Adult Acute Myeloid Leukemia Cases Research Articles

Significance of targeting DNMT3A mutations in AML.

Acute myeloid leukemia (AML) is the most prevalent form of leukemia among adults, characterized by aggressive behavior and significant genetic diversity. Despite decades of reliance on conventional chemotherapy as the mainstay treatment, patients often struggle with achieving remission, experience rapid relapses, and have limited survival prospects. While intensified induction chemotherapy and allogeneic stem cell transplantation have enhanced patient outcomes, these benefits are largely confined to younger AML patients capable of tolerating intensive treatments. DNMT3A, a crucial enzyme responsible for establishing de novo DNA methylation, plays a pivotal role in maintaining the delicate balance between hematopoietic stem cell differentiation and self-renewal, thereby influencing gene expression programs through epigenetic regulation. DNMT3A mutations are the most frequently observed genetic abnormalities in AML, predominantly in older patients, occurring in approximately 20-30% of adult AML cases and over 30% of AML with a normal karyotype. Consequently, the molecular underpinnings and potential therapeutic targets of DNMT3A mutations in AML are currently being thoroughly investigated. This article provides a comprehensive summary and the latest insights into the structure and function of DNMT3A, examines the impact of DNMT3A mutations on the progression and prognosis of AML, and explores potential therapeutic approaches for AML patients harboring DNMT3A mutations.

A nomogram based on clinical features and molecular abnormalities for predicting the prognosis of patients with acute myeloid leukemia.

The high clinical and molecular heterogeneity of acute myeloid leukemia (AML) has led to an unsatisfactory clinical prognosis, thus we sought to incorporate both clinical features and molecular abnormalities to construct a new prognostic model. A database search of the Gene Expression Omnibus (GEO) revealed 238 cases of adult AML. The independent risk factors were assessed using both univariate and multivariate Cox regression, as well as least absolute shrinkage and selection operator (LASSO) regression. The predictive accuracy, discriminatory power and clinical applicability of the nomogram were determined by the consistency index (C-index), calibration curves and decision curve analysis (DCA). In addition, a single-centre cohort of 135 cases was used for external validation. Multivariate Cox regression analysis showed that the independent influences on overall survival (OS) were age, type of disease, DNMT3A, IDH2 and TP53 mutations. The area under the curve (AUC) values for the training set were 0.755, 0.745 and 0.757 at 1, 2 and 3 years respectively; the AUC for the validation set were 0.648, 0.648 and 0.654 at 1, 2 and 3 years; and the AUC for the northwest China set were 0.692, 0.724 and 0.689 at 1, 2 and 3 years. The calibration and DCA indicated good consistency and clinical utility of the nomogram. Finally, younger (age <60 years) and elderly (age ≥60 years) patients were each divided into two risk groups with significantly different survival rates. A nomogram consisting of five risk factors was developed for forecasting the prognosis of AML with guaranteed reliability.

Clinical Features and Monitoring of Germline CEBPA-Mutated Carriers

Background: Despite being classified as an independent entity in World Health Organization (WHO) classification in 2016, familial cases of adult acute myeloid leukemia (AML) with germline-mutated CCAAT/enhancer-binding protein-α (CEBPA) gene have been rarely reported due to insufficient knowledge of the clinical features of this entity. Germline CEBPA-mutated carriers are at extremely high risk of developing AML but have long been ignored until the onset of AML. We aimed to fully identify the clinical features of the familial cases and healthy carriers for this entity to call for a guideline for its management. Methods: Whole exome sequencing (WES) data were analyzed in four members from one hereditary AML Chinese family, including two AML patients, one obligated carrier, and one non-consanguineous member. The whole coding region of CEBPA was sequenced in 21 members in this family and 6 members in another AML family. Deep sequencing targeting 248 genes related to myeloid malignancies was performed on 2 germline CEBPA-mutated carriers. To fully estimate the incidences of AML for each germline CEBPA mutation, we performed a thorough literature search for multiple members carrying germline CEBPA mutations. Results: Here, we report two Chinese families with multiple AML cases carrying germline CEBPA mutations (c.247C&amp;gt;T; p.Gln83*). One family had 11 cases in four consecutive generations and 5 healthy carriers, and the other had 2 cases in two generations. Together with these two families, we collected clinical data from 57 AML patients in 22 families with germline CEBPA-mutation in total. 48 out of 57 AML were confirmed carrying germline CEBPA mutations. 58.3% (28/48) of AML harbored double CEBPA mutations. The first hit frequently occurred at the N-terminal of CEBP/α (22/28, 78.6%), resulting in an exclusive expression of p30 at the second translation starting site of CEBPA (hereafter, CEBPA p30). The second hit normally located at C-terminal of CEBP/α (hereafter, CEBPA others). Germline CEBPA p30-carriershave higher incidences of AML compared to those with germline CEBPA others (80.36% vs 42.86%, P=0.0003). The age at onset is much younger in patients carrying germline CEBPA p30 than those carrying CEBPA others (18 vs 38.5, P=0.0093). Among cases with detailed treatment information, 10/36 did not receive standard chemotherapies according to the NCCN guidelines for AML, and 16/36 received transplantation, nearly half (7/16) of which went through autologous or synergetic (monozygotic twin) transplantation and maintained endurable complete remission. 41.7% (15/36) of AML had at least one episode of relapse, and recurrent relapses occurred in 1/3 (5/15) of the AML patients. Intriguing, despite the high rates of relapse, only 3/15 of AML patients died of relapses. Most familial AML cases with germline CEBPA mutations had a favorable overall survival (OS). We found the OS in AML patients with germline CEBPA p30 is better than those with CEBPA others (&amp;gt;25 years vs. 11 years, P=0.0125). There are 27 healthy germline CEBPA mutated carriers aged 14 to 88, 20 of which were selected for further analysis due to complete age information, 80% (16/20) are still within the range of age at onset of familial AML with germline CEBPA mutations. We detected a pre-leukemia clone harboring a pathogenic IDH2 variant (Arg140Gln) in 1 germline CEBPA-mutated carrier, who is currently healthy. Conclusions: The high penetrance accompanied by early onset in AML with CEBPA p30 mutation implies the mutation as a strong genetic risk factor for AML . The intensity of chemotherapy should be tailored for familial AML with germline CEBPA mutations. Our study is the most extensive clinical data for familial AML cases with germline CEBPA mutations and is the first to reveal precise incidences of AML with certain germline CEBPA mutations. These data should aid in genetic counsel and the management of AML patients and healthy carriers with germline CEBPA mutations.

