Acute Myeloid Leukemia M2 Research Articles

Abstract 424: Landscape of somatic mutations in drug-resistant acute myeloid leukemia

Abstract Introduction: Most patients with acute myeloid leukemia (AML) initially respond to cytarabine-anthracycline induction chemotherapy. However, in many patients, the disease recurs in a lethal drug-resistant form. Somatic mutations underlying the pathogenesis of AML have been extensively characterized by sequencing of newly diagnosed AMLs. However, the mutations driving therapy resistance and disease progression at relapse have not been well characterized. In this study, we have exome sequenced a cohort of relapsed and refractory AMLs and compared the landscape of somatic mutations at relapse to diagnosis phase AMLs to identify mutations that contribute to therapy resistance and disease progression. Materials and Methods: We performed exome sequencing of diagnosis phase AMLs (n=70) and relapsed or primary refractory AMLs (n=54). Patients with AML M3 subtype were excluded from the study. Paired diagnosis and relapse samples were available from 27 patients. A skin biopsy was used as the germline control. Nine patients had received an allogeneic hematopoietic stem cell transplant before relapse. Somatic mutations were called using varscan2 and copy number aberrations using copyCat. Since the identification of large insertions from next-generation sequencing data remains challenging using existing algorithms, FLT3 internal tandem duplications (FLT3-ITDs) were identified using a novel custom algorithm optimized for FLT3-ITD detection. Population variants were filtered out to remove donor-derived germline variants in chimeric post-transplant relapse samples. Results: Comparison of somatic mutation frequencies in diagnosis and relapse and refractory samples showed that on average relapsed tumors have a higher number of driver mutations than tumors at diagnosis. WT1, TP53, CBL, IDH1 and PTPN11 were mutated at a higher frequency in relapsed samples than at diagnosis, with 13 %, 11 %, 11 %, 9 % and 9 % of relapsed or refractory samples and 4 %, 6 %, 3 %, 4 % and 7 % of diagnosis mutated respectively. Analysis of paired diagnosis-relapse samples showed that in patients with WT1, CBL or PTPN11 mutation at diagnosis the second allele is frequently mutated or lost due to uniparental disomy occurring at relapse. Conclusions: On average relapsed AMLs have a higher number of driver mutations than diagnosis phase AMLs indicating that acquisition of additional driver mutations contributes to relapse. AMLs frequently acquire additional mutations in the same genes and pathways that already harbored mutations at diagnosis. Citation Format: Samuli Eldfors, Mika Kontro, Yevhen Akimov, Olli Kallioniemi, Kimmo Porkka, Caroline Heckman. Landscape of somatic mutations in drug-resistant acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 424. doi:10.1158/1538-7445.AM2017-424

Abstract 3659: WT1 and PRAME mRNA transfected TLR 7/8-polarized fast DC vaccines in AML patients mount specific immune responses and impact progression free survival

Abstract Patients diagnosed with acute myeloid leukemia (AML) may not be eligible for curative intensive treatment due to co-morbidity factors or age. Here we report results of 5 AML patients in morphological remission after incomplete induction/consolidation chemotherapy treated with dendritic cells (DCs) targeting WT-1 and PRAME. DCs were produced as described previously (Subklewe et al 2014), using a maturation cocktail containing the TLR 7/8 ligand R848. These DCs show a polarized release of IL-12p70 combined with low IL-10 upon stimulation. 2.5 or 5E+6 DCs per antigen were injected intradermally once weekly for 4 weeks (wks), in wk 6 and thereafter at monthly intervals. Blood and bone marrow (BM) samples were collected at regular intervals. Minimal residual disease (MRD) was measured in BM by quantitative PCR of WT-1 and PRAME expression and by morphology. Immune responses were assessed by analysis of intracellular interferon gamma expression or proliferation following stimulation with peptides spanning WT-1, PRAME, hTERT and survivin or after stimulation with autologous WT-1 and PRAME DCs. A 57 year old woman with intermediate risk M4 AML was vaccinated over 22 months after chemotherapy. Five weeks after start of vaccination she mounted strong CD8 responses against PRAME combined with an increase in hTERT response, suggesting epitope spreading. WT-1 signals in BM showed low positive levels throughout vaccination, but she remains in morphological remission 33 months after end of chemotherapy (EoC). A 50 year old man with M2 intermediate risk AML, initially not eligible for BM transplantation, showed specific immune responses against WT-1 during 10 months of vaccination. Due to Bell’s Palsy he was treated with cortisone which immediately reduced the vaccine effect, accompanied by increase of blasts in the bone marrow. Following new induction therapy he received BM transplantation and is currently in CR. A 68 year old woman with M1 intermediate risk AML is under vaccination for 24 months. WT-1 signals in BM continue to be slightly elevated without any sign of morphological relapse. CD4 responses and low CD8 responses are detected against WT-1, PRAME and hTERT. A 73 year old woman with M1 good risk AML relapsed after 6 months of DC vaccination without mounting specific immune responses. DC treatment was combined with 5-Azacytidine without any immunological and clinical effects. A 59 year old woman with good risk AML has been vaccinated for 14 months and is in remission for 21 months since EoC. WT-1 in BM remains slightly elevated whilst the initially positive PRAME signal is negative. Our results show that in 4 out of 5 AML patients fast TLR-polarized DC vaccination induced or supported specific T cell responses. 3 patients continue to be in CR after 21, 25 and 33 months respectively, following suboptimal primary chemotherapy. Immune responses are under investigation and will be presented. Citation Format: Iris Bigalke, Guri Solum, Dag Josefsen, Yngvar Fløisand, Kirsti Hønnåshagen, Lisbeth Skoge, Signe Spetalen, Stein Sæbøe-Larssen, Dolores J. Schendel, Gunnar Kvalheim. WT1 and PRAME mRNA transfected TLR 7/8-polarized fast DC vaccines in AML patients mount specific immune responses and impact progression free survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3659. doi:10.1158/1538-7445.AM2017-3659

Acute myeloid leukemia with NPM-MLF1 fusion gene: report of one case and review of literature

Objective To investigate the characteristics of NPM-MLF1 fusion gene in acute myeloid leukemia (AML). Methods The data of one AML patient with NPM-MLF1 fusion gene was analyzed,and literatures were reviewed. Results A female patient was diagnosed as AML M6. In the course of the disease, 2 hematologic relapsed, and 2 recurrences were associated with NPM-MLF1 fusion gene positive. After inductive treatment, hematologic complete remission was achieved, and NPM-MLF1 fusion genes were all negative. Survival time surpassed 6 years when the chemotherapy was performed alone. Conclusion The incidence of NPM-MLF1 fusion gene in AML is low. It is necessary to collect more clinical data to judge whether an independent disease type or not. Key words: Leukemia, myeloid, acute; Gene fusion; Translocation, genetic; Prognosis

Cell-based evaluation of changes in coagulation activity induced by antineoplastic drugs for the treatment of acute myeloid leukemia.

