Abstract: Background CEBPA mutations are found in approximately 10% of acute myeloid leukemia (AML). Recent studies revealed an association between CEBPA and GATA2 mutations. GATA2 gene encodes a transcription factor involved in hematopoiesis. In two recent studies (Grossmann et al., BJH 2013; Fasan et al., Leukemia 2013), GATA2 mutations appeared to be associated specifically with double CEBPA mutations and improved overall survival (OS). In contrast, another study failed to show any impact on OS (Green et al., BJH 2013). The aim of our study was to investigate the prognostic significance of GATA2 mutations in a large cohort of patients with CEBPA-mutated AML. Patients and methods We studied a cohort of 128 patients with CEBPA-mutated AML treated with intensive chemotherapy. The entire coding region of CEBPA, NPM1 exon 12, and GATA2 exons 4 to 6 (that encode the two multifunctional zinc finger domains of the protein) were screened on genomic DNA by PCR and direct Sanger sequencing. FLT3 internal tandem duplications (FLT3-ITD) were screened on genomic DNA by PCR and fragment analysis. Additionally, transcriptome analysis was performed with Affymetrix HG U133 Plus 2.0 array in pre-treatment samples from 72 patients for which RNA was available. Results Median age at AML diagnosis in the whole cohort was 50 years (range, 3-80). Almost all patients belonged to the intermediate cytogenetic risk-group (n=117), of which 90 had a normal karyotype. The remaining patients had favorable (n=1) or adverse cytogenetics (n=4). CEBPA mutations were distributed as follows: 29 single-mutated (sm) cases with N-terminal (N-ter) mutations, 12 sm cases with C-terminal (C-ter) mutations, 80 double-mutated (dm) cases with both N-ter and C-ter mutations, 2 cases with homozygous N-ter and 5 cases with homozygous C-ter mutations. GATA2 mutations were found in 29/128 patients (23%) and were significantly associated with CEBPA dm cases (4/41 sm vs 25/87 dm: Fisher's exact test p=0.022). NPM1 mutations were detected in 12 patients. They were specifically associated with CEBPA sm cases (12/40 sm vs 0/78 dm, p 〈 0.001) and mutually exclusive with GATA2 mutations (12/93 GATA2 wild-type vs 0/25 GATA2 mutated cases, p 〈 0.001). In contrast with previous studies, FLT3-ITD and GATA2 mutations did co-occur in our cohort, with 3 GATA2 mutants identified in 10 FLT3-ITD positive patients. Transcriptome analysis revealed that GATA2 mutations were not associated with a specific gene expression signature. As previously described, we found that CEBPA dm AML harbored a specific gene expression profile distinct from CEBPA sm cases. Since AML with homozygous CEBPA mutations were found to have a similar gene expression signature as CEBPA dm AML, we decided to pool them together for prognostic analysis. Overall, complete remission was achieved in 113 patients (88%), of whom 36 relapsed (estimated cumulated incidence of relapse [CIR] at 3 and 5 years, 39%). Neither age nor karyotype or gene mutations (including NPM1, FLT3, GATA2 and type of CEBPA mutation) significantly influenced CIR in multivariate analysis. Five-year OS was estimated at 58% in the whole cohort, with longer OS in cases with normal karyotype (p=0.05) and double CEBPAmutations (sm vs dm, p=0.03). In multivariate analysis, younger age (p=0.020) and normal karyotype (p=0.029) remained the only factors significantly associated with a longer OS. Conclusions This study confirmed the strong association between GATA2 mutations and CEBPA double mutations, in line with previous studies. However, we did not find any prognostic impact of GATA2 mutations in our cohort of CEBPA-mutated AML. Disclosures No relevant conflicts of interest to declare.

