free (EFS) and OS (Schlenk et al Leukemia 2004). More recent reports on in vitro studies indicated a synergistic action of the histone deacetylase inhibitor valproic acid (VPA) when associated with ATRA plus cytarabine and anthracyclines. In the randomized AMLSG 06-04 trial, therefore, VPA was evaluated in combination with intensive induction therapy plus ATRA in older pts ( 〉 60 years) with newly diagnosed AML. In first analyses, the addition of VPA did not provide a significant advantage in OS and EFS after a median follow-up of 47 months (Tassara et al, ASH 2010, abstract #185). This was mainly due to increased hematological toxicity by VPA after the second induction therapy. Here we provide updated analyses especially on survival outcome data based on mature follow-up. Aims To evaluate VPA in combination with intensive induction therapy and ATRA in older patients with newly diagnosed AML. Methods Between August 2004 and February 2006 186 patients were randomized (standard-arm, n=93; experimental-arm, n=93) in the AMLSG 06-04 study (ClinicalTrials.gov Identifier: NCT00151255); median age was 68 years (60-84). The first 77 pts were randomized to receive 2 induction cycles (idarubicin 12 mg/m2 i.v. days 1-3, cytarabine 100 mg/m2 cont. i.v. days 1-5, ATRA 45 mg/m2 days 3-5 and 15 mg/m2 days 6-28) with or without VPA (days 1-28; started at 400 mg bid and then adapted in order to obtain a serum level of 60-150 mg/l). After an interim analysis the study was amended; for the following 109 patients idarubicin was dose-reduced to day 1 and 3 and VPA only added during the first induction cycle. All patients were intended for consolidation. Molecular diagnostics were performed as previously published (Schlenk et al, Haematologica 2009) Results Details of the response rate and toxicity of the induction treatment have already been presented (Tassara et al, ASH 2010, abstract #185). To summarize, CR rates after double induction were in trend higher in the standard-arm (52% vs. 40%; p=0.10), and early death rate higher in the experimental-arm (14% vs. 26%; p=0.06). The main toxicities attributed to VPA were grade 3/4 infections and delayed hematologic recovery (leukocytes, neutrophils and platelets) observed after the second induction cycle. Therapy (i.e. double induction and consolidation) was completed by 37/93 (40%) of patients in the standard arm and 19/93 (20%) in the experimental arm (p=0.01) After a median follow up of 84 months, analysis of the primary endpoint EFS revealed no differences between the two arms (EFS at 5 years, standard arm 2.3%, experimental arm 7.6%; p=0.95); similarly OS was not different (OS at 5 years, standard arm 11.7%, experimental arm 11.4%; p=0.57). However, pts in the experimental arm had a significantly better relapse-free survival (RFS at 5 years, standard arm 6.4%, experimental arm 24.0%, p=0.02). In explorative subset analyses superior RFS (p=0.03) and OS (p=0.03) of CR-patients were observed in AML patients with mutated NPM1 randomized into the experimental arm (RFS at 5 years, standard arm 8%, experimental arm 42%; OS at 5 years, standard arm 37%, experimental arm 52%). In contrast no differences were seen in AML patients with NPM1 wild-type for RFS (p=0.13) and OS (p=0.87) of CR-patients (RFS at 5 years, standard arm 7%, experimental arm 20%; OS at 5 years, standard arm 15%, experimental arm 22%). Due to a low frequency of FLT3-ITD (9/72) in this patient subset meaningful analyses were not possible. Conclusion In older patients with AML, the addition of VPA to standard induction treatment was associated with severe hematological toxicity as well as higher rates of infections and did not improve EFS and OS. However, after a long follow-up VPA was associated with a significantly improved RFS, which might be related to the mutated NPM1 genotype. Disclosures: Schlenk: Celgene: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Chugai: Research Funding; Amgen: Research Funding; Novartis: Research Funding; Ambit: Honoraria.