Idarubicin (IDR), cytarabine (AraC), and tamibarotene (Am80) are effective for treatment of acute myeloid leukemia (AML). In acute leukemia, the incidence of venous thromboembolism or disseminated intravascular coagulation is associated with induction chemotherapy. Procoagulant effects of IDR, AraC, and Am80 were investigated in a vascular endothelial cell line EAhy926 and AML cell lines HL60 (AML M2), NB4 (AML M3, APL), and U937 (AML M5), focusing on tissue factor (TF), phosphatidylserine (PS), and thrombomodulin (TM). IDR induced procoagulant activity on the surface of vascular endothelial and AML cell lines. Expression of TF antigen, TM antigen, and PS were induced by IDR on the surface of each cell line, whereas expression of TF and TM mRNAs were unchanged. Conversely, Am80 decreased TF exposure and procoagulant activity, and increased TM exposure on NB4 cells. In NB4 cells, we observed downregulation of TF mRNA and upregulation of TM mRNA. These data suggest IDR may induce procoagulant activity in vessels by apoptosis through PS exposure and/or TF expression on vascular endothelial and AML cell lines. Am80 may suppress blood coagulation through downregulation of TF expression and induction of TM expression. Our methods could be useful to investigate changes in procoagulant activity induced by antineoplastic drugs.

Simultaneous Occurrence of B-Cell Lymphocytic Leukemia and Acute Myeloid Leukemia in an Elderly Female Patient

Introduction: The coexistence of B-cell chronic lymphocytic leukemia (CLL) and acute myeloid leukemia (AML) in the same patient is rare. The aim of this study is to report of simultaneous occurrence of CLL and AML M5 in an old female patient for the first time. Case Presentation: A 72-year-old woman referred to hematology clinic for evaluation of leukocytosis, anemia, and thrombocytopenia. She had no known history of chronic illness or exposure to radiation or myelotoxic drugs. Physical examination showed the existence of generalized lymphadenopathy and splenomegaly. Section of the biopsy core disclosed a diffuse infiltration of lymphoid cells with hyperchromic irregular nuclei and scant cytoplasm in the background. Immunohistochemical staining for CD20 in lymphoid cells and CD68 in blastic cells were positive but it was negative for terminal deoxynucleotidyl transferase (TdT). The flow cytometric immunophenotyping analysis was performed in the presence of CD5 positive B-cell population (CD19; CD20 dim; CD23 and CD5/CD19) with small cell size that comprised 76% of cells with abnormal phenotype was revealed. Simultaneous occurrence of CLL and AML M5 was confirmed in the patients. Conclusions: We report a case of previously untreated CLL and AML M5 with rapidly progressive course to death in less than one month from diagnosis. To the best of our knowledge, the development of AML M5 in patient with CLL has not been reported before.

Signs and symptoms of rheumatic diseases as first manifestation of pediatric cancer: diagnosis and prognosis implications

ObjectiveTo assess the prevalence and describe the clinical, laboratory and radiological findings, treatment and outcome of children with cancer initially referred to a tertiary outpatient pediatric rheumatology clinic. MethodsRetrospective analysis of medical records from patients identified in a list of 250 new patients attending the tertiary Pediatric Rheumatology Clinic, Ribeirão Preto Medical School hospital, University of São Paulo, from July 2013 to July 2015, whose final diagnosis was cancer. ResultsOf 250 patients seen during the study period, 5 (2%) had a cancer diagnosis. Among them, 80% had constitutional symptoms, especially weight loss and asthenia, and 60% had arthritis. Initially, all patients had at least one alteration in their blood count, lactate dehydrogenase was increased in 80% and a bone marrow smear was conclusive in 60% of patients. Bone and intestine biopsies were necessary for the diagnosis in 2 patients. JIA was the most common initial diagnosis. The definitive diagnosis was acute lymphoblastic leukemia (2 patients), M3 acute myeloid leukemia, lymphoma, and neuroblastoma (one case each). Of 5 patients studied, 3 (60%) are in remission and 2 (40%) died, one of them with prior use of steroids. ConclusionThe constitutional and musculoskeletal symptoms common to rheumatic and neoplastic diseases can delay the diagnosis and consequently worsen the prognosis of neoplasms. Initial blood count and bone marrow smear may be normal in the initial framework of neoplasms. Thus, the clinical follow-up of these cases becomes imperative and the treatment, mainly with corticosteroids, should be delayed until diagnostic definition.

Congenital Acute Leukemia: A Rare Hematological Malignancy

Background: Acute leukemia presenting in the neonate and the young infant is a rare hematological malignancy. It may present at birth or within few days (congenital) or it may be diagnosed in first 4-6 weeks of life (neonatal) and may be myeloid or lymphoid in origin. High index of suspicion along with bone marrow studies, cytochemistry and flow cytometry is the mainstay of diagnosis. Case presentation: We report two cases of congenital acute leukemia presenting in second month of life. The first infant presented with fever, inconsolable crying, loose stools and vomiting of two weeks duration with pallor and hepatosplenomegaly. Lab investigations revealed anemia, leucocytosis and thrombocytopenia with 80% Blasts seen on peripheral blood smear. The bone marrow aspirate was hypercellular with greater than 3% of the blasts positive for myeloperoxidase (MPO). Based on morphology and immunophenotyping (IPT) by flow cytometry a diagnosis of Acute Myeloid Leukemia M2 (AML-M2) was made. The infant was managed with BFM intermediate risk induction protocol. Maintenance therapy was given for 18 months. The infant is in complete clinical and haematological remission. The second infant presented with fever, lethargy, poor feeding and palpable organomegaly. She had similar findings of leucocytosis with 90% blasts on peripheral smear which were negative for MPO. She was diagnosed as a Calla-positive B cell Acute Lymphoblastic Leukemia (ALL). She was managed as per the Interfant collaborative group protocol but succumbed to her illness. Conclusion: Congenital leukemia is characterized by an progressive nature of illness, greater achievement of remission in AML as compared to ALL, higher relapse rate, poor prognosis and difficulty in instituting combination chemotherapy. We report two infants diagnosed with congenital AML-M2 and Calla-positive B cell ALL based on cytochemistry and IPT. Resistance of leukemic cells in congenital ALL to chemotherapeutic drugs entails using a hybrid chemotherapeutic regimen.

