Elevated frequencies of leukemic myeloid and plasmacytoid dendritic cells in acute myeloid leukemia with the <Emphasis Type="BoldItalic">FLT3</Emphasis> internal tandem duplication

Some 30% of acute myeloid leukemia (AML) patients display an internal tandem duplication (ITD) mutation in the FMS-like tyrosine kinase 3 (FLT3) gene. FLT3-ITDs are known to drive hematopoietic stem cells towards FLT3 ligand independent growth, but the effects on dendritic cell (DC) differentiation during leukemogenesis are not clear. We compared the frequency of cells with immunophenotype of myeloid DC (mDC: Lin⁻, HLA-DR⁺, CD11c⁺, CD86⁺) and plasmacytoid DC (pDC: Lin⁻, HLA-DR⁺, CD123⁺, CD86⁺) in diagnostic samples of 47 FLT3-ITD⁻ and 40 FLT3-ITD⁺ AML patients. The majority of ITD⁺ AML samples showed high frequencies of mDCs or pDCs, with significantly decreased HLA-DR expression compared with DCs detectable in ITD⁻ AML samples. Interestingly, mDCs and pDCs sorted out from ITD⁺ AML samples contained the ITD insert revealing their leukemic origin and, upon ex vivo culture with cytokines, they acquired DC morphology. Notably, mDC/pDCs were detectable concurrently with single lineage mDCs and pDCs in all ITD⁺ AML (n = 11) and ITD⁻ AML (n = 12) samples analyzed for mixed lineage DCs (Lin⁻, HLA-DR⁺, CD11c⁺, CD123⁺). ITD⁺ AML mDCs/pDCs could be only partially activated with CD40L and CpG for production of IFN-α, TNF-α, and IL-1α, which may affect the anti-leukemia immune surveillance in the course of disease progression.     

Targeting of acute myeloid leukaemia by cytokine‐induced killer cells redirected with a novel CD123‐specific chimeric antigen receptor

Current therapeutic regimens for acute myeloid leukaemia (AML) are still associated with high rates of relapse. Immunotherapy with T‐cells genetically modified to express chimeric antigen receptors (CARs) represents an innovative approach. Here we investigated the targeting of the interleukin three receptor alpha (IL3RA; CD123) molecule, which is overexpressed on AML bulk population, CD34⁺ leukaemia progenitors, and leukaemia stem cells (LSC) compared to normal haematopoietic stem/progenitor cells (HSPCs), and whose overexpression is associated with poor prognosis. Cytokine‐induced killer (CIK) cells were transduced with SFG‐retroviral‐vector encoding an anti‐CD123 CAR. Transduced cells were able to strongly kill CD123⁺ cell lines, as well as primary AML blasts. Interestingly, secondary colony experiments demonstrated that anti‐CD123.CAR preserved in vitro HSPCs, in contrast to a previously generated anti‐CD33.CAR, while keeping an identical cytotoxicity profile towards AML. Furthermore, limited killing of normal monocytes and CD123‐low‐expressing endothelial cells was noted, thus indicating a low toxicity profile of the anti‐CD123.CAR. Taken together, our results indicate that CD123‐specific CARs strongly enhance anti‐AML CIK functions, while sparing HSPCs and normal low‐expressing antigen cells, paving the way to develop novel immunotherapy approaches for AML treatment.     

Sorafenib plus all‐trans retinoic acid for AML patients with FLT3‐ITD and NPM1 mutations

Knowledge of the molecular basis of acute myeloid leukaemia has increased considerably in the past few years, and therapies targeting specific molecular defects of this disease are intensively investigated. Patients with both NPM1 and FLT3‐ITD mutations encompass 20% of cytogenetically normal AML. The multikinase and FLT3 inhibitor, sorafenib, has shown some efficacy in patients with relapsed FLT3‐ITD⁺ AML. In addition, it is suggested that all‐trans retinoic acid (ATRA) used in combination with chemotherapy has shown to improve outcome of patients harbouring NPM1 mutations. We report here the clinical course of three patients with refractory or relapsed FLT3‐ITD⁺/NPM1⁺ AML who achieved significant response upon sorafenib and ATRA combination.     