Abstract: Based on their association with certain biological and clinical features as well as their prognostic significance, mutations in the CCAAT/enhancer-binding protein-alpha (CEBPA) gene have been included as a provisional entity into the 2008 World Health Organization (WHO) classification of myeloid neoplasms. CEBPA mutations (CEBPAmut) are mainly found in acute myeloid leukemia (AML) with normal cytogenetics, and approximately 60% of the mutated patients (pts) carry biallelic mutations. Several studies showed that in particular pts with double mutant CEBPA (CEBPAdm) have a favorable outcome compared to all others. Recently, mutations in the transcription factor GATA2 were identified as genetic lesions potentially cooperating with CEBPAdm. Both, CEBPA and GATA2 are involved in the control of proliferation and differentiation of myeloid progenitors, and mutations in both genes are discussed as pre-disposing events in myeloid leukemia. Based on functional studies there is an important interplay between the two genes, e.g. through the formation of direct protein complexes. Finally, preliminary data suggest that the genotype CEBPAdm/GATA2 mutated (GATA2mut) is associated with a favorable outcome in AML pts. Aims To evaluate the frequency and the clinical impact of GATA2mut within a large cohort of CEBPAmut AML pts and to further analyze the CEBPAmut/GATA2mutgenotype within the context of other genetic alterations. Methods In total 202 AML pts (age 18 to 78 years) with CEBPA single mutations (n=89) or CEBPAdm (n=113) were analyzed for the presence of GATA2mut. All pts were enrolled on one of 6 AMLSG treatment trials applying intensive therapy [AMLHD93 n=15; AMLHD98A (NCT00146120) n=53; AMLHD98B n=13; AMLSG 07-04 (NCT00151242) n=74; AMLSG 06-04 (NCT00151255) n=25 and AMLSG 12-09 (NCT01180322) n=22]. GATA2 mutation screening was performed using a DNA-based PCR-assay covering exons 2 to 6 followed by Sanger sequencing. Results GATA2 mut were restricted to the cytogenetic intermediate-risk group; in total we detected 42 GATA2mut in 40 of the 202 pts (20.7%); 36 pts had CEBPAdm (36/113, 31.8%), 4 were CEBPA single mutated (4/89, 4.4%). All mutations were heterozygous, with 2 pts having two mutations (in exon 4 and 5, respectively). 31 (73.8%) of the 42 mutations were located in zinc-finger 1 (ZF1, exon 4) and 11 (26.1%) in ZF2 (exon 5). GATA2 sequence alterations included 39 missense and 3 frameshift mutations. The median follow-up of the 202 pts was 64.2 months (95%-CI: 60.1 – 75.1). First, we evaluated the clinical impact of GATA2mut in the whole cohort. Here, we found no differences in overall (OS), event-free (EFS), and relapse-free (RFS) survival as well as for the cumulative incidence of relapse (CIR) between GATA2mut and GATA2 wildtype pts. Next, the effects of GATA2mut in CEBPAdm pts (n=113) were analyzed without seeing any differences for the clinical endpoints OS, EFS, RFS and CIR. The same was also true when we investigated the impact of GATA2mut with respect to their location in the ZF domains; there were no differences between pts with ZF1 (n=29) and ZF2 (n=9) mutations, respectively. Finally, we evaluated the possible relevance of GATA2mut in the subgroup of CEBPAdm pts 〈 60 years with intermediate-risk cytogenetics (n=94); but again GATA2mut did not impact the endpoints OS, EFS, RFS and CIR. In contrast to recently published data, we also detected GATA2mut in a small number of pts with CEBPA single mutations (n=4); however the low pt number did not allow a meaningful analysis. In addition, in our study GATA2mut occurred in rare cases with NPM1mut, FLT3-ITD or FLT3-TKD mutations. Conclusions In our study on a large cohort of CEBPA mutated AML pts we could confirm the high coincidence of GATA2 mutations, in particular in the subgroup of pts with CEBPA double mutations. However, GATA2 mutations had no impact on clinical outcome neither in the whole cohort nor in distinct pt subgroups. Disclosures: Schlegelberger: Celgene: Consultancy. Germing:Celgene: Honoraria, Research Funding. Kindler:Novartis: Membership on an entity’s Board of Directors or advisory committees. Schlenk:Novartis: Research Funding; Amgen: Research Funding; Chugai: Research Funding; Pfizer: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Ambit: Honoraria.