Abstract: Acute myeloid leukemia (AML) with t(8;21) chromosomal translocation, leading to the RUNX1-RUNX1T1 fusion, belong to the favorable risk AML subset. However, relapse incidence may reach 30-40% in these patients. Minimal residual disease monitoring (MRD) based on the quantification of RUNX1-RUNX1T1 fusion transcript by real-time quantitative PCR (RQ-PCR) has been reported to be an independent prognostic factor for the risk of relapse. The specificity of the RUNX1-RUNX1T1 fusion and the high sensitivity of RQ-PCR techniques have made RUNX1-RUNX1T1 an ideal marker to assess treatment response in t(8;21) AML. Undetectable MRD could mean either that tumor cells persist in a latent state without RNA expression or that MRD level is below the sensitivity threshold. Studies in chronic myeloid leukemia showed that BCR-ABL DNA was still detectable in patients in long-term complete molecular response with undetectable BCR-ABL fusion transcript. Using a similar approach, we investigated the use of RUNX1-RUNX1T1 DNA as a MRD marker in t(8;21) AML, instead of RUNX1-RUNX1T1 mRNA. This approach allows linking results directly to the amount of leukemic cells, since each leukemic cell contains one copy of the RUNX1-RUNX1T1 sequence, while the level of RUNX1-RUNX1T1 mRNA may vary from a patient to another. Methods This study focuses on 17 patients with t(8;21) AML included in the CBF-2006 trial and for whom frozen material was available for further molecular analysis. Bone marrow and blood samples were collected at AML diagnosis and during follow-up, as defined in the CBF-2006 trial. Eight patients relapsed during follow-up and 9 were still in complete remission at the end of the study. Interestingly, 3 patients relapsed with a previously undetectable MRD (in blood and bone marrow samples). First, we identified the breakpoints in the RUNX1 and RUNX1T1 genes for each patient using long-range PCR approaches, coupled with next-generation sequencing (NGS) on Personal Genome Machine™ (PGM). The stability of the RUNX1-RUNX1T1 rearrangement at relapse was checked by Sanger sequencing. Then, we performed quantification of RUNX1-RUNX1T1 DNA by RQ-PCR using Taqman technology. For each patient, a primer pair and a probe were designed using the patient's unique RUNX1-RUNX1T1 breakpoint sequence. The forward and reverse primers were located in RUNX1 and RUNX1T1 genes, respectively, and the probe was located at the RUNX1-RUNX1T1 junction. Calibration curves were established using 10-fold dilutions of the diagnostic DNA of each patient in normal control DNA. Results were given as a ratio of chimeric DNA amount in the follow-up sample to chimeric DNA amount at diagnosis. Results Chromosomal breakpoints were located in RUNX1 intron 5 for all patients. RUNX1T1 breakpoints were located in intron 1b for 15 patients, and in intron 1a for 2 patients (Fig. 1). Quantification failed for 1 patient which was further leave up. Between 2 and 7 follow-up samples were studied for the other patients (median 4.5). DNA monitoring was strongly correlated with RNA monitoring (Fig. 2). Sensitivity threshold, determined by the lowest diagnostic sample dilution which gives a signal, was 10-5 for 7 patients, 10-4 for 6 patients, and only 5.10-4 for 3 patients. MRD was detectable in 31 samples and undetectable in 30 samples by both methods, whereas MRD was detectable only on RNA in 7 samples, probably because of a lack of sensitivity of the RQ-PCR assay. Interestingly, RUNX1T1-RUNX1 DNA was detected in 3 samples from 2 patients who relapsed and for whom RUNX1T1-RUNX1 transcript was undetectable, despite a good RNA quality. Conclusions Overall, RUNX1-RUNX1T1 MRD levels on DNA and RNA were quite similar. The level of mRNA expression did not seem to change during follow-up when compared with the amount of DNA. MRD monitoring on genomic DNA is a useful method, but with sensitivity variations depending on the patient's breakpoint sequence and the efficiency of the RQ-PCR assay. DNA has potential advantages: it is more stable than RNA and a best substrate for collection, processing, transport and storage. Additionally, interpretation of the results is easier because it is closely related to the number of leukemic cells. However, this method greatly increases complexity, time of implementation, and cost of monitoring MRD, which limits its interest in routine practice. Disclosures: No relevant conflicts of interest to declare.