Prognostic impact of FLT3-ITD mutation on NPM1+ acute myeloid leukaemia patients and related molecular mechanisms

The prognosis of acute myeloid leukaemia (AML) patients carrying NPM1 mutations is significantly worse when accompanied by FLT3-ITD mutations. However, accurate quantitative detection of FLT3-ITD mutations remains challenging. To identify a novel biomarker in NPM1⁺FLT3-ITD⁺ AML patients for more accurate stratification, we analysed the differential gene expression between the NPM1⁺FLT3-ITD⁺ and NPM1⁺FLT3-ITD⁻ groups in five public AML datasets and identified a biomarker by taking the intersection of differentially expressed genes. We validated this biomarker in bone marrow samples from NPM1⁺ AML patients at the Peking University Institute of Haematology and analysed its prognostic significance. BCAT1 expression was higher in the NPM1⁺FLT3-ITD⁺ group than in the NPM1⁺FLT3-ITD⁻ group in all seven cohorts. BCAT1 was able to predict the prognosis of NPM1⁺FLT3-ITD⁺ AML patients, and its predictive ability was superior to that of the FLT3-ITD allelic ratio (AR). FLT3-targeted inhibitor quizartinib reduced BCAT1 expression. BCAT1 knockdown using lentiviral vectors led to the downregulation of MYC expression. Thus, we identified BCAT1 as a novel biomarker for NPM1⁺FLT3-ITD⁺ AML patients. The FLT3-ITD/BCAT1/MYC signalling pathway may play a biological role in promoting the occurrence and development of AML in FLT3-ITD⁺ cell lines.     

Identification of emerging FLT3 ITD‐positive clones during clinical remission and kinetics of disease relapse in acute myeloid leukaemia with mutated nucleophosmin

FLT3 internal tandem duplication (ITD) mutations are frequently detected at diagnosis in cytogenetically normal acute myeloid leukaemia (CN‐AML) and predict unfavourable outcome. FLT3 ITD is an unstable aberration and may be lost or acquired at relapse. Recent whole genome sequencing studies have suggested that FLT3 ITD⁺ve AML relapse may evolve from small subclones undetectable at diagnosis by routine polymerase chain reaction (PCR). We developed a patient‐specific real‐time quantitative‐PCR (RQ‐PCR) to implement FLT3 ITD detection in six AML patients whose blasts carried wild‐type FLT3 at diagnosis and who relapsed with FLT3 ITD by routine PCR. Patient‐specific forward primers were designed after cloning and sequencing the FLT3 ITD in each case. The assay allowed retrospective detection of FLT3 ITD in diagnostic samples of 4/6 cases and to establish the kinetics of clonal evolution preceding relapse. After conventional chemotherapy, all patients had early relapse despite having been classified as NPM1⁺ve/FLT3 ITD^?ve at presentation, with shorter remissions being observed in four patients re‐classified as FLT3 ITD⁺ve by the new assay. Notably, FLT3 ITD clone became detectable by conventional PCR in three patients tested during remission after initial treatment. Our data underscore the need of identifying low FLT3 ITD levels, which are probably associated with relapse in otherwise good prognosis CN‐AML.     