Biological and Therapeutic Implications of NPM1-Mutated AML Receiving Allogeneic Hematopoietic Cell Transplantation

Background: The nucleophosmin 1( NPM1) gene mutations are the most common gene alteration in acute myeloid leukemia (AML) patients, occurring in approximately 30%-35% of adult AML cases. However, molecular and response to therapy heterogeneity are existed in this distinct group. Response to therapy, differing patterns of co-mutation and the prognosis of NPM1-mutated AML patients receving allogeneic hematopoietic stem cell transplantation (allo-HSCT) treatment were unknown. Aims: To evaluate prognostic factors in the NPM1-mutated AML who underwent allo-HSCT treatment. Methods: A total of 115 consecutive NPM1-mutated AML patients who underwent allo-HSCT from 2016 to 2022 were enrolled in this study. The primary endpoint was the cumulative incidence of relapse (CIR). Results: The median patient age was 46 (IQR: 36-50) years. The most frequent co-mutations were FLT3-ITD(58.3%) and DNMT3A (40.0%), followed by IDH2(18.3%), NRAS (13.9%) and IDH1 (11.3%). The overall 3-year cumulative incidence of relapse (CIR) and leukemia-free survival (LFS) was 17.3% and 77.2%. The 3-year CIR of patients with FLT3-ITD was significantly higher than that without FLT3-ITD (24.3% vs. 7.0%; P=0.023). The 3-year CIR of patients with DTA mutations ( DNMT3A, TET2, and ASXL1) was significantly higher than that without DTA mutations (24.7% vs. 11.2%; P=0.030). Patients who received matched sibling donor (MSD) allo-HSCT had the highest CIR compared with unrelated donor (URD) allo-HSCT and haploidentical (HRD) allo-HSCT (46.2%, 10.0% and 11.2%, P=0.002). The 3-year CIR in patient with pre-HSCT measurable residual disease negative (MRD-), MRD+ and non-remission was 10.9%, 30.0% and 72.2%(P&amp;lt;0.001). After multivariable adjustment, DTA mutations, pre-MRD+ or non-remission, MSD allo-HSCT remained statistically significantly associated with higher CIR as well as lower LFS and OS. In univariate and multivariate analyses, the cell of origin of AML did not impact the outcomes of NPM1-mutated AML patients accept allo-HSCT treatment . Conclusion: The DTA mutations ( DNMT3A, TET2, and ASXL1) were identified a distinct group of patients with adverse prognosis in NPM1-mutated AML patients who received allo-HSCT. Our results indicate that NPM1-mutated patients accept HRD/URD donors can achieve lower CIR than MSD donors in transplantation. HRD allo-HSCT may suitable for high risk patients within this group.

One-Pot Era-CRISPR/Cas12a System for Rapid and Sensitive Detection of Multiple NPM1 Mutations

Nucleophosmin 1 (NPM1) gene c.863_864 4-bp insertion mutations occur in 30% of adult acute myeloid leukemia (AML) patients, which is associated with a unique pathological phenotype and good prognostic significance. The detection of NPM1 mutations is important for risk stratification, treatment guidance, and minimal residual disease monitoring in AML. However, the current detection methods have their limitations in terms of equipment requirements, sensitivity, turnaround time, price, etc. Moreover, simultaneous detection of multiple NPM1 mutation types is challenging. Here, we developed a one-pot ERA-CRISPR/Cas12a-based assay in which only one guide crRNA is needed to detect hundreds of NPM1 gene variants, covering over 97% of NPM1-mutated AML cases. By optimizing multiple parameters, this assay finally achieved a sensitivity of as low as 0.01% within 30 min detection under constant 39℃, without wild-type related cross signal. In clinical detection, 21 NPM1 mutation positive cases from 70 AML patient were successfully screened out using our method, including two cases not detected by Sanger sequencing. This method is one-step, rapid, sensitive, specific, inexpensive, and without the need of Sequencer or temperature cycler. It holds the potential to assist clinicians in point of care testing.

Utility of Next-Generation Sequencing in the Detection of RNA Fusion Genes in Myeloid Malignancies in Singapore

Background Oncogenic fusion genes play an instrumental role in the pathogenesis of haematological malignancies and the identification of these genes is crucial for diagnosis, risk stratification and treatment decisions. Conventional diagnostic methods such as karyotyping and fluorescence in situ hybridization (FISH) are the current standard of care but are limited in the sensitivity and availability of different FISH probes for detection of fusion genes. In our institution, we have routinely integrated a targeted next-generation sequencing (NGS) panel in our diagnostic workflow for patients with suspected myeloid malignancies. Here we aim to review the cases with RNA fusion genes detected via NGS and compare these results with that of conventional diagnostic methods. Methods We analysed bone marrow samples from patients with suspected myeloid malignancies at the time of diagnosis in Singapore General Hospital (SGH). Extracted genomic DNA and RNA were amplified, and library preparation was performed with the Oncomine TM Myeloid Research assay (Thermo Fisher Scientific) to interrogate 40 DNA genes and 29 fusion driver genes. NGS libraries were sequenced on the Ion GeneStudio S5 System (Thermo Fisher Scientific) and analyzed using in-house bioinformatics pipelines and the Ion Reporter software. Limit of detection was determined as 3% allele frequency for DNA variants and 10 -2 for RNA fusions. RNA fusions detected by the NGS panel were consolidated and compared with cytogenetics (karyotyping and FISH) results. Results A total of 436 samples from adult patients with suspected myeloid malignancies were received for NGS testing from October 2021 to March 2023 in SGH, the largest tertiary hospital in Singapore. 306 had paired cytogenetics and NGS testing performed. 27 of the 306 cases (9%) had RNA fusions detected by NGS as summarized in Table 1 and Figure 1; their disease subgroups comprised of 16 acute myeloid leukaemia (AML), 5 chronic myeloid leukaemia (CML), 3 BCR-ABL1-negative myeloproliferative neoplasm (MPN), 2 acute promyelocytic leukaemia (APL) and 1 myelodysplastic syndrome (MDS). Eight AML cases had RUNX1-RUNX1T1.R3R3 fusions, with 6/8 (75%) harbouring additional DNA single nucleotide variants or indel mutations. Two AML cases had CBFB-MYH11 fusions, with 1 associated with KIT D419 deletion, and the other with U2AF1, NF1, and ASXL1 mutations. We also identified 2 adult AML cases (aged 49 and 65 years respectively) with DEK-NUP214.D9N18 fusions, both of which had associated FLT3-ITD mutations. There were 5 cases with KMT2A fusions, with a variety of partners including MLLT3.K9M6 (n=2), ELL.K9E2 (n=1), MLLT1.K8M4 (n=1), and MLLT6.K8M9 (n=1). Two patients with APL had PML-RARA detected for different variants: P3R3 and P6R3. Amongst the 8 MPN cases, 5 had BCR-ABL1 fusions consistent with diagnosis of CML. There was 1 case of MPN-Eo with FLP1L1-PDGFRA fusion, and the remaining 2 with ETV6-ABL1 and KMT2A-ELL.K9E2 rare fusions. The detected RNA fusions had clinical impact in establishing diagnosis (93%), prognosis (96%), and guidance of treatment decisions (89%). Out of the 27 cases with RNA fusions detected by NGS, 23 (85%) had concordant karyotype and FISH results. Four RNA fusions, namely KMT2A-ELL, CBFB-MYH11, FIP1L1-PDGFRA and ETV6-ABL1, were not detected by karyotyping or conventional FISH probes performed in our institution. It likely reflects the sensitivity of the NGS panel in detecting RNA fusions in cases of cryptic translocations. Conclusion We have demonstrated that NGS has a high sensitivity for identification of RNA fusion genes, is complementary to conventional cytogenetics testing, and has vast clinical impact in terms of diagnosis, prognostication and clinical management. We advocate for the integration of NGS with DNA and RNA sequencing into routine investigation of suspected myeloid malignancies for a more precise and comprehensive diagnostic approach.