Chromothripsis in Oncology: Literature Review and Case Report

The article presents a clinical case and literature review dwelling on the recently discovered chromothripsis phenomenon in oncology. Chromothripsis is a type of complex genome changes when a chromosome is first torn into dozens and even thousands of fragments, and then these fragments are bound in a random manner. Sometimes, several chromosomes are involved in the restructuring. As a result, genome mutant zones are formed which trigger malignancies and congenital diseases. In other words, the use of certain methodological approaches (multicolor fluorescence in situ hybridization, SKY technique, and some others) permits to observe under a microscope the splitting of two or more chromosomes and further reunification of these fragments into new unusual two- or multicolor structures, chromosomal markers. Chromothripsis is a rare phenomenon with a peculiar pattern observed in clones of cells of various tumors including hematopoietic and lymphoid tissue malignancies. There are published data on a higher incidence of this phenomenon in patients with myelodysplastic syndromes and bone tumors. TP53 gene mutations play an important role in the development of chromothripsis. The use of paired-sequencing DNA or SNP approaches in oncology is promising both in theoretical and clinical application. The first subject cohort should include patients with TP53 and MLL gene mutations, complex chromosomal aberrations, EVI-1 gene overexpression, and some others. The article presents the chromothripsis phenomenon in an 8-month-old girl with M7 acute myeloid leukemia.

A rare case of acute megakaryoblastic leukemia with orbital chloroma in a non-Down syndrome child

Acute megakaryoblastic leukemia (AMKL) is a rare disease accounting for 7%–10% of acute myeloid leukemia (AML) in children. It is uncommon in children without Down syndrome (DS). Orbital chloroma is usually associated with AML M2, M4, and M5. Herein, we report the case of a 22-month-old female who was diagnosed to have AMKL with orbital chloromas and without DS. Morphology and the initial panel of immunophenotyping were inconclusive and the presence of orbital chloromas added to the ambiguity. The presence of CD61 expression and marrow fibrosis supported by hyperdiploidy helped us clinch the diagnosis. Hence, comprehensive analysis of morphology, immunophenotyping, and cytogenetics is warranted to make an accurate diagnosis of AMKL.

The pheno-genotypic characteristics of infantile acute leukemia in a regional cancer center from South India

Introduction: Acute leukemia (AL) is uncommon in infants, with an annual incidence of 30 per million live births. They have peculiar biological characteristics. Although remarkable progress is seen in treatment of childhood AL, infantile AL remains a resistant subset with a dismally low 4-year survival of 35%. Objectives: To study the morphological, immunophenotypic, and cytogenetic features of infantile AL. A retrospective study of AL cases in children from birth up to 1 year of age, presenting to the departments of pediatric oncology and hematopathology between January 2010 and April 2015, was conducted. Results: Thirty-eight cases of infantile AL were included. The mean age at presentation was 10.2 months, and a female preponderance (M–F ratio: 0.65:1) was noted. Hyperleukocytosis (total white cell count >50 × 109/mm3) was seen in 13 (39.4%) cases. Immunophenotyping done in 31 cases showed pre-B acute lymphoblastic leukemia (B ALL) in 18 (58%), pre-T ALL in three (9.7%), and acute myeloid leukemia (AML) in 10 (32.3%). CD10 positivity was seen in 12 (57.1%) cases of ALL. Cytogenetic study done in 34 cases showed AML with recurrent genetic abnormalities in four. Mixed lineage leukemia (11q23) abnormality was seen in three cases of ALL. Two cases of AML were associated with trisomy 21. One case with features of AML M7 in a 4-day-old baby turned out to be transient abnormal myelopoiesis on follow-up. Conclusion: Literature on infantile AL from Indian studies is scarce compared to the available Western literature. Hence an epidemiological study of AL cases was done with review of literature, in an attempt to understand their pheno-genotypic features that influence their behavior. This may help in standardizing the treatment of these rare cases.

Sinais e sintomas sugestivos de doenças reumáticas como primeira manifestação de doenças neoplásicas na infância: implicações no diagnóstico e prognóstico

ObjectiveTo assess the prevalence and describe the clinical, laboratory and radiological findings, treatment and outcome of children with cancer initially referred to a tertiary outpatient pediatric rheumatology clinic. MethodsRetrospective analysis of medical records from patients identified in a list of 250 new patients attending the tertiary Pediatric Rheumatology Clinic, Ribeirão Preto Medical School hospital, University of São Paulo, from July 2013 to July 2015, whose final diagnosis was cancer. ResultsOf 250 patients seen during the study period, 5 (2%) had a cancer diagnosis. Among them, 80% had constitutional symptoms, especially weight loss and asthenia, and 60% had arthritis. Initially, all patients had at least one alteration in their blood count, lactate dehydrogenase (LDH) was increased in 80% and a bone marrow smear was conclusive in 60% of patients. Bone and intestine biopsies were necessary for the diagnosis in 2 patients. JIA was the most common initial diagnosis. The definitive diagnosis was acute lymphoblastic leukemia (2 patients), M3 acute myeloid leukemia, lymphoma, and neuroblastoma (one case each). Of 5 patients studied, 3 (60%) are in remission and 2 (40%) died, one of them with prior use of steroids. ConclusionThe constitutional and musculoskeletal symptoms common to rheumatic and neoplastic diseases can delay the diagnosis and consequently worsen the prognosis of neoplasms. Initial blood count and bone marrow smear may be normal in the initial framework of neoplasms. Thus, the clinical follow‐up of these cases becomes imperative and the treatment, mainly with corticosteroids, should be delayed until diagnostic definition.