T‐cell replete haploidentical stem cell transplantation attenuates the prognostic impact of FLT3‐ITD in acute myeloid leukemia: A report from the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation

Acute myeloid leukemia (AML) patients harboring the FLT3‐ITD mutation are considered a high risk patient subset preferentially allocated for allogeneic stem cell transplantation in first remission. Whether FLT3‐ITD retains a prognostic role in haploidentical stem cell transplantation (haplo‐SCT) is unknown. To analyze the prognostic impact of FLT3‐ITD in haplo‐SCT, we performed a retrospective analysis of the multicenter registry of the acute leukemia working party of the European Society for Blood and Marrow Transplantation. We included all adult AML patients with known FLT3 status who underwent a first T‐cell replete related haplo‐HCT in first complete remission from 2005 to 2016. We evaluated 293 patients of whom 202 were FLT3^wt and 91 were FLT3‐ITD mutated. FLT3‐ITD patients were more likely to be NPM1 mutated as well as be in the intermediate risk cytogenetic risk category. In multivariate analysis, patients with FLT3‐ITD had comparable rates of relapse incidence [Hazard ratio (HR) = 1.34, confidence interval (CI) 95%, 0.67‐2.7; P = .9] and leukemia‐free survival (HR = 0.99, CI 95%, 0.62‐1.57; P = .9) to those of FLT3^wt patients. Overall survival, the incidence of nonrelapse mortality, and graft versus host disease‐free/relapse‐free survival were not significantly impacted by FLT3‐ITD status. Furthermore, relapse and overall survival were comparable between FLT3‐ITD patients transplanted from various donor pools, namely matched siblings, unrelated donors, haplo‐SCT). Finally, subset analysis of patients with intermediate risk cytogenetics confirmed the absence of a prognostic impact of FLT3‐ITD also for this patient segment. In AML patients undergoing T‐cell replete haplo‐SCT, the FLT3‐ITD mutation possibly does not retain its prognostic significance.     

Abnormal expression of apoptosis-related genes in haematological malignancies: overexpression of MYC is poor prognostic sign in mantle cell lymphoma

The expression of apoptosis-related genes BCL2, BAX, BCL2L1, BCL2A1, MCL1, DAPK1 and MYC was studied by quantitative real-time polymerase chain reaction on total RNA samples from patients with acute lymphoblastic leukaemia (ALL, n = 16), acute myeloid leukaemia (AML, n = 27), chronic myeloid leukaemia (CML, n = 12), mantle cell lymphoma (MCL, n = 19) and chronic lymphoid leukaemia (CLL, n = 32). BCL2, BAX, BCL2A1, MCL1, DAPK1 and MYC were overexpressed in all patient groups. BCL2L1 was underexpressed in CLL and CML, but not in AML, ALL and MCL. MCL1 levels were significantly higher in CD13 and CD33-positive ALL, and in CD56-positive AML samples. BCL2, BCL2L1, BCL2A1 and MCL1 were overexpressed and DAPK1 was underexpressed in CLL samples with a 11q23 deletion. MYC overexpression was significantly associated with shorter overall survival in MCL (P < 0.01). AML patients with a normal karyotype showed a higher frequency of BCL2A1 overexpression (P < 0.001) than those with an abnormal karyotype.     

Acute myeloid leukaemia with myelodysplastic features in children: a report of Japanese Paediatric Leukaemia/Lymphoma Study Group

The clinical characteristics and prognostic relevance of acute myeloid leukaemia (AML) with myelodysplastic features remains to be clarified in children. We prospectively examined 443 newly diagnosed patients in a multicentre clinical trial for paediatric de novo AML, and found ‘AML with myelodysplasia‐related changes’ (AML‐MRC) according to the 2008 World Health Organization classification in 93 (21·0%), in whom 59 were diagnosed from myelodysplasia‐related cytogenetics alone, 28 from multilineage dysplasia alone and six from a combination of both. Compared with 111 patients with ‘AML, not otherwise specified’ (AML‐NOS), patients with ‘AML‐MRC’ presented at a younger age, with a lower white blood cell count, higher incidence of 20–30% bone marrow blasts, unfavourable cytogenetics and a lower frequency of Fms‐like tyrosine kinase 3 internal tandem duplication (FLT3‐ITD), NPM1 and CEBPA mutations. Complete remission rate and 3‐year probability of event‐free survival were significantly worse in ‘AML‐MRC’ patients (67·7 vs. 85·6%, P < 0·01, 37·1% vs. 53·8%, P = 0·02, respectively), but 3‐year overall survival and relapse‐free survival were comparable with ‘AML‐NOS’ patients. By multivariate analysis, FLT3‐ITD was solely associated with worse overall survival. These results support the distinctive features of the category ‘AML‐MRC’ even in children.     