Abstract: In a recent update on MRD monitoring in 407 NPM1 mutated (NPM1mut) AML patients (pts) we could confirm the results from our previous study showing that achievement of RQ-PCR negativity after double induction (DI), after completion of therapy (CT) as well as during the follow-up period (FUP) is significantly associated with a lower cumulative incidence of relapse (CIR) and superior overall survival (OS) [Döhner K, Annals of Hematol; 2013;Suppl.1,92:S39] . In addition, in pts with concurrent FLT3-ITD (FLT3-ITDmut) or DNMT3A (DNMT3Amut) mutations, we also showed that the median NPM1mut transcript levels after each treatment cycle were significantly higher. Aim To evaluate the impact of concurrent FLT3-ITD and DNMT3Amut on MRD kinetics and clinical outcome in NPM1mutAML pts. Methods For this analysis we included all pts enrolled on one of two AMLSG treatment trials [AMLHD98A (NCT00146120) n=46; AMLSG 07-04 (NCT00151242) n=199] for whom the FLT3-ITD and DNMT3A mutation status at the time of diagnosis was determined. MRD levels (ratio NPM1mut/ABL1 transcripts x 104) were detected by NPM1mut specific RQ-PCR using TaqMan technology; the sensitivity of the assays was 10-5 - 10-6. DNMT3A and FLT3-ITD mutation status was assessed by standard PCR-based methods Results In total, 1588 samples [bone marrow n=1564; peripheral blood n=24] from 245 NPM1mut pts were analyzed [at diagnosis, n= 240; during therapy, n= 807; during FUP, n= 541] . FLT3-ITD and DNMT3A mutation status was available in 245/245 (FLT3-ITDmut n=94) and in 234/245 (DNMT3Amut n= 122) pts, respectively. Pre-treatment NPM1mut transcript levels did not correlate with clinical characteristics, DNMT3A or FLT3-ITD mutation status and had no impact on event-free survival, relapse-free survival and OS. Multivariable analyses stratified for FLT3-ITD mutation status after DI and CT revealed RQ-PCR negativity as a significant factor for longer remission duration (hazard ratio (HR) 15.15 and 8.95, respectively) and better OS (HR 6.13 and 4.27, respectively); DNMT3A mutation status had no significant impact in these models. Subgroup analyses showed that the proportion of pts achieving RQ-PCR negativity after DI, after CT and during FUP was significantly lower in DNMT3Amut compared to the DNMT3A wildtype pts (8.6% vs 33.3%, p= 〈 0.001; 36,3% vs 61.9%, p=0.009; 33% vs 51%, p=0.04, respectively) whereas for FLT3-ITDmut pts this effect was only significant after DI (8.3% vs 25%, p=0.022). Based on these findings we further investigated the impact of RQ-PCR negativity in the context of concurrent FLT3-ITD and DNMT3A mutations. After DI, there was no significant difference in CIR and OS for RQ-PCR negative pts with respect to FLT3-ITD or DNMT3A mutation status. After CT, RQ-PCR negative pts with DNMT3Amut had a significantly higher CIR compared to DNMT3A wildtype pts (34% vs 8% at 4 years; p=0.007). This adverse prognostic impact was consistently seen during the FUP (CIR 21% vs 3% at 4 years; p=0.01); there was no difference in CIR rates between pts with and without FLT3-ITD mutations. Conclusions We demonstrate a significant correlation between the DNMT3A mutation status and the achievement of RQ-PCR negativity at all clinically relevant time points i.e. after DI, and CT, and during FUP while this strong correlation was not observed for FLT3-ITDmut. Within the NPM1mut RQ-PCR negative group the presence of DNMT3Amut allows the identification of pts at high risk of relapse. Based on our findings DNMT3A mutation status should be determined in NPM1mut pts to further refine MRD monitoring. The establishment of DNMT3Amut specific MRD assays might provide additional information on MRD status in these pts. Disclosures: Schlegelberger: Celgene: Consultancy. Lübbert:Johnson and Johnson: Advisory Board Other. Kindler:Novartis: Membership on an entity’s Board of Directors or advisory committees. Germing:Celgene: Honoraria, Research Funding. Schlenk:Novartis: Research Funding; Amgen: Research Funding; Pfizer: Honoraria, Research Funding; Chugai: Research Funding; Ambit: Honoraria; Celgene: Honoraria, Research Funding.

Abstract: To evaluate the prognostic value of minimal residual disease (MRD) in patients with acute myeloid leukemia (AML) with NPM1 mutation (NPM1 mut ). Patients and Method RNA-based real-time quantitative polymerase chain reaction (RQ-PCR) specific for the detection of six different NPM1 mut types was applied to 1,682 samples (bone marrow, n = 1,272; blood, n = 410) serially obtained from 245 intensively treated younger adult patients who were 16 to 60 years old. Results NPM1 mut transcript levels as a continuous variable were significantly associated with prognosis after each treatment cycle. Achievement of RQ-PCR negativity after double induction therapy identified patients with a low cumulative incidence of relapse (CIR; 6.5% after 4 years) compared with RQ-PCR–positive patients (53.0%; P 〈 .001); this translated into significant differences in overall survival (90% v 51%, respectively; P = .001). After completion of therapy, CIR was 15.7% in RQ-PCR–negative patients compared with 66.5% in RQ-PCR–positive patients (P 〈 .001). Multivariable analyses after double induction and after completion of consolidation therapy revealed higher NPM1 mut transcript levels as a significant factor for a higher risk of relapse and death. Serial post-treatment assessment of MRD allowed early detection of relapse in patients exceeding more than 200 NPM1 mut /10 4 ABL copies. Conclusion We defined clinically relevant time points for NPM1 mut MRD assessment that allow for the identification of patients with AML who are at high risk of relapse. Monitoring of NPM1 mut transcript levels should be incorporated in future clinical trials to guide therapeutic decisions.