GATA Factor-Dependent Positive-Feedback Circuit Controls Acute Myeloid Leukemia Cell Proliferation: Mechanisms and Targets for Therapeutic Intervention

Abstract Recent studies have established a relationship between GATA-2, a master regulator of hematopoiesis, and acute myeloid leukemia (AML). Loss-of-function mutations and elevated GATA2 expression are implicated in AML. While inhibitory GATA2 mutations occur in patients with primary immunodeficiency, which progresses to AML, GATA2 can be overexpressed in AML patients with poor prognosis. Many questions remain unanswered regarding mechanisms by which GATA2 deregulation results in AML. We reported that the Ras-p38α pathway induces multi-site GATA-2 phosphorylation in proerythroblasts, and Ser192 is required for the phosphorylation. Because RAS is mutated in 20% of AML patients, and elevated GATA2 expression correlates with poor prognosis, we tested whether this pathway functions in AML. GATA-2 was phosphorylated in the steady-state in AML cell lines, and oncogenic Ras(G12V) expression induced GATA-2 hyperphosphorylation in a p38/ERK-dependent manner. GATA-2 contains a sequence that conforms to the MAPK docking site termed "DEF motif". Mutation of S192 or the DEF motif abrogated Ras(G12V)-mediated GATA-2 hyperphosphorylation. We analyzed function using a Mouse Aortic Endothelial (MAE) cell line, in which GATA-2 and Ras(G12V) synergistically induce expression of specific GATA-2 target genes, e.g. Hdc. The DEF mutation abrogated GATA-2 hyperphosphorylation by Ras(G12V) and attenuated GATA-2 activity to induce Hdc expression in the presence of Ras(G12V) in MAE cells (50% of control, p < 0.05). GATA-2 hyperphosphorylation was also induced by expression of constitutively-active (ca) p38α and caMEK-1. caMEK-1-induced hyperphosphorylation was suppressed by the p38 inhibitor, SB203580 (p < 0.05) while the MEK inhibitor U0126 did not influence cap38α-induced hyperphosphorylation. The phosphatase inhibitor okadaic acid induced GATA-2 hyperphosphorylation, and p38 was more important than ERK in this context. These data support a model in which a hierarchical network of kinases controls GATA-2 phosphorylation and function in AML. Phospho-proteomics is being used to establish interconnectivity between network components and temporal aspects of the multi-step signaling mechanism. ChIP-Seq analysis in Kasumi-3 AML cells revealed GATA-2 occupancy at IL1B and CXCL2. IL1B and CXCR2, encoding the CXCL2 receptor, are implicated in AML. GATA-2 downregulation decreased expression of these genes (IL1B: 81.5% decrease, p < 0.001, CXCL2: 73% decrease, p < 0.001). Inhibition of ERK or p38 attenuated GATA-2 phosphorylation, decreased GATA-2 occupancy at GATA2, IL1B, and CXCL2, and lowered mRNA expression of these genes. To determine the consequences of the reduced occupancy, FAIRE was used to quantitate accessibility at the occupancy sites. Under conditions of reduced GATA-2 occupancy, accessibility decreased 50-65% at the GATA2 -77 enhancer and GATA-2 occupancy sites of IL1B and CXCL2 (p < 0.05). IL-1β and CXCL2 stimulated GATA-2 phosphorylation via p38 and ERK activation in Kasumi-1 and primary AML cells and increased GATA-2 chromatin occupancy and target gene expression. These results reveal a Ras-p38/ERK-GATA-2-IL-1β/CXCL2 axis that constitutes a positive-feedback circuit. We tested whether this circuit impacts Kasumi-1 cell proliferation. GATA2 downregulation reduced the proliferative rate (p < 0.001), which was partially rescued with recombinant CXCL2 (p < 0.01). To determine whether the signal-dependent mechanism can be extrapolated to human AML patients, we analyzed AML datasets from TCGA and GEO. GATA2 and CXCL2 expression was elevated in AML (p < 0.01). GATA2, IL1B, and CXCL2 mRNA levels correlated in M5 AML (GATA2 vs. IL1B: p=0.0015, GATA2 vs. CXCL2: p=0.0046, IL1B vs. CXCL2: p=0.000001). Among the patients with normal karyotype AML, high GATA2 and CXCL2 expression correlated with significantly reduced survival (p=0.003). In aggregate, our results establish a Ras-p38/ERK-GATA-2-IL-1β/CXCL2 axis in AML. It is attractive to propose that the GATA-2-dependent positive-feedback circuit is deployed upon ectopic elevation of GATA-2 activity in certain AML contexts and may harbor potential therapeutic targets. Given the profound capacity of this mechanism to alter GATA-2 activity, it is crucial to consider the GATA-2 activation state, rather than mRNA or total GATA-2 protein levels in clinical scenarios. Disclosures No relevant conflicts of interest to declare.

Alterations in Pathways Regulating Phosphatidil Inositol 3 Phosphate (PI3P) Produce Both Cell Proliferation and Therapy Resistance, and Define a Group of Patients with Poor Prognosis in Acute Myeloid Leukemia (AML)