Expression of the leucocyte common antigen (LCA,CD45) isoforms RA and RO in acute haematological malignancies: possible relevance in the definition of new overlap points between normal and leukaemic haemopoiesis

Summary. The membrane expression of CD45RA and CD45RO on fresh leukaemic cells taken from 529 cases of acute haemopoietic malignancies, including 117 B-origin acute lymphoblastic leukaemia (B-origin ALL), 3 7 T-origin acute lymphoblastic leukaemia (T-origin ALL), 297 de novo acute myeloid leukaemia (AML), 42 refractory anaemia with excess of blasts in transformation (RAEB-T) and 36 myeloid blastic phase of chronic myelogenous leukaemia (CML-BP-my), was analysed. B-origin ALLs were characterized by the lack of the RO isoform along with the consistent presence of RA. Conversely, a differential expression of the two isoforms was detected in different subsets of T-origin ALL, in that T-stem cell leukaemias (T-SCL: CD7⁺, CD4^?, CD8^?, CD1^?) preferentially expressed CD45RA whereas conventional T-acute lymphoblastic leukaemias (T-ALL: CD7⁺, CD4⁺ and/or CD8⁺ and/or CD1+) were consistently marked by CD45RO. Within myeloid malignancies, most of AMLs displayed CD45RA, while a substantial group of CML-BP-my preferentially exhibited CD45RO. As a general rule, a reciprocal exclusion of the two isoforms was observed in AML as well as in ALL. Nevertheless, a frequent coexpression of CD45RA and CD45RO was observed in CD14⁺ AML. In vitro treatment with all-trans retinoic acid (ATRA) was able to promote a switch from CD45RA to CD45RO expression in 2 7 de novo AML, independently from morphological subtyping. To our knowledge, this is the first report on CD45 isoform expression in a large series of patients with acute leukaemia. The knowledge of the differential expression of CD45RA and CD45RO can ameliorate our classificative approach to haematological malignancies, as well as disclose new multiple overlap points between normal and leukaemic cell differentiation.     

Synergistic antitumor effects of lenalidomide and rituximab on mantle cell lymphoma in vitro and in vivo

Rituximab (RTX), a chimeric anti‐CD20 antibody, is associated with direct induction of apoptosis and antibody‐dependent cell‐mediated cytotoxicity (ADCC) with clinical efficacy in mantle cell lymphoma (MCL). Lenalidomide (LEN), a novel immunomodulatory agent, sensitizes tumor cells and enhances ADCC. Our study attempted to elucidate the mechanism of LEN‐enhanced RTX‐mediated cytotoxicity of MCL cells. We found that LEN and RTX induced growth inhibition of both cultured and fresh primary MCL cells. LEN enhanced RTX‐induced apoptosis via upregulating phosphorylation of c‐Jun N‐terminal protein kinases (JNK), Bcl‐2, Bad; increasing release of cytochrome‐c; enhancing activation of caspase‐3, ‐8, ‐9 and cleavage of PARP. Meanwhile, LEN activated NK cells and increased CD16 expression on CD56^lowCD16⁺ NK cells. Whole PBMCs but not NK cell‐depleted PBMCs treated with LEN augmented 30% of RTX‐dependent cytotoxicity. Daily treatment with LEN increased NK cells by 10‐folds in SCID mice, and combination of LEN and RTX decreased tumor burden and prolonged survival of MCL‐bearing SCID mice. Taken together, our study demonstrates that LEN plus RTX provides a synergistically therapeutic effect on MCL cells by enhancing apoptosis and RTX‐dependent NK cell‐mediated cytotoxicity and may be an optimal combination in the clinical trial of relapsed or refractory MCL. Am. J. Hematol. 2009. © 2009 Wiley‐Liss, Inc.     