Identification of a novel NPM1 mutation in acute myeloid leukemia.

e19019 Background: Nucleophosmin (NPM1) is predominant nucleolar localization. NPM1-mutant acute myeloid leukemia (AML) account for 25-35% of adult AML patients, has been defined as a distinct AML entity in the 2022 WHO classification. The majority of NPM1 mutations reported affect exon 12 (classical type A mutation), account for 75-80% of adult NPM1-mutated AML cases, which generate an extra nuclear export signal (NES) in the C-terminal leading to aberrant cytoplasmic localization. Similar aberrant cytoplasmic translocation has also been reported in NPM1 mutants that identified in exons 9 and 11. Identification of novel non-type A mutants is paramount for diagnosis, risk stratification, and prognosis for targeted population. Methods: Somatic mutation analysis is performed on 566 AML patients using next-generation sequencing (NGS). Immunofluorescence and immunohistochemistry were used to identify the cytological localization of mutant NPM1. HEK-293T cells transiently transfected with plasmids for ectopic expression of the GFP-mutated NPM1 fusion proteins were treated with the specific Crm1/XPO1 inhibitor leptomycin B and to evaluate the NES dependence of their subcellular localization. Results: We identified one AML patient with NPM1 mutant located on exon 5. This patient was a 59-year-old female with de novo AML, with an 18-nucletide in-frame insertion at position 405 (c.405_406insGCCCTGGAACTGGGGAAC, named MutSong) in the middle of exon 5 [variant allele fraction (VAF), 11.9%]. It generated a new NPM1 mutant protein (p.135insALELGN), and containing a leucine-rich NES. Notably, different with all NPM1 mutant in other exons reported so far, our exon 5 mutant retained the functional c-terminal but insert one leucine-rich NES, similar finding was recently reported in NPM1 mutants that identified in exon 5 but on different position. This NPM1 mutant in exon 5 resulted in cytoplasmic localization both by confocal and immunohistochemistry in AML patient sample. HEK-293T overexpressing the new GFP-NPM1 exon 5 fusion protein indicated the aberrant localization in the cytoplasm and partially in nucleoli. Moreover, NES-dependent cytoplasmic localization was inhibited by exportin-1 inhibitor leptomycin B, indicated that the novel NES generated by exon 5 mutant is responsible for cytoplasmic localization. This patient carried WT1, IKZF1, JAK2, and NUP98 mutations, and was intravenously treated with daunorubicin plus cytarabine for induction followed by 3 cycles of intermediate-dose cytarabine as consolidation. Then the patient received allogeneic hematopoietic stem cell transplantation (allo-HSCT). Unfortunately, the patient died of severe pulmonary infection and viral encephalitis 14 months after the initial remission, 7 months after allo-HSCT. Conclusions: Our finding of the novel NPM1 mutation in exon 5 supported that beside exon 12, exon 5 mutant is another NPM1 “born to be exported” mutant critical for leukemogenesis.

RAS Mutations in Adult Acute Myeloid Leukemia (AML). Frequency, Mutational Spectrum, and Identification of a Comutation Bias for KRASK117 (TET2/ASXL1)

Introduction RAS mutations are subclonal events associated with both good and bad prognosis AML categories (Papaemmanuil E et al.N Engl J Med 2016; 374). An adverse prognostic meaning of RAS mutations in treated AML with myelodysplastic-related changes has also been reported.In solid tumors, it has been shown that patients with KRASQ61K do not confer resistance to therapy, unlike other mutations: KRASQ61K introduces a cryptic splice donor site and causes a short inactive isoform (Kobayashi Y et al. Nature 2022 ;603). However, most KRASQ61K patients acquired the silent mutation G60G, which removes the new splice donor site and increases therapy resistance. This splicing mechanism was not present in cases with NRASQ61K and HRASQ61K. We investigated the frequency of RAS mutations, their molecular associations, and the prevalence of splicing mechanisms involving KRASQ61K and NRASQ61K in AML. Methods We included a consecutive series of adult de novo AML enrolled in the CETLAM trials, a total of 620 patients who were diagnosed between 2017 to 2022. DNA and RNA were extracted from bone marrow or peripheral blood. Next Generation Sequencing (NGS) was carried out with three panels containing at least 42 AML-related genes, including KRAS and NRAS. Only variants with a VAF (variant allele frequency) >4% and ranked as pathogenic or likely pathogenic were used for the analysis. Conventional karyotype was routinely assessed. cDNA was obtained from 7 patients with NRASQ61K, and was amplified through a conventional PCR. cDNA products were observed in gel electrophoresis and sequenced by the Sanger method. IBM SPSS Statistics was used to perform Chi-Square tests. In statistical tests regarding molecular associations, patients with co-mutated NRAS and KRAS are excluded. Results Our study found RAS mutations in 16.9% (105 patients) of adult AML cases. 11.9% of AML patients had mutated NRAS, while 6.1% mutated KRAS (74 and 38 patients, respectively). In our study, Q61 was the third most common mutation in NRAS with an incidence of 26%, while it was the fourth most common in KRAS with a frequency of 12%. The distribution of mutations in KRAS and NRAS is shown in figure 1. Regarding the Q61 codon, the distribution of each amino acid change in our cohort is shown in Figure 1b. Nine NRASQ61K were found, and cDNA could be obtained from 7 patients. NRASQ61K and non-mutated NRAS samples presented the standard large transcript in the gel electrophoresis. These findings ruled out the presence of exon skipping in AML with NRASQ61K mutations. No single case of KRASQ61K was observed in this extensive series of AML, so we could not identify alternative transcripts described in solid tumors nor compensatory KRASG60G mutations.1.1% of patients (7 cases) presented co-mutated NRAS and KRAS. In 9 subjects, more than one NRAS mutation was found, whereas, in 3 patients we detected two KRAS mutations. Lastly, we focused on molecular associations of RAS mutated patients (Figure 2). Of the total RAS-mutated cohort (105 patients), DNMT3A and NPM1 had the highest mutation frequency. Notably, 40.3% of NRAS-mutated patients had NPM1 mutations, significantly higher (p=0.018) than KRAS (16.1%). Another interesting finding was that KRASK117N mutations were mutually exclusive with DNMT3A and NPM1. All KRASK117N (6 cases) were associated with TET2 and or ASXL1 mutations. Cytogenetics information was available for 87 patients with RAS mutations (59 NRAS and 35 KRAS). NRAS-mutated patients showed a higher frequency of core binding factor (CBF) alterations than KRAS (18.1% vs. 3.6%, respectively) (p=0.063). Complex and miscellaneous karyotypes had higher rates of KRAS mutations than NRAS mutations (57.1% vs. 32.7%, respectively, p=0.032).In 26 cases, RAS mutations were the single mutation or the one with the highest VAF; notably, 42.3% had an NRASQ61 mutation. We had cytogenetic information of 20 patients, and twelve had inv(16) (60%). Conclusions In this cohort of adult AML cases, RAS mutations were present in 16.9%. Mutations at codon G12 were the most prevalent lesions in AML. NRAS mutations were more frequent than those of KRAS. Up to 17% of RAS mutated AML cases showed more than one mutation. CBF chromosomal abnormalities were commonly associated with NRAS mutations. Complex and miscellaneous karyotypes showed KRAS mutations more often. KRASK117N mutations were consistently associated with TET2 and ASXL1 mutations and mutually exclusive with NPM1 and DNMT3A. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Proteomics-Based Strategy for the Recommendation of Conventional Chemotherapy Vs Venetoclax Plus Hypomethylating Agents in Adult Acute Myeloid Leukemia (AML)