IntroductionPI3P is a key regulator of cell growth, and mediates cell proliferation via PI3K/AKT/mTOR in response to various growth signals. Abnormal activation of genes in its pathway is associated to oncogenic activity and poor Overall Survival (OS). PI3P is also a core activator of autophagy. Which role autophagy plays in cancer is not well established; it can function as a pro-apoptotic mechanism, or it can improve survive to stresses clearing damaged mitochondria and proteins accumulation, preventing apoptosis. Levels and activity of pro-apoptotic and anti-apoptotic proteins, particularly bcl-2 and p53, membrane signaling via mTOR, high levels of cAMP, a complex made by pink/park, promote a switch from apoptotic autophagy toward a mechanism that augment cell resiliency.Our study aims to define the role of PI3P pathways in AML, and to establish if autophagy could reduce the patients' chance to respond to induction, and to worsen OS.MethodsWe analyzed 208 consecutive newly diagnosed non M3 AML patients, screened for TP53, FLT3, NMP1, IDH1, IDH2, and DNMT3A mutations. Remission status was assessed with bone marrow biopsy.In all the patients, we perform Microarray-based Comparative Genomic Hybridization with Affymetrix SNP array 6.0 or Cytoscan HD; we perform Whole Exome Sequencing (WES)in 80/208 patients.Survival data were collected prospectively, with a median follow-up of 18 months. Survival analysis was performed with Kaplan Meyer method using log rank test. Univariate and multivariable regression and Cox Hazard Ratio(HR) model was performed. Correlation between variables was assessed with Fisher's exact test.ResultsWe analyzed 4 pathways (Table 1); we selected genes in pathways basing on literature and GO data. Alterations in these pathways involved 103/209 patients (48%).PI3K/AKT/mTOR pathway alterations (both gains or losses) were shown to confer worst OS (p = .035, Figure 1a) when compared with unaltered patients; events in these pathways did not affect therapy response.Autophagy pathway alterations were shown to confer worst OS (p<.001, Figure 1b); alterations of autophagy were related to lower Complete Remission rate (CR%) after induction in univariate (p<.001) and multivariable regrassion with age, karyotype, secondary AML, TP53 mutation (p=0.014). Autophagy was significantly altered in patients with complex karyotype and TP53 mutation (p<.001).AMPK pathway alterations were shown to confer worst OS (p<.001, Figure 1c); Alteration of regulators in cAMP were related to lower CR% after induction in univariate (p<.001) and multivariable analysis with age, karyotype, secondary AML, TP53 mutation (p=0.009). AMPc pathway alteration was significantly associated with complex karyotype and TP53 mutation (p<.001).Autophagy switch pathway confer worst OS to patients(p<.001, figure 1d); autophagy switch was related to lower CR% after induction in univariate (p<.001) and multivariable analysis with age, karyotype, secondary AML (p<0.001). Autophagy switch was an independent risk factor in optimal Cox-HR model (p<.001, HR 2.996, CI 95% 2.101-4.271). Alterations in PINK or PARK did not showed to affect prognosis alone.Having at least an altered pathway is associated with worst prognosis (p <.001), and poor CR% after induction in univariate(p=.009) and multivariable analysis (p=.014).WES in a sub-cohort of patients did not found any significant mutation in genes we analyzed. This data is consistent with literature.ConclusionsOur work investigates for the first time the role of PI3P pathways and autophagy in AML. Surprisingly, it showed that both positive and negative alterations in these pathways are associated with poor prognosis. Significantly, alterations in cAMP and autophagy pathways were associated with therapy resistance.These results point out that both positive and negative regulation of autophagy could worsen patients OS; a diminished autophagy could be linked to a hyper-proliferative state via activation of AKT/mTOR but an augmented autophagy could give cell resiliency, favoring cytoplasm turnover, damaged mitochondria elimination, and neutralizing oxidative damages to proteins. A pan-PI3K inhibitor could target these mechanisms and improve chemo-sensitivity in high risk AML.Acknowledgment:ELN, AIL, AIRC, PRIN, Progetto Regione-Università 2010-12 (L. Bolondi), FP7 NGS-PTL project. [Display omitted] [Display omitted] DisclosuresGuadagnuolo:CellPly S.r.l.: Employment. Soverini:Ariad: Consultancy; Bristol-Myers Squibb: Consultancy; Novartis: Consultancy. Martinelli:Novartis: Speakers Bureau; MSD: Consultancy; Ariad: Consultancy, Speakers Bureau; BMS: Speakers Bureau; Celgene: Consultancy, Speakers Bureau; Pfizer: Consultancy, Speakers Bureau; Genentech: Consultancy; Roche: Consultancy, Speakers Bureau; Amgen: Consultancy, Speakers Bureau.

RUNX1-ETO-Dependent Transcriptional Repression of RASSF2 Contributes to t(8;21) Leukemia through Evasion of MST1-Driven Apoptosis Signaling

The t(8;21) chromosomal translocation is among the most frequent recurring cytogenetic abnormalities associated with acute myeloid leukemia (AML), found in 8-12% of de novo AML patients. The t(8;21) results in the stable fusion of the RUNX1 and RUNX1T1 genes, and formation of the oncofusion protein RUNX1-ETO (AML1-ETO). RUNX1-ETO is composed of the N-terminal DNA-binding domain of RUNX1 and nearly the entire ETO protein. RUNX1-ETO promotes leukemia development via the recruitment of transcription factor/transcriptional repression complexes (including NCOR, HDACs, p300, etc.) to regulatory regions of RUNX1 target genes known to be critical for myeloid differentiation and function, such as CEBPA, SPI1 (PU.1), NFE2, and CSF1R. Despite this knowledge, additional RUNX1-ETO target genes remain poorly characterized, and the complete molecular mechanism through which RUNX1-ETO leads to leukemic transformation remains to be elucidated. We propose that a better understanding of additional RUNX1-ETO target genes will lead to the potential for development of novel therapeutics to treat these patients.One such gene that we initially identified as markedly downregulated in RUNX1-ETO leukemia cells using a mouse model of t(8;21) AML is RASSF2 (Lo et al, Blood, 2012). Assessment of publicly available gene expression data revealed that RASSF2 is specifically downregulated in the bone marrow of t(8;21) AML patients compared to patients of different cytogenetic subtypes or to non-t(8;21) FAB subtype M2 AML patients. Additionally, RT-qPCR analysis confirmed that RASSF2 transcript is downregulated 10-100-fold in the t(8;21) AML cell lines, Kasumi-1 and SKNO-1, compared to non-t(8;21) AML cell lines and normal CD34+ hematopoietic cells. Expression of RUNX1-ETO in a non-t(8;21) AML cell line led to a reduction in RASSF2 mRNA expression, while knockdown of RUNX1-ETO in Kasumi-1 cells resulted in a ~5-fold increase in RASSF2 expression. Assessment of published ChIP-seq data showed that RUNX1-ETO directly binds at two regulatory regions within the RASSF2 genomic locus in t(8;21) AML cell lines and patient samples.Re-expression of RASSF2 at physiological levels in t(8;21) AML cell lines resulted in a modest negative growth phenotype, and greatly sensitized these cells to apoptosis following stimulation with various pro-apoptotic agents. Re-expression of RASSF2 in RUNX1-ETO-transduced primary mouse bone marrow caused these cells to lose their long-term self-renewal ability after 3 weeks in a serial replating/colony formation assay. This loss of self-renewal ability in co-transduced cells was accompanied by a marked increase in apoptosis during each of the first three weeks of replating. Mechanistically, re-expression of full-length RASSF2, but not of a deletion mutant lacking the SARAH heterodimerization domain (RASSF2ΔSARAH), in t(8;21) AML cell lines resulted in increased protein amount of the pro-apoptotic kinase, MST1. This suggests that RASSF2 may be a critical regulator of MST1 protein stability in AML cells. Importantly, modest (2-3-fold) overexpression of MST1 in t(8;21) AML cell lines resulted in a significant increase in apoptosis and caused growth arrest. The effects of RASSF2 or MST1 expression in non-t(8;21) AML cell lines were greatly reduced, suggesting that the cellular context of RUNX-ETO-driven leukemias makes them highly susceptible to MST1-dependent apoptosis.Overall, we have identified the importance of a MST1-driven pro-apoptotic signaling axis in t(8;21) leukemia. RUNX1-ETO-dependent transcriptional repression of RASSF2 may be essential for evasion of this apoptosis signaling during leukemic transformation via reduction of MST1 protein stability. MST1, perhaps better known as the mammalian orthologue of the drosophila Hippo kinase, is a critical tumor suppressor in many solid tumor types; and we believe our studies warrant the continued investigation of this pathway in hematological malignancy. DisclosuresNo relevant conflicts of interest to declare.