An immunophenotypic pre‐treatment predictor for poor response to induction chemotherapy in older acute myeloid leukaemia patients: blood frequency of CD34+ CD38low blasts

Many older patients with acute myeloid leukaemia (AML) that receive standard intensive chemotherapy fail to achieve complete remission (CR). Upfront identification of patients unlikely to benefit from standard induction chemotherapy would be important for exploration of novel therapies. This study evaluated if a flow cytometric assay measuring pre‐treatment CD34⁺ CD38^low blast frequency could predict therapeutic‐resistance in 736 AML patients entered into the UK National Cancer Research Institute AML16 trial. High peripheral blood CD34⁺ CD38^low blast frequency (>7% of leucocytes), present in 18% of assessable patients, conferred significantly reduced CR rates (38% vs. 76%, P < 0·0001) and poor survival, and was independently prognostic for all endpoints of treatment resistance by multivariate analysis.     

AXL/MERTK inhibitor ONO-7475 potently synergizes with venetoclax and overcomes venetoclax resistance to kill F LT 3-ITD acute myeloid leukemia

FMS-like Tyrosine Kinase 3 (FLT3) mutation is associated with poor survival in acute myeloid leukemia (AML). The specific Anexelekto/MER Tyrosine Kinase (AXL) inhibitor, ONO-7475, kills FLT3-mutant AML cells with targets including Extracellular-signal Regulated Kinase (ERK) and Myeloid Cell Leukemia 1 (MCL1). ERK and MCL1 are known resistance factors for Venetoclax (ABT-199), a popular drug for AML therapy, prompting the investigation of the efficacy of ONO-7475 in combination with ABT-199 in vitro and in vivo. ONO-7475 synergizes with ABT-199 to potently kill FLT3-mutant acute myeloid leukemia cell lines and primary cells. ONO-7475 is effective against ABT-199-resistant cells including cells that overexpress MCL1. Proteomic analyses revealed that ABT-199-resistant cells expressed elevated levels of pro-growth and anti-apoptotic proteins compared to parental cells, and that ONO-7475 reduced the expression of these proteins in both the parental and ABT-199-resistant cells. ONO-7475 treatment significantly extended survival as a single in vivo agent using acute myeloid leukemia cell lines and PDX models. Compared to ONO-7474 monotherapy, the combination of ONO-7475/ABT-199 was even more potent in reducing leukemic burden and prolonging the survival of mice in both model systems. These results suggest that the ONO-7475/ABT-199 combination may be effective for AML therapy.     

Relapse risk and survival in patients with FLT3 mutated acute myeloid leukemia undergoing stem cell transplantation

In patients with AML with FMS‐like tyrosine kinase 3 (FLT3) mutations, the significance of minimal residual disease (MRD) detected by PCR before allogeneic stem cell transplantation (SCT) on outcomes after transplant remains unclear. We identified 200 patients with FLT3‐AML who underwent SCT at our institution. Disease status at transplant was: first or second complete remission (CR1/CR2, n = 119), high‐risk CR (third or subsequent CR, marrow hypoplasia, or incomplete count recovery) (CR‐HR, n = 31), and morphological evidence of active disease (AD, n = 50). The median follow‐up was 27 months, and the 2‐year overall and progression‐free survival were 43% and 41%, respectively. Relapse was highest in the AD group (85%) and the CR‐HR FLT3 MRD positive group (72%), followed by CR‐HR FLT3 MRD negative (58%), CR1/CR2 FLT3 MRD positive (39%), and lowest in the CR1/CR2 FLT3 MRD negative group (23%). On multivariate analysis, independent factors influencing the risk of relapse were detectable morphological disease and FLT3 MRD by PCR pre‐transplant. Factors that did not influence the relapse risk included: age, graft type, graft source, type of FLT3 mutation, or conditioning intensity. Morphologic and molecular remission status at the time of transplant were key predictors of disease relapse and survival in patients with FLT3‐AML.     