Abstract: Context. The prognostic value of gene mutations in older AML patients (pts) treated intensively remains unclear. Only one study has explored the role of mutation patterns determined by NGS in older AML pts prospectively treated with various chemotherapies in years 2000-2010 (Eisfeld Leukemia 2018). Methods. Pts older than 60y enrolled in the ALFA-1200 trial (NCT01966497) between 09/2012 and 06/2016 were sequenced with a 37-gene myeloid panel. Pts received one 7+3 course followed by 2 intermediate-dose cytarabine courses. Pts with non-favorable risk were eligible for allogeneic stem cell transplantation (SCT). Variable selection for multivariate analyses was performed by lasso penalized regression including age, gender and log(WBC) as covariates. Results. Sequencing was done in 471 (93%) of the 509 enrolled pts. Median age and WBC count were 68y and 5.3x109/L, respectively (resp). CR (including CRp) was achieved in 341 (72%) pts and 90 underwent RIC-SCT in first CR. With a median follow-up of 25.4 months, median OS was 20.7 months. Pts had a median of 3 mutations (range 1-10). The 17 mostly frequently mutated genes (≥5% of pts, by decreasing frequency: DNMT3A, NPM1, TET2, ASXL1, FLT3, SRSF2, IDH2, RUNX1, NRAS, IDH1, STAG2, BCOR, TP53, PTPN11, U2AF1, EZH2 and KRAS) were retained for prognostic analyses. Genes belonging to a common pathway (eg. NRAS and KRAS) may have divergent prognostic values, preventing biology-informed grouping of mutations. Cytogenetic risk (derived from ELN 2017, Döhner Blood 2017, not considering gene mutations) was favorable (fav), intermediate (int), adverse (adv) and missing in 3%, 72%, 18% and 7% resp. Because of the few pts with fav cytogenetics in our cohort, pts were further grouped into non-adv and adv cytogenetics. CR rates and median OS were 75.6% vs 56.6% and 24.8 vs 9.5 months in pts with non-adv and adv cytogenetics, resp (both p 〈 0.0001). Because of difference in mutational patterns and gene-gene interactions, the prognostic role of mutations was considered independently in these two non-adv and adv subgroups. In the 388 pts with non-adv cytogenetics, NPM1 mutations independently predicted improved CR rate (Odds Ratio [OR]=2.3, p=0.014), while mutations in ASXL1 (OR=0.46, p=0.012), RUNX1 (OR=0.46, p=0.013) and NRAS (OR=0.49, p=0.04) had independent adverse predictive value. In univariate analysis the shorter OS of FLT3-ITD pts was confined to allele ratios≥ 0.5 (FLT3-ITDhigh, p=0.02). In a multivariate analysis accounting for clinical covariates, mutations in NPM1 (Hazard Ratio [HR] =0.45, p 〈 0.0001) and in SRSF2 (HR=0.64, p=0.03) predicted improved outcome, while FLT3-ITDhigh (HR=2.00, p=0.03), mutations in DNMT3A (HR=1.74, p=0.001), ASXL1 (HR=1.84, p=0.002) and NRAS (HR=1.70, p=0.009), but not RUNX1 or TP53, independently predicted worse OS. Significant interactions (eg. NPM1 - SRSF2, p=0.009, NPM1 - DNMT3A, p=0.03) precluded a simple NPM1-based stratification of pts with non-adv cytogenetics. This led to define a new prognostic hierarchy (Figure). The 49 NPM1mut pts with SRSF2 mutation and/or without adverse co-mutations (FLT3-ITDhighDNMT3A, ASXL1 and NRAS) had a median OS of 49.7 months, defining very low risk. NPM1wt pts without adverse co-mutations (n=114) had a median OS of 30.7 months and were considered at low risk. Among pts with ≥1 adverse co-mutation, NPM1 status had no significant prognostic influence (p=0.18). Regardless of NPM1 status, pts with a single (n=187) or ≥2 (n=38) adverse co-mutations (FLT3-ITDhighDNMT3A, ASXL1 or NRAS) had a median OS of 21.0 and 12.0 months, resp, and were considered at intermediate and high risk, resp. In the 83 pts with adv cytogenetics, TP53 mutations predicted shorter OS (p=0.004). Among pts with adv cytogenetics, those without TP53 mutation had a median OS of 12.6 months and were thus classified as high risk while the median OS of the 30 pts with adv cytogenetics and TP53 mutations was only 5.4 months, defining very high risk disease. This stratification resulted in improved OS prediction compared to the full molecular ELN 2017 (C-index 0.63 vs 0.58, resp). This stratification also predicted Relapse-Free Survival (RFS, Figure, p 〈 0.0001). Censoring at SCT did not affect these results. Conclusion. In AML patients older than 60y treated intensively, mutations in 7 genes (NPM1, SRSF2, FLT3, DNMT3A, ASLX1, NRAS and TP53) can refine the prognosis of cytogenetic sub-groups. Figure Figure. Disclosures Cluzeau: MENARINI: Consultancy; CELGENE: Consultancy; JAZZ PHARMA: Consultancy.