Suggestion of MUC16 (c.39169C>T) Mutation As a Potential Candidate for Predisposition to Myeloid Malignancy Associated with Trisomy 21

While the majority of leukemia cases occur in the absence of any known predisposing factor, there are germline mutations that significantly increase the risk of developing hematopoietic malignancies in childhood. Germline testing for the predisposition to myeloid malignancies is becoming more common with the recognition of multiple familial syndromes. In USA, Clinical Laboratory Improvement Amendments (CLIA) approved testing exists for mutations in RUNX1, GATA2, CEBPA and the other genes of inherited bone marrow failure syndromes. Meanwhile, GATA1 mutation is almost associated with Down syndrome and in acute myeloid leukemia (AML) with acquired trisomy 21. We experienced a case of infant AML with trisomy 21 and coexisting mutation of MUC16.We experienced infant leukemia (FAB classification, AML M7, Acute megakaryoblasticleukemia) with trisomy 21 and coexisting MUC16, SCRIB, and CEBPA mutation in a 7-month-old boy. Peripheral blood (PB) was taken from the boy every day during hospitalization. Bone marrow (BM) examination was performed at initial diagnosis and after remission. All of PB and BM samples were collected with informed consent, and the study was reviewed and approved by the Institutional Review Board of Seoul National University College of Medicine. The selected five PB samples are demonstrated on Fig. 1. G-banding revealed trisomy 21 and fluorescent in situ hybridization (FISH) for chromosome enumeration 21 revealed 71% with trisomy 21 cells in his diagnosis. At first, we suspected transient abnormal myeloproliferative disease associated with mosaic Down syndrome and serially monitored the proportion of cells with trisomy 21 by FISH and the percentage of blast during hospitalization. Cells with trisomy 21 decreased as blast disappeared in PB during chemotherapy, suggesting trisomy 21 is acquired abnormality in leukemic cells.We performed target gene sequencing of 359 genes related to the hematologic neoplasm, bone marrow failure syndrome, and cancer susceptibility to selected five PB. In addition, to search for the inherited predisposition gene to AML, we also performed whole genome sequencing (WGS) with BM specimen at initial diagnosis and after achieving remission. These next generation sequencing (NGS) with the Illumina HiSeq2500 platform revealed MUC16 (c.39169C>T) is the only significant mutation that persisted throughout from initial diagnosis to post-remission status. Mutations which were seen at initial diagnosis but disappeared after achieving remission were SCRIB (c.2197G>A, p.Arg733Trp) and CEBPA (c.371G>A, p.Ala124Val).To investigate whether MUC16 (c.39169C>T) mutation is rare variant which can be detected in normal person, we screened MUC16 mutation in healthy control (n=365) and in patients with other hematologic diseases (adult myelodysplastic syndrome, n=155; adult aplastic anemia, n=57; adult myeloproliferative neoplasm, n=44; childhood myeloid neoplasm, n=26; inherited bone marrow failure syndrome, n=21; idiopathic eosinophilia, n=4; familiar hemophagocytic lymophohistiocytosis, n=10; congenital neutropenia, n=1) using allele-specific PCR. MUC16 (c.39169C>T) was found in none of them. Discovered site of MUC16 (c.39169C>T) was not reported in solid tumor as well, though the other sites of MUC16 were frequently reported. It is generally known that AML with trisomy 21 accompanies GATA1 mutation, but the infant AML in this study did not accompany GATA1 mutation, but rather, MUC16 mutation.In this patient, GATA1 mutation was not found throughout hospital course. Instead, here we suggest that the constitutional MUC16 (c.39169C>T) mutation coexerts with acquired trisomy 21 in the development of infant AML and that MUC16 (c.39169C>T) mutation is a potential candidate gene for predisposition to AML. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

CSF3R Mutations Are Predominantly Subclonal Events in Intermediate Risk Karyotype AML and Prevalently Occur with CEBPA Mutations