INTRODUCTION: Newer AML therapies with Hypomethylating Agents & Venetoclax (HV) provide an alternative to Conventional Chemotherapy (CC), with anthracycline and cytosine arabinoside, but criteria to select which to use are lacking. We investigated whether proteomic classification could provide guidance. METHODS: Using Reverse Phase Protein Array (RPPA) methodology, the level of 405 proteins (327 total; 78 post-translational modified) were measured in 818 samples from newly diagnosed adult AML patients and normalized to normal CD34+ cells. Patients treated with HV (N=85) or CC (N=335) were analyzed. Overall Survival (OS) and Complete Remission Duration (RD) analysis was performed with pairwise LogRank tests (p < 0.05), adjusted for False Discovery Rate (FDR), to identify individual proteins predictive of outcome. Patients were clustered based on protein expression. Categorical clinical variables were compared with Fisher's Exact test with simulated p-values (10000 replicates), and continuous ones with Kruskal-Wallis test. Univariate (UV) and Multivariate (MV) models were built with Cox proportional hazards model (CoxPH). Deep neural network analysis using random forest was performed to create a Protein Classifier for treatment recommendation. RESULTS: Individual protein analysis found 55 prognostic proteins (Protein Selector 1 - PS1), which clustered patients into 3 groups. A preferred therapy for cluster 2 was not identified with PS1, so a second Protein Selector 2 (PS2) was generated, identifying 45 predictive proteins in this subset. From the PS1/2 analysis, the 95 proteins formed 4 distinct expression profiles (C1=red, C2=gold, C3=magenta, C4=green) which were stratified by treatment (HV or CC), as shown in Fig. 1. Patients in C2 were enriched for secondary AML (57% vs C1=41%, C3=33%, C4=32%; p<0.001) and unfavorable cytogenetics (41% vs C1=32%, C3=39%, C4=35%; p<0.001), and were older (60yo vs C1=61yo, C3=54yo, C4=56yo; p<0.001). Mutations in CEBPA, GATA2, FLT3 and NPM1 were more common in C4 (22%, 13%, 35%, 35% vs C1=12%, 1.7%, 5.7%, 6%; C2=7.8%, 6.2%, 18%, 15%; C3=9.3%; 1.4%, 29%, 21%; p=0.037, p=0.013, p<0.001, p<0.001, respectively), highlighting the heterogeneity between groups. We evaluated OS and RD (Fig. 2) in each cluster (C1=red, C2=gold, C3=magenta, C4=green), considering each treatment (HV= solid line, CC=dashed line). Patients in C1 and C2 treated with HV had a diametrically different and superior response compared to those treated with CC (C1-HV: Median OS (MS)>65mo, Median RD (MRD)>60mo; C2-HV: MS=15mo, MRD=24mo vs C1-CC: MS=20mo, MRD=39mo; C2-CC: MS=10mo, MRD=17mo). C3 showed the opposite, with CC outperforming HV (MS=19mo, MRD>102mo vs MS=7mo, MRD=9mo). Similarly, in C4, CC patients did dramatically better, (MS=40mo, MRD>96mo vs MS=9mo, MRD=7mo). In the UV CoxPH model, we included variables for PS cluster and treatment and all groups predicted OS, except for C3-CC and C4-CC, as did age, secondary AML, complex karyotype, and CEBPA, FLT3, KIT, MLL, NPM1, PTPN11 and TP53 mutations. In the MV model, all clusters were predictive for OS, together with age, secondary AML, complex karyotype and NPM1 mutation. Regarding RD, in the UV model, only C2-HV, C2-CC, and C4-HV predicted RD, as did age, secondary AML, complex karyotype, unfavorable cytogenetics, and FLT3, RUNX1 and TP53 mutations. In the MV model of RD, all clusters, except for C3-HV were predictors, together with age. Importantly, optimized treatment triaging based on PS1/2 would have reassigned 52% (218/420) of cases, and is estimated to increase the 3yrs OS of the whole cohort from 35.6% to 48.8%, a 37.2% increase. Moreover, the 2yrs Relapse Free Survival would increase from 57,9% to 63,9%, a 10.39% increment. Furthermore, our Protein Classifier identified a set of 3 proteins (SPI1, NOTCH1.cle, and TGM2) whose expression can be used to rapidly classify patients and recommend either HV or CC. This classifier has a very high c-index (a measure of individual patient discriminatory power) of 0.93 (>0.7 is considered predictive, 1 would indicate perfection) demonstrating that it robustly predicts optimal therapy choice. CONCLUSION: Protein profiling identified protein expression patterns which had diametrically different responses according to treatment (HV or CC). Given the US annual incidence of 20000 newly diagnosed AML cases, proteomic triaging could result in ~2600 more cures using existing therapy. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