Comparative quantitative expression of bcl-2 by normal and leukaemic myeloid cells

Summary. Expression of the bcl-2 oncoprotein by AML blasts has previously been demonstrated to be heterogenous with high levels of bcl-2 expression being associated with a low complete remission rate and poor survival. We have quantified bcl-2 expression in AML blasts in relation to expression of the CD34 antigen and in comparison to CD34-positive cells from normal bone marrow. When expressed as molecules of equivalent soluble fluorochrome (MESF) per cell, AML blast cell bcl-2 expression varied from 11.1 to 99.9 × lO³ (median 39.4 × 10³, n=56) with 28.5% of patients expressing high MESF values (>50 × 10³) and 16% of patients expressing low MESF values (<20 × 10³), the remainder expressing intermediate values. There was no significant difference between intensity of bcl-2 expression and FAB classification in the de novo AML cases, and there was no significant differences between de novo and secondary AML cases. Blasts from CD34⁺ AML patients expressed significantly higher levels of bcl-2 (mean MESF 43.6 × 10³, n =36) than CD34^? AML patients (mean MESF 31.7 × 10³, n=19). In five cases of CD34⁺ AML, bcl-2 expression was determined on purified CD34⁺ and CD34^? blast cell populations. In all cases CD34⁺ blasts were found to express significantly higher bcl-2 MESF values compared to the CD34^? fraction. Purified CD34⁺ cells from normal bone marrow consistently expressed high levels of bcl-2 (MESF >75 × 10³ n = 4), which was comparable to that found on CD34⁺ AML cells. Our results suggest that the poor prognosis previously associated with AML blasts expressing the CD34 antigen may in part be related to high expression of bcl-2. Also the ability to measure bcl-2 in AML blasts quantitatively by flow cytometry and to categorize patients into discrete groups may be of value as a prognostic indicator in AML.     

hMICL and CD123 in combination with a CD45/CD34/CD117 backbone – a universal marker combination for the detection of minimal residual disease in acute myeloid leukaemia

Real‐time quantitative polymerase chain reaction (qPCR) has been extensively validated for the detection of minimal residual disease (MRD) in acute myeloid leukaemia (AML). Meanwhile, multicolour flow cytometry (MFC) has received less attention because the so‐called leukaemia‐associated immunophenotypes (LAIPs) are generally of lower sensitivity and specificity, and prone to change during therapy. To improve MRD assessment by MFC, we here evaluate the combination of human Myeloid Inhibitory C‐type Lectin (hMICL, also termed C‐type lectin domain family 12, member A, CLEC12A) and CD 123 (also termed interleukin‐3 receptor alpha, IL3RA) in combination with CD34 and CD117 (KIT), as an MRD assay in pre‐clinical and clinical testing in 69 AML patients. Spiking experiments revealed that the assay could detect MRD down to 10^?4 in normal bone marrow with sensitivities equalling those of validated qPCR assays. Moreover, it provided at least one MFC MRD marker in 62/69 patients (90%). High levels of hMICL/CD123 LAIPs at the post‐induction time‐point were a strong prognostic marker for relapse in patients in haematological complete remission (P < 0·001). Finally, in post induction samples, hMICL/CD123 LAIPs were strongly correlated (r = 0·676, P = 0·0008) to applied qPCR targets. We conclude the hMICL/CD123‐based MFC assay is a promising MRD tool in AML.     