Background: The colony stimulating factor 3 receptor(CSF3R) plays an important role in granulocyte differentiation and acquired mutations have been described in different myeloid malignancies. Besides chronic neutrophilic leukemia (CNL) and severe congenital neutropenia (SCN), mutations in CSF3R (CSF3Rmut) have also been reported in acute myeloid leukemia (AML) at low frequency and recently have been associated with CEBPA mutations (Lavallèe et al., Blood 2016; Maxson et al., Blood 2016) in adult and pediatric AML. First studies suggest sensitivity of CSF3Rmut CNL against JAK kinase inhibitors suggesting the possibility for targeted therapy for a subset of CSF3Rmut AML patients.Aims: The evaluation of CSF3Rmut in AML with intermediate-risk karyotype for frequency and association with other mutations with special focus on CEBPA mutation status.Methods: We analyzed 274 cases with intermediate risk de novo AML for CSF3Rmut by next generation sequencing (Illumina, San Diego, CA) of exon 14 and 17 (sensitivity: 1%), which contain the mutational hotspot regions of CSF3R. As CSF3R mutations were associated with CEBPA mutations in the above-mentioned studies, we analyzed a selected cohort of 179 CEBPA mutated cases in comparison to 95 cases with CEBPA wild type.202 cases (74%) had normal karyotype (CN-AML) and 72 (26%) had intermediate-risk aberrant cytogenetics according to MRC criteria. Female/male ratio was 127/147 and age ranged from 16- 88 y (median: 64 y). Mutation status of the following genes were available in all cases: ASXL1, CEBPA, FLT3-ITD, FLT3-TKD, GATA2, IDH1/2, KRAS, KMT2A-PTD, NPM1, NRAS, RUNX1 and WT1. All mutations with a variant allele frequency (VAF) < 10% were defined as subclonal. 95 cases had wild type CEBPA (CEBPAwt), 92 cases were CEBPA single mutated (CEBPAsm), 81 cases were CEBPA double mutated (CEBPAdm), 6 cases showed CEBPA mutations withloss of heterozygosity (CEBPA LOH).Results: Overall, in 7/274 patients (3%) CSF3Rmut were detected. This is slightly higher than expected in an unselected AML cohort (Sano et al., Br J Haematol 2015). Cytomorphology revealed AML M0 (n=1), AML M1 (n=2), AML M2 (n=3) and AML M4 (n=1). With regard to immunophenotype CSF3Rmut cases did not differ from CSF3Rwt cases which may be due to limited case numbers. All CSF3Rmut cases had CN-AML. The activating transmembrane mutation p.Thr618Ile, which has been identified as common mutation in CNL was the most frequent mutation (4/7). In addition two mutations leading to a truncated receptor (p.Ser715* and p.Gln749*) and a previously not described frame-shift mutation leading to a defecting splice variant (p.Ser783Glnfs*6) were identified. The median CSF3R VAF was 7% (range 2-45%). In 5/7 cases, CSF3Rmut was subclonal (VAF <10%, see figure). CSF3Rmut cases showed a median of 2 concomitant mutations (range 1-4). The most frequent concomitantly mutated gene was CEBPA (5/7 cases; 2 cases CEBPAsm, 1 case CEBPAdm and 2 cases with CEBPA LOH), followed by NRAS (3/7), ASXL1 (2/7) and RUNX1 (2/7). IDH1, TET2 and WT1 were mutated in one case each (see figure). Mutations in NPM1, FLT3, KMT2A or GATA2 were never detected. Interestingly, in CEBPAdm and CEBPA LOH cases, CSF3R was the only concomitantly mutated gene, whereas CEBPAsm cases harbored mutations in at least 1 gene in addition to CSF3R. In 2/7 CSF3Rmut cases we analyzed changes in the patterns of genetic lesions between diagnosis and relapse. Case 1 harbored CEBPAdm and a subclonal CSF3Rmut (VAF: 7%) at diagnosis. At relapse the CEBPAdm persisted, whereas CSF3Rmut was lost. Case 2 harbored two germline CEBPAmut and one somatic CEBPAmut at diagnosis. In addition, mutations in CSF3R and WT1 were identified. At relapse both the somatic CEBPAmut and WT1mut persisted, whereas CSF3Rmut was lost. These results indicate again that the CSF3Rmut was present only in a subclone of CEBPAmut AML in both cases.Due to the limited number of CSF3Rmut cases, we did not perform survival analysis in this study.Summary: 1) CSF3R mutations are rare and predominantly subclonal events in AML and therefore may not be a suitable target for directed therapy.2) CSF3Rmut prevalently occur with CEBPA mutations but are not limited to CEBPAdm. [Display omitted] DisclosuresFasan:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Perglerová:MLL2 s.r.o: Employment. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Other: Part Owner MLL Munich Leukemia Laboratory.

The Genomic Landscape of Childhood and Adult Acute Erythroid Leukemia

Introduction: The genetic basis of several acute myeloid leukemia (AML) subtypes remains poorly characterized, such as that of acute erythroid leukemia (AEL, AML M6) which is currently subclassified by morphology alone, and is associated with poor outcome. Here we sought to perform a definitive genomic analysis of AEL and translate these findings into faithful experimental models and novel therapeutic approaches.Methods: We studied 151 AEL cases (19.2% pediatric, 4.6% young adult, 21.9% adult and 54.3% older adult). Diagnosis of AEL was centrally confirmed and subclassified according to WHO 2008 and revised 2016 criteria. Whole exome and/or genome sequencing, RNA-sequencing and SNP array analysis were performed on all cases and in 2 AEL cell lines (TF-1 and Hel). Genomic data were compared to those from non-M6 childhood and adult AML from TARGET (n=192) and TCGA (n=197) studies. The functional effects of fusion transcripts and mutated genes were examined in IL-3 dependent Ba/F3 cells, NIH3T3 cells for focus formation assays and/or mouse lineage negative hematopoietic stem cells (lin- HSC) for colony forming and transplantation assays. Avatars of human AEL were established in immunocompromised NSGS and MISTRG mice for preclinical studies.Results: a) Genomic landscape of AEL. We identified 2,250 non-synonymous clonal and subclonal somatic mutations in 1,723 genes with a mean of 16.4 per case (range 2-88) and with missense and frameshift mutations accounting for 47.1% and 22.5% of all mutations, respectively. 78 genes were recurrently mutated in at least 3 cases. In frame fusions were detected in 31% of childhood and 27.5% of adult cases, and were more frequent in cases with complex karyotype. 124 potential driver genes were identified by statistical analysis or known pathogenic role in cancer, 9 of which were recurrent novel targets of mutation, most commonly involving chromatin modification (60.3%), cell cycle/tumor suppression (TP53, 33.8%), DNA methylation (28.5%), transcription regulation (26.5%), splicing (15.9%), NPM1 (11.9%), Ras (11.3%), JAK-STAT signaling (9.9%), the cohesin complex (8.6%), ALK/NTRK1 (4.6%) and PI3K signaling (3.3%). Overall, 33% of cases harbored a mutation in signaling genes amenable to inhibition by tyrosine kinase/Ras inhibitors. Mutations in TP53 and DNA methylation genes were significantly more frequent in adults while mutations in transcription regulators and Ras pathway were more frequent in children. Splicing mutations correlated with MDS phenotype and PI3K alterations with therapy-related AEL. Based on the co-occurrence and exclusivity of mutations 7 main distinct AEL genetic subtypes were defined: 1) pediatric AEL with NUP98-rearrangements (3.3% of all cases); 2) adult complex karyotype AEL with TP53 mutations (33.8%); 3) AEL with MLL-rearrangements (12.6%); 4) NPM1-mutated AEL (11.9%); 5) DNA-methylation/splicing mutated AEL (17.8%); 6) splicing/Ras/transcription regulation mutated AEL (21.2%) and 7) Other (8.6%) (Fig.1A). Mutations of chromatin modifiers occurred independently of karyotype, age and subtype (Fig.1B). NUP98-fusions and mutations in PTPN11, UBTF and GATA1 were more frequent in pediatric AEL compared to non-M6 AML. Among adults, mutations in TP53 and MLL were more frequent in AEL while FLT3, NPM1, DNMT3A and IDH1 were more in frequent in non-M6 subtypes. A complex karyotype, therapy-related AEL, TP53 mutations and NUP98-rearrangements were associated with poor outcome.b) Functional AEL modeling and therapeutic translations. Expression of NUP98-JARID1A in lin- HSC resulted in sustained self-renewal and development of an aggressive transplantable leukemia. At least three classes of signaling pathway mutations are targetable in AEL. ALK mutations in the extracellular MAM domain transformed Ba/F3 cells which were sensitive to crizotinib in vitro. Mutations in the tyrosine kinase domain of NTRK1 transformed NIH3T3 cells and were sensitive to entrectinib in vitro. Targeting of JAK-STAT, mTOR and PI3K pathways were examined in xenografts and sensitivity to JAK2 inhibitor ruxolitinib was confirmed in vivo.Conclusions: We provided the first comprehensive landscape of genomic alterations in AEL and defined distinct genomic groups with unique patterns of mutation occurrence compared to non-M6 AML. Finally, we showed that several pathogenic pathways are amenable to inhibition by approved targeted compounds. [Display omitted] DisclosuresMeggendorfer:MLL Munich Leukemia Laboratory: Employment. Wei:Novartis: Honoraria, Research Funding. Loh:Abbvie: Research Funding; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Mullighan:Amgen: Speakers Bureau; Incyte: Membership on an entity's Board of Directors or advisory committees; Loxo Oncology: Research Funding.