PB1801: CHROMOSOMAL ABNORMALITIES DURING THE CLINICAL COURSE OF ADULT ACUTE MYELOID LEUKEMIA

Background: Using conventional cytogenetics, an abnormal karyotype can be detected in 60-70% of adult acute myeloid leukemia (AML) patients. Taking into consideration the identified cytogenetic abnormalities AML patients are currently classified into three risk groups: favorable, intermediate and adverse. These genetic abnormalities are the most important factors in determining response to chemotherapy as well as outcome in AML. Aims: Aim of this study was to detect the diagnostic and prognostic significance of chromosomal abnormalities in leukemic cells at different phases of AML. Methods: Cytogenetic investigations of bone marrow and/or peripheral blood cells from 140 adult patients were performed during clinical evolution of AML: in newly diagnosed patients, in remission and at relapse. The methods of conventional cytogenetics (GTG) and fluorescence in situ hybridization (FISH) were used. Cytogenetic methods were performed using standard techniques and karyotypes were described according to the International System for Human Cytogenetic Nomenclature. Results: Cytogenetic investigations of 140 adult AML patients were performed in newly diagnosed patients, in remission and at relapse. Structural (del(5q), del(7q), rearrangements of 3q, 12p, 17p and 11q23, t(8;21)(q22;q22), t(9;22)(q34;q11), t(15;17)(q22;q21), t(16;16)(p13;q22), inv(16)(p13q22), marker and ring chromosomes, acentric structures and others) and numerical (-5, -7, -Y, +8, monosomal karyotype and others) chromosomal abnormalities in leukemic cells were found. Some genetic abnormalities (BCR/ABL1, PML/RARa, AML1/ETO, CBFβ/MYH11 fusion genes) were detected by molecular genetic methods (FISH). Spectrum of cytogenetic abnormalities had an important diagnostic and prognostic significance. Diagnosis of AML was possible due to the presence of specific genetic markers that can confirm some types of AML (AML with t(8;21)(q22;q22), t(9;22)(q34;q11), t(15;17)(q22;q21), t(16;16)(p13;q22)/inv(16)(p13q22), 3q21q26 rearrangements, BCR/ABL1, PML/RARa, AML1/ETO, CBFβ/MYH11 fusion genes). Taking into consideration the identified cytogenetic abnormalities AML patients were classified by risk groups: the group of patients with adverse cytogenetic markers (3q21q26 rearrangements, -5, -7, t(9;22)(q34;q11), BCR/ABL1 fusion gene, multiple changes (≥3), monosomal karyotype), the intermediate risk group without significant prognostic markers and the group with favorable prognostic factors (t(8;21)(q22;q22), t(15;17)(q22;q21), t(16;16)(p13;q22)/inv(16)(p13q22), AML1/ETO, PML/RARa, CBFβ/MYH11 fusion genes). In remission cytogenetic investigations showed a normal karyotype without cytogenetically visible changes in all patients. Cytogenetic investigations at relapse were performed in 5 of 140 AML cases. Additional/secondary chromosomal aberrations in leukemic cells or new cells clones with multiple structural and numerical changes at relapse was not detected. Summary/Conclusion: Cytogenetic investigations had an important significance for diagnosis, prognosis and selection the optimal treatment strategy of AML. Besides the analysis of chromosome banding patterns it is necessary for patients with AML to apply molecular genetic studies, namely FISH and polymerase chain reaction (PCR). Taking into consideration the identified cytogenetic abnormalities AML patients were classified by risk groups. Presence of the additional/secondary chromosomal aberrations in leukemic cells or new cells clones with multiple structural and numerical changes at AML relapse is the one sign of clonal evolution and progression of disease.

Nucleophosmin 1: from its pathogenic role to a tantalizing therapeutic target in acute myeloid leukemia

ABSTRACT Nucleophosmin 1 (NPM1, also known as B23) is a multifunctional protein involved in a variety of cellular processes, including ribosomal maturation, centrosome replication, maintenance of genomic stability, cell cycle control, and apoptosis. NPM1 is the most commonly mutated gene in adult acute myeloid leukemia (AML) and is present in approximately 40% of all AML cases. The underlying mechanisms of mutant NPM1 (NPM1mut) in leukemogenesis remain unclear. This review summarizes the structure and physiological function of NPM1, mechanisms underlying the pathogenesis of NPM1-mutated AML, and the potential role of NPM1 as a therapeutic target. It is reported that dysfunctional NPM1 might cause AML pathogenesis via its role as a protein chaperone, inhibiting differentiation of leukemia stem cells and regulation of non-coding RNAs. Besides conventional chemotherapies, NPM1 is a promising therapeutic target against AML that warrants further investigation. NPM1-based therapeutic strategies include inducing nucleolar relocalisation of NPM1 mutants, interfering with NPM1 oligomerization, and NPM1 as an immune response target.

A real-world study of infectious complications of venetoclax combined with decitabine or azacitidine in adult acute myeloid leukemia.

The purpose of this study was to identify the incidence, sites and main pathogens, and risk factors for infectious complications occurring in patients with adult acute myeloid leukemia (AML) during the first course of venetoclax combined with decitabine or azacitidine. A retrospective cohort analysis was performed of 81 patients with AML older than 14years who received the first cycle of venetoclax combined with a hypomethylating agent (HMA) between March 2018 and March 2021 at our institution. Infectious complications, if any, were documented. Among a total of 81 cases of AML, 59 (72.8%) patients occurred infections, including fever without an identifiable source (28.8%), clinically documented infections (40.7%), and microbiologically documented infections (30.5%). The most commonly isolated organism in culture was Candida albicans, followed by Klebsiella pneumonia, and Pseudomonas aeruginosa. The 4-week and 8-week mortality rates were 3.7% and 7.4%, respectively. In multivariate analysis, a high proportion of blasts in bone marrow, decreased hemoglobin level, and fever with or without a documented infection at baseline were significant independent risk factors for infectious complications. Compared with conventional chemotherapy, the incidence of infectious complications of venetoclax combined with decitabine or azacitidine significantly decreased. Pretreatment high leukemia burden and fever were independent risk factors for infections.

Independent prognostic significance of TP53 mutations in adult acute myeloid leukaemia with complex karyotype.

Adult acute myeloid leukaemia (AML) patients with complex karyotype (CK) generally have unfavourable outcomes. CK commonly co-exists with characteristic chromosomal and genetic abnormalities such as monosomal karyotype (MK), -17 or 17p- [abn(17p)] and TP53 mutations. Their individual prognostic significance needs to be clarified. Seventy-three adult CK-AML patients and eleven adult non-CK-AML patients with TP53 mutations (non-CK/TP53mu ) who were diagnosed and received therapy at our institute were enrolled. One hundred and fifty-seven AML cases retrieved from the cancer genome atlas (TCGA) for validation. Among CK-AML patients, those with TP53 mutations (CK/TP53mu ) had significantly lower rates of 1-course induction complete remission (CR), 2-year relapse-free survival (RFS) and 2-year overall survival (OS) than those without TP53 mutations (CK/TP53wt ); whereas, abn(17p) did not have the above impacts; MK was significantly associated with a lower 2-year OS rate but was not related to the rates of CR and RFS. Multivariate analysis showed that it were TP53 mutations and treating with chemotherapy alone but not MK and abn(17p) that independently predicted the adverse prognosis for RFS and OS in CK-AML. Furthermore, non-CK/TP53mu patients showed similar rates of CR, RFS and OS to CK/TP53mu patients. Validation using the TCGA cohort showed that CK/TP53mu patients had a significantly lower 2-year OS rate than CK/TP53wt patients, whereas abn(17p) and MK did not impact OS; the 2-year OS rate of patients with CK/TP53wt was similar to that of patients with intermediate-risk cytogenetics. Adult CK-AML patients have varied risks and TP53 mutations seem to be an independent adverse prognostic factor.