Abstract: Background : Point mutations in isocitrate dehydrogenase (IDH) genes are seen in 20% of adult patients (pts) with acute myeloid leukemia (AML). Prognostic significance of each IDH1/2 mutation (mut) analyzed with co-occurring mutations treated with intensive chemotherapy (IC) remains inconsistent, particularly with the advent of IDH inhibitors. Furthermore, the role of allogeneic stem cell transplantation (SCT) in IDH-mutated without favorable-risk features is not known. Patients & Methods: Between 2009 and 2016, 262 pts with IDH1/2 mutated AML (101 IDH1mut, 115 IDH2R140Qmutand 46 IDH2R172mut) were treated with IC in younger ALFA-0702 (NCT00932412, n = 133) and older ALFA-1200 (NCT01966497, n = 129) prospective trials. Median age was 50 [42-54] and 67 y [64-71] , respectively (resp). Targeted 37-gene next-generation sequencing (NGS) information was available for all pts. According to ELN 2010 classification, non-favorable CR/CRp pts were eligible for SCT if they had a sibling or matched unrelated donor. Correlation between IDHmutand covariates was realized by Pearson correlation coefficient and point biserial correlation for continuous and dichotomic variables, resp. Impact on response and survival was assessed for all covariates present in at least 10% of patients. SCT was considered as a time-dependent variable. Informative variables selected by LASSO were included in multivariate logistic regression for response and multivariate Cox model for survival. All analyses were stratified on the clinical trial. Results: IDH1 mut was significantly associated with NPM1mut(p=0.025), DNMT3Amut(p=0.009) and mutually exclusive with TET2mut(p=0.009). 80% (81/101) of IDH1 mutated pts achieved CR/CRp [96 % (46/48) if concomitant NPM1mut vs 66% (35/53) if not (p=0.0009)]. With a median FU of 39 months, overall median OS was not reached and median EFS was 15 months (Fig 1A). Presence of NPM1mutwas the only variable associated with longer OS (HR=0.33, p=0.001) and EFS (HR = 0.4, p = 0.001) in multivariate analysis. At 5 years, OS was estimated at 68% and 35% and EFS at 55% and 26%, resp (Fig 1B). Effect of concomitant NPM1mutwas reinforced in the absence of DNMT3Amut(HR=0.14, p=0.0006 and HR=0.16, p & lt;0.0001, for OS and EFS resp). At 5 years, EFS was estimated at 70% in IDH1+/NPM1+/DNMT3A- AML pts vs 30% for other IDH1+ AML pts (Fig 1B). IDH2R140Q mut was significantly associated with NPM1mut(p=0.0004) and SRSF2mut(p & lt;0.0001) and normal karyotype (p=0.002), but negatively correlated with complex karyotype (p=0.01). 91% (105/125) pts with IDH2R140Q mutated AML achieved CR/CRp [100 % (58/58) if concomitant NPM1mutvs 82% (47/57) if not (NS)]. With a median FU of 40 months, overall median OS was not reached and median EFS was 25 months (Fig 1A). Again, the presence of NPM1mutwas associated with a longer OS and EFS (HR=0.47, p=0.02 and HR= 0.24, p & lt;0.0001, resp). Presence of DNMT3Amutwas associated with shorter OS (HR = 2.1, p=0.02) and EFS (HR = 2, p=0.008) along with high WBC (HR = 1.9, p=0.005) for decreased EFS. At 5 years, OS was estimated at 67% in IDH2R140Q+/NPM1+ AML pts vs 40% in those with IDH2R140Q+/NPM1- AML. At 5 years, EFS was estimated at 56% vs 29% in these two subgroups, resp (Fig 1B). Again, the effect of concomitant NPM1mutwas reinforced in the absence of DNMT3Amut(HR = 0.26, p=0.0009 and HR = 0.15, p & lt;0.0001, for OS and EFS resp). At 5 years, EFS was estimated at 72% in IDH2R140Q+/NPM1+/DNMT3A- AML pts vs 29% in other IDH2R140Q+ AML pts (Fig 1C). IDH2R172K mut was significantly associated with DNMT3Amut(p=0.0004) and BCORmut(p & lt;0.001), as well as +11 (p=0.002), but negatively correlated with NPM1mut(p=0.001). 78% (36/46) pts with IDH2R172Kmutachieved CR/CRp. No genetic alteration was associated with outcome, perhaps due to limited number of pts. With a median FU of 43 months, overall median OS and EFS were 60 and 14 months, resp (Fig 1A). Finally, in non-favorable ELN 2010 pts (74, 74 and 46 with IDH1mut, IDH2R140Qmutand IDH2R172Kmut, resp), SCT in first CR only benefited to pts with IDH1mut(p=0.004 for OS) or with IDH2R172Kmut(p=0.03 for EFS). Conclusion: In a large prospective series, NPM1mutis the main better risk factor in the IDH1mutand IDH2R140Qmutsubgroups and may be used as stratification factor in clinical trials testing frontline specific IDH inhibitors with IC. Allogeneic SCT in first CR appears to improve the outcome of pts with non-favorable IDH1 or IDH2R172K mutated AML. Figure 1 Disclosures Micol: Jazz Pharmaceuticals: Consultancy; AbbVie: Consultancy. Thomas:ABBVIE: Honoraria; DAICHI: Honoraria; INCYTE: Honoraria; PFIZER: Honoraria. Braun:Institut de Recherches Internationales Servier (IRIS): Research Funding. Ades:Novartis: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Agios: Membership on an entity's Board of Directors or advisory committees; Jazz: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Helsinn Healthcare: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Silence Therapeutics: Membership on an entity's Board of Directors or advisory committees. Berthon:PFIZER: Other: DISCLOSURE BOARD; JAZZPHARMACEUTICAL: Other: DISCLOSURE BOARD; CELGEN: Other: DISCLOSURE BOARD. Boissel:NOVARTIS: Consultancy. Vey:Janssen: Honoraria; Novartis: Consultancy, Honoraria. Pigneux:Astellas: Honoraria; Amgen: Honoraria; Novartis: Honoraria; Jazz: Honoraria; Abbvie: Honoraria; Roche: Honoraria; Pfizer: Honoraria; Daichi: Honoraria; F. Hoffmann-La Roche Ltd: Honoraria. Recher:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Incyte: Honoraria; Jazz: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Abbvie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Sunesis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Macrogenics: Consultancy, Membership on an entity's Board of Directors or advisory committees. Dombret:CELGENE: Consultancy, Honoraria; AGIOS: Honoraria; Institut de Recherches Internationales Servier (IRIS): Research Funding. De Botton:Daiichi: Consultancy; Janssen: Consultancy; Agios: Consultancy, Research Funding; Astellas: Consultancy; Pierre Fabre: Consultancy; Servier: Consultancy; Pfizer: Consultancy; Novartis: Consultancy; Forma: Consultancy, Research Funding; Syros: Consultancy; AbbVie: Consultancy; Celgene Corporation: Consultancy, Speakers Bureau; Bayer: Consultancy.

Abstract: Acute myeloid leukemia (AML) is a heterogeneous group of malignancies sensitive to natural killer (NK) cell anti-tumor responses. However, NK cells are frequently defective in AML. Here, we found in an exploratory cohort (n = 46) that a different expression of activating receptors (NCR) on NK-cells identified a subgroup of patients with reduced expression of DNAM-1, NKG2D, NCRs and perforin especially in CD56bright NK-cells. Patients with a deficient NK-cell profile had a significantly higher risk of relapse (P = 0.03) independently of cytogenetic classification in multivariate analysis. We hypothesized that the transcription patterns in AML cells could give a clue on the NK/AML crosstalk. Analysis of the gene expression in AML blasts following classification according to their NK-cell status showed extensive differences in immune-related and oncogenic pathways. A favorable NK status was associated with pro-inflammatory AML transcription patterns whereas patients with defective NK cells showed a profound gene expression decrease in AML blasts for cytokine and chemokine signaling, antigen processing and adhesion molecule pathways. A set of 388 leukemic classifier genes defined in the exploratory cohort was independently validated in a multicentric cohort of 194 AML patients (ALFA-0701). In total, these data evidenced the prognostic value of NK-cell responses at diagnosis of AML and the relationships between NK-cells and AML independently of known cytogenetic status.

Abstract: Introduction: In a recent French FRALLE versus LALA comparison, we have demonstrated a large benefit in outcome when adolescents and young adults were treated in a pediatric rather than an adult ALL protocol (Boissel JCO 2003). Similar provocative results have been observed in three other pediatric versus adult ALL studies (Stock ASH 2000, de Bont Leukemia 2004, Testi ASH 2004). One explanation may be the larger amounts of steroids, vincristine (VCR), and L-asparaginase (L-aspa) administered in pediatric patients. The GRAALL-2003 study was thus designed to offer a pediatric approach in adults with Ph-negative ALL until 60 years of age. Methods: Treatment included a 5-drug induction, high dose-intensity consolidation blocks, delayed intensification, and 2-year maintenance. The comparison with the former LALA-94 protocol showed a 8.6-fold, 3.7-fold, and 16-fold increase in cumulative doses of prednisone, VCR, and L-aspa, respectively. One difference with childhood ALL therapy remained indication of allogeneic SCT in first CR which was offered to all patients with high-risk factor and a donor. In addition, induction was reinforced with a hyper-cyclophosphamide sequence (HyperC) in case of poor early response (cortico- and/or chemoresistant ALL). In the present report, 212 GRAALL-2003 patients with Philadelphia-negative ALL aged 15–55 years with a median follow-up of 18 months were compared to 712 patients previously treated in the LALA-94 trial. Results: Cohorts were comparable in terms of prognostic factors. CR rate was significantly higher in GRAALL patients (93 vs 88%, P=0.02) due to a reduction in resistant disease (0.5 vs 8%, P 〈 0.001) with comparable induction death (6 vs 5%, P=0.37) rate. When compared to the LALA-94 study, 2-year EFS and overall survival were markedly improved (56 vs 41% and 66 vs 54%; P=0.0002 and 0.02, respectively). Similar differences were observed after censoring SCT patients at transplant time, underlining the role played by chemotherapy modification. Because of a worse tolerance to induction and post-remission therapy, the benefit associated with the GRAALL approach decreased with advanced age, becoming not statistically significant over 40 years of age. Finally, outcome improvement was higher in T- than in B-lineage ALL (2-year overall survival, 75 vs 51% and 61 vs 56%; P=0.016 and 0.25, respectively). Among patients with T-ALL, poor early responders (allocated to HyperC induction) tended to have a better EFS than good early responders, suggesting a major role of HyperC in this ALL subset. Conclusion: In conclusion, a pediatric approach benefits to younger adults with ALL.