Acute myeloid Leukemia

Acute myeloid leukemia (AML) has been genetically characterized extensively and can now be subdivided into 9 to 11 pathogenetically different subtypes according to their profile of driver mutations. In clinical practice karyotyping and molecular analysis of NPM1, cEBPa and FLT3-ITD are required for treatment stratification and potentially genotype specific treatment. Some markers such as NPM1 not only offer prognostic information but can also serve as markers of minimal residual disease and thus have the potential to guide therapy in the future.The basis of curative treatment is intensive combination chemotherapy comprizing cytarabine and an anthracycline ("7 + 3" regimen). The prolonged duration of aplasia can be reduced significantly by accelerated therapy ("S-HAM" regimen). Following achievement of a complete remission patients with a low risk of relapse - based on genetic and clinical features - receive chemotherapy based consolidation therapy whereas high risk patients - and potentially also those with an intermediate risk - receive an allogeneic stem cell transplantation. Whereas adding the rather unspecific tyrosinekinase inhibitor sorafenib to standard treatment in unselected AML patients has not improved overall survival (OS), the addition of midostaurin to standard therapy in the selected group FLT3 mutated patients has resulted in a moderate but significant OS benefit.Real world data show that in patients below 50 years a cure rate of ca. 50 % can be achieved. However less than 10 % of patients above the age of 70 will be alive after five years even after intensive treatment. Therefore when curative and intensive treatment is deemed impossible the therapeutic standard in elderly and unfit patients used to be low-dose cytarabine with an average OS of 4 months. This has now been replaced by a new standard of care of hypomethylating agents - azacytidine and decitabine - which both achieve higher remission rates and show strong trends towards a prolonged OS of between 8 and 10 months.The paradigm for genotype-specific therapy is acute promyelocytic leukemia (APL - or AML M3 in the former FAB classification). This entity used to be a problematic AML subgroup because of its frequent coagulation disturbances and potentially fatal bleeding problems. Today patients with APL can be treated with a chemotherapy free combination of ATRA - a differentiating agent - and Arsenic Trioxide - an apoptosis inducing agent. In patients with a leukocyte count < 10 000 / µl a cure rate of > 90 % can now be achieved.

Brazilian guidelines on hematopoietic stem cell transplantation in acute myeloid leukemia.

Acute myeloid leukemia (AML) accounts for 90% of all cases of acute leukemia in adults. In Brazil, the mortality from myeloid leukemia is 1.74/100 000 men and 1.37/100 000 women. Our aim was to review and update guidelines of the Brazilian Society of Bone Marrow Transplantation on indications of hematopoietic stem cell transplantation (HSCT) for the treatment AML. (i) Allo-HSCT is recommended for high-risk AML (IA); (ii) allo-HSCT is recommended for AML of intermediate risk (IA); (iii) allo-HSCT is recommended for AML relapsed/refractory (C4); (iv) auto-HSCT is recommended for AML after 1 consolidation (C4); (v) auto-HSCT is recommended for AML in CR1 (higher than QT in the Brazilian experience) (C4); (vi) auto-HSCT is accepted for AML M3 in second molecular complete remission (2B); (vii) peripheral blood instead of Bone Marrow HSC for advanced disease (2A); (viii) recommended conditioning protocols: Bu-Cy/Bu-Mel, Bu-Flu, TBI-Cy. In umbilical cord HSCT, consider ATG-based protocols (2A); (ix) allogeneic HSCT for the treatment of AML can be used in patients between 60 and 80 yr with good performance status and the absence of significant comorbidities (C4).

AC220 and AraC cause differential inhibitory dynamics in patient-derived M5-AML with FLT3-ITD and, thus, ultimately distinct therapeutic outcomes.

Engrafting the bone marrow cells of a patient with M5 acute myeloid leukemia into immunocompromised mice (AM7577) resulted in serially transferrable stable AML and eventual mortality. The disease starts in the bone marrow and then expands to peripheral areas, which is typical of M5 leukemogenesis, where high leukemic burden in blood is coincident with symptoms/mortality. The leukemic cells in the mice had myeloid morphology, phenotypes, and genotypes (including the internal tandem duplication of FMS-like tyrosine kinase receptor 3 gene [FLT3-ITD]) similar to those of the original patient. Autocrine mechanisms of human granulocyte-macrophage colony-stimulating factor/interleukin-3 likely support AM7577 growth in mice. Treatment with FLT3 TKI (AC220) caused complete remission in peripheral blood, spleen, and bone, along with relief of symptoms and extended life, hinting that FLT3-ITD may be a key leukemogenic driver maintaining the disease. Interestingly, withdrawal of AC220 (high dose) did not result in relapse of disease, suggesting cure. These results, however, are in contrast to cytarabine (AraC) induction treatment: First, although AraC significantly suppresses the diseases in blood, and to a lesser degree in bone marrow and spleen, the suppression is temporary and does not prevent eventual onset of disease/death. Second, the withdrawal of AraC always resulted in rapid relapse in peripheral blood and eventually death. Our observation in this patient-derived model may provide useful information for clinical applications of the two drugs.