FLT3 Inhibitors Upregulate CXCR4 and E-Selectin Ligands and CD44 Via ERK Suppression in AML Cells, and Blockade of CXCR4 and E-Selectin Signaling with GMI-1359 Overcomes AML Resistance to Quizartinib in Vitro and In Vivo

FLT3 Inhibitors Upregulate CXCR4 and E-Selectin Ligands and CD44 Via ERK Suppression in AML Cells, and Blockade of CXCR4 and E-Selectin Signaling with GMI-1359 Overcomes AML Resistance to Quizartinib in Vitro and In Vivo

High EVI1 Expression Predicts Adverse Outcomes in Children With De Novo Acute Myeloid Leukemia

BackgroundA high ecotropic viral integration site 1 (EVI1) expression (EVI1 high) is an independent prognostic factor in adult acute myeloid leukemia (AML). However, little is known of the prognostic value of EVI1 high in pediatric AML. This study aimed to examine the biological and prognostic significance of EVI1 high in uniformly treated pediatric patients with AML from a large cohort of seven centers in China.MethodsA diagnostic assay was developed to determine the relative EVI1 expression using a single real-time quantitative polymerase chain reaction in 421 newly diagnosed pediatric AML patients younger than 14 years from seven centers in southern China. All patients were treated with a uniform protocol, but only 383 patients were evaluated for their treatment response. The survival data were included in the subsequent analysis (n = 35 for EVI1 high, n = 348 for EVI1 low).ResultsEVI1high was found in 9.0% of all 421 pediatric patients with de novo AML. EVI1 high was predominantly found in acute megakaryoblastic leukemia (FAB M7), MLL rearrangements, and unfavorable cytogenetic aberrance, whereas it was mutually exclusive with t (8; 21), inv (16)/t (16; 16), CEBPA, NPM1, or C-KIT mutations. In the univariate Cox regression analysis, EVI1 high had a significantly adverse 5-year event-free survival (EFS) and overall survival (OS) [hazard ratio (HR) = 1.821 and 2.401, p = 0.036 and 0.005, respectively]. In the multivariate Cox regression analysis, EVI1 high was an independent prognostic factor for the OS (HR = 2.447, p = 0.015) but not EFS (HR = 1.556, p = 0.174). Furthermore, EVI1 high was an independent adverse predictor of the OS and EFS of patients with MLL rearrangements (univariate analysis: HR = 9.921 and 7.253, both p < 0.001; multivariate analysis: HR = 7.186 and 7.315, p = 0.005 and 0.001, respectively). Hematopoietic stem cell transplantation (HSCT) in first complete remission (CR1) provided EVI1 high patients with a tendential survival benefit when compared with chemotherapy as a consolidation (5-year EFS: 68.4% vs. 50.8%, p = 0.26; 5-year OS: 65.9% vs. 54.8%, p = 0.45).ConclusionIt could be concluded that EVI1 high can be detected in approximately 10% of pediatric AML cases. It is predominantly present in unfavorable cytogenetic subtypes and predicts adverse outcomes. Whether pediatric patients with EVI1 high AML can benefit from HSCT in CR1 needs to be researched further.

Prognostic Nomogram for Acute Myeloid Leukemia Patients With Biallelic CEBPA Mutations.

Adult acute myeloid leukemia (AML) patients with biallelic mutations of CEBPA (biCEBPA) displays a favorable clinical outcome, and is defined as a unique entity in the 2016 World Health Organization classification. However, due to the intrinsic characteristics of the mutation, existence of co-occurring mutations and diversified gene expression signature, the prognosis of these patients needs to be analyzed in a more systematic way. In this study we evaluated the genetic characteristics and clinical outcome in a cohort of 137 biCEBPA AML cases, and proposed a prognostic nomogram to predict the overall survival (OS) of based on the clinical variables selected by multivariate Cox regression model in training cohort, including age, white blood cell count, co-existence of DNMT3A and CSF3R mutation and whether patients could achieve complete remission after induction therapy. The area under the receiver operating characteristic (ROC) curves for 3 and 5-year OS were 0.833 and 0.863, respectively. RNA sequencing of 4 relapsed patients showed that over-expression of VMP1 was an indicator of poor prognosis of biCEBPA AML patients. In conclusion, this prognostic nomogram might provide a more accurate prediction of the clinical outcomes of biCEBPA AML patients.

C-Terminal BRE Overexpression in 11q23-Rearranged and t(8;16) Acute Myeloid Leukemia Is Caused By Intragenic Transcription Initiation

Abstract TextOverexpression of the BRE (Brain and Reproductive organ-Expressed ) gene defines a distinct pediatric and adult acute myeloid leukemia (AML) subgroup (Balgobind et al, Leukemia 2010, and Noordermeer et al, Blood 2011 and 2012). BRE overexpression specifically associates with 11q23/MLL/KMT2A-rearranged AML, occurring in 70% and 50% of 11q23 childhood and adult AML cases, respectively. In adult 11q23-rearranged AML, BRE overexpression significantly associates with younger age. BRE functions in BRCA1-A (Breast Cancer 1) complex mediated DNA damage repair, BRISC (BRCC3 isopeptidase complex) complex mediated mitotic spindle assembly and in protection from apoptosis and senescence. Another constituent of the BRCA1-A/BRISC complexes is BRCC3 which is recurrently mutated in AML, especially in cases with a TET2 mutation (Huang et al., Haematologica, 2015). BRE is relatively highly expressed in blood cells in general but its exact function in hematopoiesis is unknown.By performing genome-wide ChIP-sequencing analyses on primary KMT2A-MLLT3 patient material (n=9) we identified a promoter enriched for active chromatin marks (H3K4me3/H3K27Ac) in BRE intron 4, exclusively in AML cases with BRE overexpression. Both 5' RACE end RNA-sequencing identified strong biallelic expression of a previously unknown C-terminal BRE transcript. This transcript starts near the intragenic promoter, with BRE intron 4 sequences spliced to exon 5 and downstream sequences. RNA-seq and qRT-PCR showed that the downstream transcript is expressed 5-8-fold higher compared to the full-length transcript, explaining BRE overexpression. Of note, two start codons in exon 5 of the downstream BRE transcript may cause the formation of N-terminally truncated BRE isoforms.By performing a qRT-PCR that specifically recognizes the new BRE transcript on samples from a large cohort of pediatric and adult AML (n=195) we observed that it was highly expressed in 60% of 11q23/ KMT2A -rearranged (n=58) and 70% of t(8;16)/ KAT6A-CREBBP cases (n=20), while it was virtually absent from other AML subsets (n=101) and normal hematopoietic tissues (n=14). In gene reporter assays, the AML-specific fusion protein KMT2A-MLLT3 transactivated the intragenic BRE promoter. Further epigenome analyses revealed 97 additional intragenic promoter marks frequently bound by KMT2A in AML with C-terminal BRE overexpression. The bound genes may be part of a context-dependent KMT2A-MLLT3-driven oncogenic program, because they were higher expressed in this AML subtype compared to other cytogenetically-defined AML groups (p<0.01). C-terminal BRE might be a contributor to this oncogenic program because in a case with relapsed AML, we observed an ins(11;2) fusing CHORD1 in frame to BRE exon 5. The CHORDC1 fusion to BRE exon 5 is remarkable because the alternate BRE transcript starts with exon 1b also fused to exon 5. This means that if the CHORDC1-BRE and alternate BRE transcripts would be translated, they would contain a similar C-terminal part of the BRE protein. The CHORDC1 disruption may also contribute to AML pathogenesis because it was recently shown that heterozygous Chord1 (a.k.a. Morgana) inactivation in mice results in the spontaneous development of atypical chronic myeloid leukemia (Di Savino, Blood, 2015). We conclude that abnormal expression of C-terminal alternate BRE sequences specific for AML is caused by distinct, non-random events; by a chromosomal abnormality and by intragenic transcription initiation in 11q23/ KMT2A -rearranged and t(8;16)/ KAT6A-CREBBP AML. These findings warrant further studies to determine whether leukemia-specific alternate BRE contributes to disease pathogenesis and how abnormal intragenic transcription activation in AML is brought about. DisclosuresHaferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Clonal Interference of Signaling Mutations Holds Prognostic Relevance in Core Binding Factor (CBF) Acute Myeloid Leukemia (AML)