Abstract: Abstract 2125 Background: Over the last three decades, progress in the treatment of childhood acute lymphoblastic leukemia (ALL) has considerably improved the outcome of children, leading to 5-year OS of more than 80%. Numerous comparisons, including the French LALA/FRALLE (Boissel et al. JCO 2001), have reported a better outcome in teenagers treated with pediatric as compared to standard historical adult ALL protocols. Even if modern pediatric-inspired adult ALL protocols have recently reported impressive improvements, especially in younger patients (Huguet et al. JCO 2009), the issue of whether younger adults (YAs) should be treated according to pediatric or adult protocols remains an open one. The aim of this study was first to evaluate the feasibility and the results of a non-modified pediatric protocol (the French FRALLE 2000) in adolescents and younger adults (AYAs, aged 15–29 years) treated in adult departments. Methods: From February 2001 to June 2010, 72 AYAs with Ph-negative ALL were treated according to the pediatric FRALLE 2000-BT protocol in 12 adult hematology units in France and Belgium. After a prednisone prephase and a four-drug induction (prednisone, daunorubicin, vincristine and L-asparaginase), patients in CR received a consolidation, a 1st delayed intensification, an interphase, a 2nd delayed intensification, and a maintenance chemotherapy during two years. Results: The median age was 19 years (range, 15–29 years). The cohort was separated in 2 subgroups: 44 adolescents aged 15–19 years and 28 young adults (YAs) aged 20–29 years. There were no significant differences between the adolescent and the YA populations in term of sex ratio, white blood cell count (WBC), central nervous system involvement, and phenotype (BCP- vs T-ALL). As expected, few recurrent cytogenetical abnormalities were identified in this population and did not differed between both subgroups. In the adolescent group, we identified 2 patients with t(4;11), 1 patient with t(1;19), and 3 patients with hypodiploïdy and/or neartriploïdy, whereas this repartition was 2/2/1 in YAs. Rates of good early response to prednisone were in 68% in adolescents and 61% in YAs (p=.52), while rates of good early response to chemotherapy were 80% and 86%, respectively (p=.51). No patient died during induction. Complete remission (CR) rate did not differ between subgroups (98% vs 100%, p=.42). With a median follow up of 4.8 years, 5-year EFS was 57% (41% in adolescents vs 79% in YAs, p=.03) and 5-year OS was 67% (56% and 82% respectively, p=.09). In patients with BCP-ALL, 5-year EFS was 60% (43% in adolescents vs 91% in YAs, p=.02) and 55% in T-ALL (57% vs 50% respectively, p=.81). Twelve patients (17%) received an allogeneic stem cell transplantation (SCT) in first CR (5 adolescents and 7 YAs). Four patients died in first CR, all after SCT, (2 adolescents and 2 YAs). In univariate analysis, a high WBC (continuous variable, p=.02) and a poor early response to chemotherapy (33% vs 63%, p=.02), but not phenotype or poor early response to prednisone, were significantly associated with a shorter EFS. In multivariate analysis, age (adolescents vs YA, p=.04), WBC (continuous variable, p=.0005), and poor early response to chemotherapy (p=.006) had still an impact on EFS. The poor outcome of adolescents compared to YAs, also observed in the French adult GRAALL protocol (not published), was not explained by differences in ALL characteristics, early response to therapy, or treatment-related toxicity. Conclusion: The pediatric protocol FRALLE 2000 is effective and safe for the treatment of selected AYAs with Ph-negative ALL referred to adult departments. The results observed in the YA population are promising, warranting prospective comparisons with the more recent pediatric-inspired adult protocols. The unexpected poorer outcome of adolescents deserves further investigations to explore a potential impact of the quality of care delivered in an adult environment. Disclosures: No relevant conflicts of interest to declare.