Variability in responsiveness to lovastatin of the primitive CD34+ AML subfraction compared to normal CD34+ cells

In the present study, we questioned whether the cholesterol synthesis inhibitor lovastatin potentiates the cytotoxicity of chemotherapeutic agents in the primitive CD34⁺ subpopulation of acute myeloid leukemia (AML) cells. AML mononuclear cells (n = 17) were sorted in CD34⁺ and CD34⁻ fractions and compared to normal CD34^+/− cells (n = 7). The percentage of surviving cells upon exposure to lovastatin (25–100 μM) and/or chemotherapeutics (cytarabin or daunorubicin) was determined with a luminescent cell viability assay. The results demonstrate that the primitive CD34⁺ subpopulation of normal and AML cells displayed a higher sensitivity to lovastatin than the more mature CD34⁻ subpopulation. The combination of lovastatin and chemotherapeutics resulted in a more pronounced inhibitory effect on both subpopulations. In contrast to the homogeneous results in normal CD34⁺ cells, a distinct heterogeneity in lovastatin sensitivity was found in AML samples. Therefore, a group of normal (n = 11) and abnormal (n = 6) responders were identified based on a reduced or increased cell survival compared to normal CD34⁺ cells. This distinction was not only observed in the CD34⁺ AML subfraction but also in CD34⁺CD38⁻AML cells. In the abnormal responder group, 50% of patients presented with unfavorable cytogenetics and significant higher peripheral blast cell counts, which coincided with poor treatment results. In summary, the findings indicate that the primitive subfraction of CD34⁺ AML cells is in the majority of cases affected by lovastatin treatment, which is potentiated when combined with chemotherapeutics. Heterogeneity of the response observed in AML patients allowed identification of a subgroup with poor prognosis.     

Differential antigen expression and aberrant signaling via PI3/AKT,MAP/ERK,JAK/STAT,and Wnt/β catenin pathways in Lin−/CD38−/CD34+ cells in acute myeloid leukemia

Acute myeloid leukemia is often called as stem cell disease that presents with treatment failure and poor disease outcome. Leukemic stem cells in acute myeloid leukemia (AML) are enriched in Lineage‐/CD38?/CD34+ compartment of CD34?positive AML. Many markers important for stem cell biology have been reported for their association with leukemic stem cell population, but what remains clinically most important is a rapid identification of prognostic information. In this study, we evaluated four signal transduction pathways and thirteen markers on Lin?/CD38?/CD34+ population in AML. Expressions were compared in different AML subtypes, survival, and treatment outcome groups. We observed that markers important in homing, cell quiescence, and signal propagation such as CD44, CD96, CD90, WT‐1, CD123 and CD25 were most significantly differentially expressed on Lin‐/CD38?/CD34+ population in AML from their normal counterparts (P < 0.05, Mann–Whitney). Constitutive activation of phospho ERK, AKT, and STAT5 in these cells was associated with poor outcome. Also, an increased frequency of putative leukemic stem cell population shows negative impact on treatment outcome and overall survival, suggesting that initial evaluation of AML samples for pLSC frequency and constitutively activated signaling pathway can provide prognostic and therapeutic information at the time of diagnosis.     

The genotype nucleophosmin mutated and FLT3‐ITD negative is characterized by high bax/bcl‐2 ratio and favourable outcome in acute myeloid leukaemia

Nucleophosmin gene (NPM1) mutations characterize acute myeloid leukaemia (AML) with normal karyotype and frequently co‐exist with FLT3 internal tandem duplications (ITD). We evaluated bcl‐2, bax, NPM1 and FLT3‐ITD in 222 AML patients. Bax/bcl‐2 ratio >0·35 and NPM1 without FLT3‐ITD were significantly associated (P = 0·0001). NPM1‐mutated (mt)/FLT3‐ITD negative patients showed a higher complete remission (CR) rate (90%, P = 0·0002) and a longer overall survival (OS, P = 0·00007). NPM1‐mt/FLT3‐ITD negative plus bax/bcl‐2 > 0·35 subset showed a very high CR rate (96%), very long OS (P = 0·00005) and disease‐free survival (P = 0·004). The favourable prognosis of NPM1‐mt/FLT3‐ITD negative patients might be explained by a higher bax/bcl‐2 ratio.