Context. Mutations affecting cytokine signal transduction genes (‘signaling’ genes) are frequent in CBF AML and increase the risk of relapse. Clonal heterogeneity heralds poor prognosis in many tumors. Clonal interference is a specific type of clonal heterogeneity defined by the coexistence of independent clones affecting a common pathway, and has been linked to resistance to selection pressures in bacteria and viruses (Hughes, Nat Rev Genet 2015). Single-cell data has shown that in CBF AML with several signaling mutations, these mutations occur in independent clones (Hirsch, Nat Comm 2015). We investigated the prognostic role of clonal interference of signaling mutations in intensively-treated CBF AML.Methods. Diagnostic bone marrow (BM) samples from CBF AML cases from 2 multicenter adult and pediatric trials underwent targeted high-throughput sequencing (HTS) in Lille (n=215) as reported (Duployez Blood 2016). Diagnostic samples from adult CBF AML cases referred to Munich Leukemia Laboratory were analyzed with an independent HTS platform, including previously published RUNX1 - RUNX1T1 cases (CBFA, Krauth, Leukemia 2014) and additional CBFB - MYH11 (CBFB) cases (n=230 in total). All analyses were stratified on the HTS platform. We defined the number of ‘signaling clones’ as the total number of variants found in signaling genes (KIT, FLT3, NRAS, KRAS, JAK2 and CBL) by HTS in each patient.Results. The study cohort included 445 patients (pts, M/F 251/194, median age 42y [IQR 24 - 54]) with CBFA (n=230) and CBFB (n=215) AML. Median WBC was 14.6 x109/L. Sequencing data for all 445 pts was available for 6 signaling genes, with the following mutations frequencies: KIT 31%, NRAS 29%, FLT3 16% (TKD 10%, ITD 5%, both 1%), KRAS 13%, CBL (2%) and JAK2 (1%). FLT3-TKD, NRAS and KRAS mutations were significantly more frequent in CBFA AML, and FLT3-ITD in CBFB AML. Recurrent co-mutations in genes covered in >90% of cases included TET2 (6%) and ASXL1 (5%). At least one mutation in KIT, NRAS, KRAS, FLT3, CBL or JAK2 was found in 293 (66%) pts, of whom 126 harbored clonal interference, defined as ≥2 signaling clones, because of mutations in different genes (n=76), different variants in one gene (n=27), or both (n=23, Figure A). Gene redundancy, defined as the presence of ≥2 variants in one gene, particularly affected NRAS (n=27/50 cases) and KIT (n=12/50).Among cases with at least one signaling clone, clonal interference was more frequent in CBFB AML (48% vs 36% in CBFA, p=0.025), but was not associated with age, gender, leukocytosis, or to mutations in ASXL1 or TET2 . Importantly, the total allelic burden of signaling clones was comparable in cases with and without clonal interference (p=0.2, Figure B).All pts were treated with different 7+3 based intensive chemotherapy regimens as previously published (Duployez Blood 2016, Krauth Leukemia 2014). Median follow-up was 4.7 years, 47 (11%) pts received an allogeneic transplant in first CR. Censoring at transplant did not change our results. Three-year event-free survival (EFS) was 59.8% (95% CI, 51.2 - 67.3) in pts with no signaling mutation, 60.0% (52.0 - 67.2) in pts with a single signaling clone, and 46.4% (37.2 - 55.2) in pts with clonal interference (p=0.001, Figure C). This difference in EFS was seen in CBFB and to a lesser extent in CBFA AML. Clonal interference increased the cumulative incidence of failure (hazard ratio [HR]=1.54 compared to pts with a single signaling clone; p=0.03) translating into a trend towards poorer overall survival (HR=1.55; p=0.07). Martingale residuals indicated that each additional signaling clone worsened EFS (HR=1.30; p<0.0001). Presence of gene redundancy (≥2 variants in one gene) predicted worse EFS (p=0.04; Figure D) independently of the number of mutated signaling genes (p=0.10), further strengthening the adverse prognostic value of intratumoral heterogeneity in CBF AML. In a multivariate Cox model, clonal interference predicted a worse EFS (HR=2.32, p=0.002) independently of older age (p<0.0001), higher WBC (p<0.0001), CBF subtype (CBFA vs CBFB, p=0.004), and total allelic burden of signaling clones (p=0.13).Conclusion. Our results in a large cohort of CBF AML patients demonstrate that presence of two or more genetic lesions in recurrently mutated cytokine signaling genes predicts poor prognosis in CBF AML independently of clinical features. Our analyses suggest that clonal interference contributes to chemoresistance in AML. [Display omitted] DisclosuresItzykson:Janssen: Research Funding; Novartis: Research Funding. Fasan:MLL Munich Leukemia Laboratory: Employment. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Jourdan:NOVARTIS: Consultancy, Honoraria. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Autologous Transplantation Is Associated with Better Outcomes Compared to Chemotherapy Alone in a Minority-Rich Non-High-Risk Acute Myeloid Leukemia Cohort