Abstract: Introduction: In core binding factor (CBF) acute myeloid leukemia (AML), the disruption of CBFa (AML1) or CBFb genes through t(8;21) or inv(16) impairs normal hematopoietic differentiation, but is considered to be insufficient to induce leukemogenesis. The need of an additionnal oncogenic event that promotes cellular proliferation has been suggested. This proliferative signal may be delivered by mutated receptor tyrosine kinases (RTKs) or RAS proto-oncogenes. In a retrospective study, we evaluate the incidence and the prognosis of RTKs (FLT3/D835, KIT/D816 and KIT/Ex8 insertion/deletion) and N/K-RAS mutations in patients with CBF AML. Patients: Genomic DNA were obtained from the bone marrow or blood of 81 patients under 60 years with CBF AML (t(8;21)/inv(16) : 48/33) : 21 treated in the pediatric LAME trial (27/6) and 60 in the adult ALFA trial (33/15) between 1990 and 2004. The median follow-up is 5 years. Results: RTKs mutations are found in 15/74 (20%) patients. KIT mutations are present in 11/74 (15%) patients, at a lower rate than previously described. KIT/D816 mutations are observed in 2/49 (4%) t(8;21) patients and 1/32 (3%) inv(16) patients. KIT/Ex8 mutations are slightly more present in inv(16) than in t(8;21) patients (5/32 vs 3/42, 16% vs 7%). One of the 5/81 patients (6%) with FLT3/D835 mutations also presents a KIT/Ex8 mutation. RAS mutations rates are significantly higher in inv(16) than in t(8;21) patients (11/33 vs 4/38, 33% vs 11%), with mostly N-RAS mutations (75%). No correlation between RTKs or RAS mutations and age or WBC is observed. Four patients did not achieve complete remission (CR rate : 95%), 2 with FLT3/D835 and 3 with RAS mutations. In univariate analysis, RTKs mutations are significantly associated with a shorter OS and EFS (p=.001 and p=.02, resp.) KIT mutations are associated with a shorter EFS and RFS (p=.002 and p=.003) in t(8;21) patients but do not affect the outcome of inv(16) patients in our study. Patients with FLT3/D835 mutations have significantly shorter OS, EFS and RFS (p 〈 .0001, p 〈 .0001 and p=.0007, resp.). The outcome of each subgroup of CBF AML is affected by FLT3/D835 status. RAS mutations have no impact on outcome. Multivariate analysis show a persistent significant prognostic impact of RTKs mutations on OS, RFS and EFS. Conclusion: Inv(16) and t(8;21) cytogenetics are classically associated with a low risk of relapse. Patients with CBF AML are no more considered by many teams for allogeneic stem cell transplantation, even with a related donor. However, screening for RTKs mutations may identify patients with a more adverse outcome and thus susceptible to benefit from intensified protocols or tyrosine kinase inhibitors.

Abstract: Introduction: Philadelphia chromosome (Ph)-negative acute lymphoblastic leukemia (ALL) with high-risk genetics and/or measurable residual disease (MRD) are at high-risk of disease recurrence. In the previous GRAALL-2005 study, we identified KMT2A rearrangements (KMT2A-r), IKZF1 intragenic deletion (IKZF1del) and post-induction (TP1, week 6) MRD ≥ 0.01% as independent factors to predict relapse in Ph-negative B-cell precursor (BCP) ALL (Beldjord K, Blood 2014). In the GRAALL-2014 trial, high-risk (HR) patients were thus defined by the presence of at least one of these three factors. Among them, only those with higher MRD levels defined as TP1-MRD ≥ 0.1% and/or week 12 (TP2) MRD ≥ 0.01% were considered at very high risk (VHR) and proposed allogeneic hematopoietic stem cell transplant (alloSCT) in first remission (Dhedin et al., Blood 2015). Since October 2018, all these patients were eligible to be included in the GRAALL-2014-QUEST phase 2 study to receive blinatumomab as part of consolidation and maintenance phases or as a bridge to transplant. Methods: From October 2018 to December 2020, 95 patients with high-risk Ph-negative BCP-ALL without central nervous system involvement at diagnosis and in continuous complete remission after induction and consolidation 1, were prospectively included to start blinatumomab at week 12. One patient was excluded because of T-ALL phenotype (with CD19 aberrant expression). Patients with alloSCT indication and a stem cell source received blinatumomab 28 microg/d administered by continuous intravenous infusion (cIV) until transplant. A minimum of 4 weeks blinatumomab was recommended before proceeding to transplantation. All other patients received 5 cycles of blinatumomab 28 microg/day cIV (for 28 days), during consolidation 2 and 3 and at months 1/3/5 of the maintenance phase respectively. The primary objective was disease-free survival (DFS). Secondary objectives included post-blinatumomab MRD response at TP3 (after consolidation 2 or before alloSCT), overall survival (OS), and safety. Early results are reported here. Results: Median age was 35 years old (range, 18-60). Median white blood cell count (WBC) at diagnosis was 12 G/L (range, 1-449). Oncogenetic analyses allowed classifying ALL as Ph-like (18%), KMT2A-r (17%), DUX4/ERGdel (13%), ZNF384-r (11%), low hypodiploidy/near triploidy (7%), B-other (26%) or unknown (9%). An IKZF1del was found in 37/93 (40%). A TP1-MRD ≥ 0.01% was found in 46/94 patients (49%). Final risk group was HR for 45 patients and VHR for 49 patients. Last pre-blinatumomab MRD was & lt;0.01% in 49/88 (56%) of evaluable patients. A total of 40 patients (42%) received an alloSCT. The median number of blinatumomab cycles received in patients not proceeding to alloSCT was 4 cycles (range, 1-5). Thirty-nine severe adverse events (SAEs) were reported: 1 CRS (grade 2), 8 neurotoxicities (1 grade 2, 3 grade 3, 3 grade 4, 1 grade 5), 19 infections, and 11 others. The only grade 5 SAE occurred after alloSCT (seizures). After blinatumomab, a complete MRD response (with at least 0.01% sensitivity) was achieved in 61/82 (74%) evaluable patients and in evaluable patients with pre-blinatumomab detectable MRD. MRD response to blinatumomab was lower in patients with high pre-blinatumomab MRD level, while not impacted by age, WBC, or oncogenic subgroup. With a median follow-up of 20 months, 18-month DFS and OS was 78.8% (95% CI [66.9-86.8]) and 92.1% (95% CI [83.2-96.4] ) respectively (Figure 1). Patients with VHR diseases had a worse DFS (68.8%, 95% CI [51.1-81.2]) as compared to other patients (90.6%, 95% CI [72.1-97.1] ); p=0.018). This difference of DFS was abrogated by censoring patients at transplant (VHR 88.1%, 95% CI [65.5-96.3] versus others 90.6%, 95% CI [72.1-97.1%], p=0.10). Other factors significantly associated with better DFS were DUX4/ERGdel subgroup, low pre-blinatumomab MRD, and complete MRD response after blinatumomab. Conclusion. In patients wih high-risk BCP-ALL, blinatumomab added to consolidation is safe and gives promising results. A comparison to similar patients treated in the same GRAALL-2014 study before October 2018 is planned with a longer follow-up. Figure 1 Figure 1. Disclosures Boissel: Novartis: Consultancy, Honoraria, Research Funding; Incyte: Honoraria; Amgen: Consultancy, Honoraria, Research Funding; SANOFI: Honoraria; Servier: Consultancy, Honoraria; Bristol-Myers Squibb: Honoraria, Research Funding; CELGENE: Honoraria; JAZZ Pharma: Honoraria, Research Funding; PFIZER: Consultancy, Honoraria. Huguet: Novartis: Other: Advisor; Jazz Pharmaceuticals: Other: Advisor; Celgene: Other: Advisor; BMS: Other: Advisor; Amgen: Other: Advisor; Pfizer: Other: Advisor. Rousselot: Incyte, Pfizer: Consultancy, Research Funding. Chalandon: Incyte, BMS, Pfizer, Abbie, MSD, Roche, Novartis, Amgen: Other: Advisory Board; Incyte: Speakers Bureau; Incyte, BMS, Pfizer, Abbie, MSD, Roche, Novartis, Gilead, Amgen, Jazz, Astra Zenec: Other: Travel EXpenses, Accomodation. Delabesse: Astellas: Consultancy; Novartis: Consultancy. Dombret: Abbvie: Honoraria; Amgen: Honoraria, Research Funding; Incyte: Honoraria, Research Funding; Jazz Pharmaceuticals: Honoraria, Research Funding; NOVARTIS: Research Funding; pfizer: Honoraria, Research Funding; servier: Research Funding; BMS-Celgene: Honoraria; Daiichi Sankyo: Honoraria. OffLabel Disclosure: Blinatumomab in frontline high-risk acute lymphoblastic leukemia