Background:The role of autologous stem cell transplantation (autoSCT) in non-high-risk acute myeloid leukemia (AML) is controversial. It remains a frequent consolidation alternative for AML in first complete remission (CR1) in Europe and Asia, especially for intermediate risk disease, but has fallen out of favor over allogeneic transplantation (alloSCT) in the United States (U.S.). Donor availability, however, affects feasibility of alloSCT in ethnic and racial minorities that remain disproportionately underrepresented in donor registries. We explored the survival experience in our predominantly minority non-high-risk AML patients treated with autoSCT in comparison to those treated with chemotherapy alone, the current standard of care in the U.S. for favorable-risk AML as well as for intermediate-risk disease when alloSCT is not available.Methods:All adult patients diagnosed with AML between the years 1997 - 2015 were identified within the Montefiore Medical System in the Bronx, NY. For the purpose of our analysis we only included AML cases with favorable or intermediate cytogenetic risk that received induction with aggressive chemotherapy, achieved CR1 and underwent consolidation with high-dose chemotherapy alone or chemotherapy followed by autoSCT. We used descriptive statistics to compare baseline characteristics of patients treated with one of the two consolidation modalities. We used the Kaplan Meier survivor function to compare overall survival (OS) and relapse-free survival (RFS) between the two groups and multivariable Cox Proportional Hazard models to adjust for key covariates.Results:We identified 149 newly diagnosed adult AML cases with favorable (n=14) or intermediate-risk (n=135) cytogenetic risk. Among the 126 patients treated with aggressive chemotherapy, 96 (76%) achieved CR1 after induction chemotherapy, among which 10 experienced early relapse, 3 were lost to follow up and 83 proceeded to consolidation with either high-dose chemotherapy alone (n=58) or chemotherapy followed by autoSCT (n=25).The cohort was predominantly non-white (n=50/83, 60%). The median age was 57 years. Distribution of age, sex, race and cytogenetic-risk was similar between the two treatment groups (table 1). Median follow up was 17.7 months.Median OS was significantly longer for patients consolidated with autoSCT compared to those treated with chemotherapy alone (1.9 vs. 1.2 years; p=0.046). Recipients of autologous transplantation also had longer RFS compared to those consolidated with chemotherapy alone (1.6 vs. 0.6 years; p<0.01) (Figure 1). No treatment-related deaths were observed in the autoSCT group.In multivariable analysis adjusted for age, sex, race and cytogenetic risk, patients who underwent consolidation with autoSCT had a significantly better OS (HR 0.28; 95%CI 0.11 - 0.72; p<0.01) and RFS (HR 0.20; 95%CI 0.09 - 0.45; p<0.001) compared to those treated with chemotherapy alone.Conclusions:In this minority-rich non-high-risk AML cohort, we showed that consolidation with autologous transplantation provided OS and RFS benefit compared to chemotherapy alone. Survival rates in our multiethnic cohort were comparable to those historically reported in predominantly white AML cohorts. Autologous transplantation remains a beneficial treatment option in non-high risk AML in first remission and is a particularly valuable therapeutic strategy for ethnic/racial minorities when an allogeneic donor is not available. Integration of minimal residual disease assessment may help refine the selection of those more likely to benefit from this approach. [Display omitted] DisclosuresSteidl:Novartis: Research Funding; Aileron Therapeutics: Consultancy, Research Funding; GlaxoSmithKline: Research Funding; Celgene: Consultancy; Bayer Healthcare: Consultancy.

Identification of Relapse Risk Using a Multiparameter Flow Cytometry-Based Detection of Minimal Residual Disease in Adult Patients with Acute Myeloid Leukemia at ELN Intermediate-Risk

Background: Minimal Residual Disease (MRD) measurement is a useful tool to refine prognosis in adult Acute Myeloid Leukemia (AML) patients in morphological complete remission (CR). Multiparametric flow cytometry (MFC) may provide independent prognostic information particularly in intermediate (Int)-I and Int-II ELN 2010 risk groups.Patients and methods: Analysis was performed on bone marrow (BM) aspirates of 120 ELN Int-I/II patients (pts) from 2 independent institutions, separately analyzed in two MFC labs. Treatment consisted of one-two idarubicin-based induction courses followed by High-dose-Ara-C consolidation and by allogeneic transplantation (NILG, n: 18/51; GIMEMA, n: 20/69). The median age was 49yr (18-73yr), F/M ratio was 49/71; de-novo and secondary AML cases were 94.2% and 5.8%, respectively. FLT3 gene was mutated in 41.7% of the patients (50/120), being wild type in 56.6% of the cases (68/120). FLT3-ITD was detected in 35.0% of the patients (42/120) and FLT3-D835 was detected in 7.5% of the patients (9/120). NPM1 gene was mutated in 31.6% of the patients (38/120) being wild type in 65.8% of the cases (79/120). ELN 2010 int risk groups were distributed as follows: 79.2% Int-I (n: 95) and 20.8% Int-II (n: 25). At diagnosis, a panel of 22 monoclonal antibodies, combined in four 8-color tubes, was employed to detect leukemia-associated immunophenotypes (LAIPs). MRD was measured after the first cycle (TP1) and after the second cycle (TP2). MRD was considered positive if residual leukemic cells were equal or higher than 0.1% and 0.035% at TP1, and than 0.035% at TP2, respectively. Relapse-free survival (RFS) and overall survival (OS) were analyzed using Kaplan-Meier curves and the log-rank test.Results: One-hundred-eighteen and 116 patients were tested at TP1 and TP2, respectively. At TP1, MRD-positivity was detected in 72.9% (86/118) at a threshold of 0.1% and 83.9% pts (99/118) at a threshold of 0.035%. Both thresholds were significant in predicting relapse (P=0.0002 and P=0.0158 for the 0.1% and the 0.035% threshold, respectively). We observed a shorter RFS in MRD-positive versus MRD-negative patients: median RFS was 10.8 months (0.1% threshold) and 11.3 months (0.035% threshold) in MRD-positive pts as compared to 28.5 months (0.1% threshold) and 19.5 months (0.035% threshold) in MRD-negative patients. Furthermore, MRD-positive pts presented with a significantly shorter OS as compared to MRD-negative patients (0.1% threshold, P=0.0185), with a median OS of 18.2 months as compared to 33.7 months for MRD-negative pts. When the 0.035% threshold was used, OS was not different between the two groups of pts. At TP2, MRD-positivity was detected in 75.8% of the pts (88/116) and significantly predicted relapse (P=0.0023, 0.035% threshold): the median RFS was 10.8 months in MRD-positive pts vs 26.3 months in MRD-negative. Moreover, we observed a significant improvement in terms of OS in MRD-negative vs MRD-positive pts (P=0.0411), the median OS being 37.0 and 18.2 months, respectively. Notably, the proportion of MRD-positive and MRD-negative patients did not differ in terms of age, sex, ELN risk classification, FLT3 mutational status, and number of allogeneic transplanted patients.Conclusions: Overall, our findings demonstrate a correlation between MRD levels by MFC and relapse risk in ELN-intermediate risk patients. The use of an allogeneic transplantation approach rather than other consolidation strategies in ELN-intermediate risk AML patients is often debated. MFC-based MRD potentially allows for a more appropriate and tailored post-remission therapeutic strategy in these AML patients. Furthermore, harmonization of MRD analysis between different labs and different protocols sharing a common technical approach seems to be feasible. Future prospective studies are warranted for prospective evaluation of MRD in ELN-intermediate AML patients. DisclosuresRossi:AbbVie: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Teva: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees.