Effects of Decitabine on the proliferation of K562 cells and the expression of DR4 gene

Objective

To investigate the role of DR4 gene in the occurrence, development and prognosis of acute myeloid leukemia (AML), find a new regulatory gene of Decitabine for the treatment of AML, namely DR4 gene, and explore the molecular mechanism of AML in the treatment of AML.

急性髓细胞性白血病（AML）中IL-3Rα 基因的表达及意义

目的:检测白介素-3受体α亚基(IL-3Rα)在急性髓细胞性白血病(acute myeloid leukemia,AML)中的表达,研究其在AML发病和预后评估中的意义。方法:应用逆转录-聚合酶链反应(RT-PCR)法检测57例AML患者IL-3Rα的表达,其中包括52例初治和复发患者(6例M1、28例M2、10例M3、3例M4、5例M5)及5例缓解患者,另10例正常人作为对照。结果:①10例正常人和5例AML缓解患者未检测到IL-3Rα表达。②52例初治和复发的AML患者中IL-3Rα阳性表达21例,占40.38%,M1～M5各型患者的阳性率差异无显著性(P>0.05)。初治和复发AML患者之间IL-3Rα阳性表达无明显差异(P>0.05)。③AML患者的IL-3Rα阳性表达与外周血白细胞计数、骨髓中原始细胞比例、CD34阳性表达有关(P<0.05)。④AML患者中IL-3Rα阳性表达的患者CR率低于阴性者(P<0.05)。结论:①IL-3Rα在AML发病中有一定作用。②IL-3Rα基因可作为AML预后不良的一个指标。     

Comparative proteome analysis of acute myeloid leukemia with and without maturation

Acute myeloid leukemia (AML) is a severe, rapidly progressing disease triggered by blocking granulocyte or monocyte differentiation and maturation. Because of its heterogeneity, AML is divided into a number of subtypes. Unfortunately, so far very few correlations have been found between AML classification and its clinical course or patient response to treatment. In addition, as yet only a few subtype-specific AML biomarkers have been discovered. To solve these problems here, we focused on two AML subtypes M1 and M2 that are especially difficult to differentiate. Using 2D electrophoresis and mass spectrometry, we analyzed the protein profiles of peripheral blood (PB) and/or bone marrow (BM) samples collected from 38 AML-M1/M2 patients and 17 healthy volunteers. Comparative analysis of AML-M1/M2 and control PB/BM cells revealed 25 proteins that accumulated differentially. Hierarchical clustering of proteomic results clearly divided the AML samples into 2 groups (M1 and M2). Annexin III, L-plastin and 6-phosphogluconate dehydrogenase were found only in the M2 group. We also observed that the levels of annexin I and actin gamma 1 were correlated with resistance to treatment and the time of relapse. It appears that these five proteins can serve as potential AML biomarkers.     

Extramedullary Relapse of the AML Transformed from MDS Following Auto-HSCT: A Case Report

The treatment for AML (Acute myeloid/myelogenous leukemia) transformed from MDS (myelodysplastic syndrome) is difficult and controversial clinically, especially in elder patients. In this case report, we diagnosed a 59-year-old female patient with AML-M2a transformed form MDS which might be caused by her chemotherapy for mastocarcinoma. After achieving complete remission (CR) through combined chemotherapy, autologous peripheral blood stem cell transplantation (auto-PBSCT) was attempted. Following auto-HSCT, marrow showed continuous CR but the patient later developed extramedullary bone-infiltration relapse. Then local radiotherapy has been applied, and the patient now has prolonged survival. This is the first (or a successful) case report of auto-HSCT in an elderly patient with AML transformed from MDS.     

An abnormal CD34+ myeloid/CD34+ lymphoid ratio at the end of chemotherapy predicts relapse in patients with acute myeloid leukemia.

During conventional follow-up of patients with acute myeloid leukemia (AML), the emergence of cytopenias is considered to be a sign of impending relapse, and it represents an example of how leukemic hematopoiesis affects normal hemopoietic differentiation. In the present study, we have explored the possible value of the analysis of the distribution of CD34+ myeloid and CD34+ lymphoid progenitor cells in follow-up complete remission bone marrow samples from de novo AML patients as a prognostic parameter for predicting relapse. A total of 213 bone marrow samples from 36 AML patients in morphological complete remission, obtained at the end of induction, consolidation, and intensification therapy and every six months thereafter were analyzed. The normal CD34+ myeloid/CD34+ lymphoid ratio ranged between 2.4 and 8.9. In contrast, in most AML cases an abnormally high ratio (> or =10) was observed at the end of induction and consolidation therapy: 96% and 75% of cases, respectively. On the other hand, at the end of intensification, 70% of the patients displayed a normal CD34+ ratio. Patients with a myeloid/lymphoid CD34+ ratio higher than 10 at the end of intensification showed a significantly lower overall survival (median survival of 19 months versus median not reached, P = 0.05), as well as a lower disease-free survival (median of 7 months versus 30 months, P = 0.0001). Regarding sequential studies, 67% of the relapses were preceded by the re-appearance of an abnormal CD34 ratio, whereas relapse was not predicted in four patient with leukemia classified as M3 undergoing maintenance therapy. From the remaining 18 patients who are still in continuous complete remission, all except 3 cases (17%) displayed a normal CD34 myeloid/lymphoid ratio. In summary, the present study shows that the persistence at the end of chemotherapy of an abnormally high (> or =10) ratio between CD34+ myeloid and CD34+ lymphoid progenitors in the bone marrow of AML patients is associated with high risk of relapse and a shorter overall survival.     

Treatment of poor-risk myelodysplastic syndromes and acute myeloid leukemia with a combination of 5-azacytidine and valproic acid

5-azacytidine (AZA) has become standard treatment for patients with higher-risk myelodysplastic syndrome (MDS). Response rate is about 50% and response duration is limited. Histone deactylase (HDAC) inhibitors are attractive partners for epigenetic combination therapy. We treated 24 patients with AZA (100?mg/m(2), 5?days) plus valproate (VPA; continuous dosing, trough serum level 80-110?μg/ml). According to WHO classification, 5 patients had MDS, 2 had MDS/MPD, and 17 had acute myeloid leukemia (AML). Seven patients (29%) had previously received intensive chemotherapy, and five had previous HDAC inhibitor treatment. The overall response rate was 37% in the entire cohort but significantly higher (57%) in previously untreated patients, especially those with MDS (64%). Seven (29%) patients achieved CR (29%) and two PR (8%), respectively. Hematological CR was accompanied by complete cytogenetic remission according to conventional cytogenetics in all evaluable cases. Some patients also showed complete remission according to FISH on bone marrow mononuclear cells and CD34(+) peripheral blood cells, as well as by follow-up of somatic mitochondrial DNA mutations. Four additional patients achieved at least marrow remissions. Factors influencing response were AML (vs. MDS), marrow blast count, pretreatment, transfusion dependency, concomitant medication with hydroxyurea, and valproic acid (VPA) serum level. This trial is the first to assess the combination of AZA plus VPA without additional ATRA. A comparatively good CR rate, relatively short time to response, and the influence of VPA serum levels on response suggest that VPA provided substantial additional benefit. However, the importance of HDAC inhibitors in epigenetic combination therapy can only be proven by randomized trials.     

急性髓系白血病ERG、MDR1及BAALC基因的表达水平及临床意义

目的:探讨急性髓系白血病(acutemyeloidleukemia,AML)患者骨髓单个核细胞的ETS相关基因(ETS related gene,ERG)、多药耐药基因1(Multidrug resistance gene 1,MDR1)、脑和急性白血病胞质(Brain and acute leukemia cytoplasmic,BAALC)基因的表达水平及临床意义。方法:选取90例成人AML患者为研究对象,均接受蒽环类药物诱导化疗联合阿糖胞苷等进行初步治疗,采用实时荧光定量检测PCR技术检测治疗后的骨髓单个核细胞ERG、MDR1、BAALC基因表达,并分析其与患者临床特征、危险度分层、治疗疗效、生存率的关系。结果:AML患者ERG、MDR1、BAALC基因均有强表达,三因子表达强弱同白细胞、血小板等临床指标无明显相关性(P0.05),不同危险度分层对应的ERG、MDR1、BAALC表达之间有明显差异,中危组、高危组患者表达强度均高于低危组(P0.05)。ERG、MDR1低表达组对应的疗效CR(93.1%%,89.6%)、OS(56.5%, 66.7%)较高表达组CR(61.4%, 81.3%)、OS(56.5%,66.7%)值更高(P0.05),不同BAALC表达者疗效CR及生存率OS比较差异无统计学意义(P0.05)。结论:AML患者单个核细胞的ERG、MDR1基因表达水平与其危险度分层、疗效和生存率呈负相关,以REG和MDR1同AML关系敏感,BAALC敏感度较低,联合检测REG、MDR1、BAALC基因表达可能提高AML患者危险度分层、疗效及预后判读的准确度。     

Distribution of the cytoskeletal protein,Nestin, in acute leukemia

Abstract

Nestin is a neuroepithelial stem cell marker that is expressed in some types of tumor cells. Recent reports suggest that Nestin may be closely related to malignant cell proliferation and migration. Acute leukemia (AL) is characterized by a lack of differentiation, which results in uncontrolled proliferation in the bone marrow and accumulation of immature cells. The expression and function of Nestin in AL is unclear. We investigated Nestin immunohistochemical patterns of 87 patients that included 47 cases of acute myeloid leukemia (AML) and 40 cases of acute lymphoblastic leukemia (ALL), and 20 patients in complete remission (CR) from AML or ALL. We also investigated the clinico-pathological features of 87 cases of AL and their CR and overall survival (OS). Nestin was expressed in leukemic blasts and mature granulocytic cells in most cases (39/47) of AML. Conversely, Nestin was expressed in mature granulocytic cells in fewer cases (6/40) of ALL, but not in blasts. Nestin expression appeared in leukemic blasts of AML, but not ALL. Nestin expression in AML blast cells was not associated with CR or OS. We provide evidence that Nestin is expressed in AL and might be a useful immunohistochemical marker for identifying AML and ALL.     

Cytogenetic analysis and clinical significance of chromosome 7 aberrations in acute leukaemia

The analysis was performed on bone marrow cells derived from 96 patients with acute leukaemia (AL): 76 with acute myelogenous leukaemia (AML) and 20 with acute lymphoblastic leukaemia (ALL). Aberrations of chromosome 7 were revealed in 20 (21%) of 96 analysed cases: in 14 (18%) with AML and in six (30%) with ALL. Structural aberrations, present in 13 patients (eight with AML and five with ALL), were unbalanced and led to partial monosomy (12 cases) or trisomy (four cases) of chromosome 7. Twelve (86%) out of 14 AML and all the ALL patients with chromosome 7 aberrations had complex karyotypes in their bone marrow cells. Monosomy 7 and 7q losses were frequently observed in the AML group, whereas, in the ALL group, gains in 7q and losses in the short arms constituted most chromosome 7 aberrations. The occurrence of monosomy, or of losses in 7q, results in a worse response to induction therapy in AML patients. The complete remission (CR) rate was significantly lower in this group in comparison to the group of AML patients with a normal karyotype (p = 0.01) in bone marrow cells.     

Detection of minimal residual disease (MRD) after bone marrow transplantation (BMT) by multi-parameter flow cytometry (MPFC)

Multi-parameter flow cytometry (MPFC) was used to detect minimal residual disease (MRD) following bone marrow transplantation (BMT) in 21 patients. Bone marrow (BM) was analyzed pre-transplant and 3–4 months post-BMT while the patients were in clinical and morphological remission. MRD was detected by identifying cells with aberrant antigen expression and/or leukemia-associated phenotype (LAP) using MPFC. Prior to BMT, 8 out of 21 patients exhibited normal antigen expression based on normal BM samples while 13 BM aspirates had abnormal MPFC. Pre-BMT MPFC was abnormal in all 10 patients who were not in complete remission (CR) (>5% blasts in BM) as well as 3 patients acute lymphoblastic leukemia (ALL) who were in CR. In BM from ALL patients, an abnormal uniform B cell population was observed however antigen expression patterns varied greatly between patients. BM from acute myeloblastic leukemia (AML) patients showed an abnormal distribution of CD34+ cells. In addition, a correlation was observed between pre-BMT cytogenetics and MPFC. Only 2 out of 8 (25%) patients with normal MPFC pre-autologous bone marrow transplantation (ABMT) relapsed (AML), while 6 out of 13 (46%) patients with abnormal pre-BMT MPFC relapsed including 2 out of 3 patients who were transplanted in clinical CR. Pre-BMT MPFC may thus be an effective tool for detection of MRD by detection of a pre-transplant MPFC abnormality.     

Hematopoietic stem cell expansion and distinct myeloid developmental abnormalities in a murine model of the AML1-ETO translocation

The t(8;21)(q22;q22) translocation, which fuses the ETO gene on human chromosome 8 with the AML1 gene on chromosome 21 (AML1-ETO), is one of the most frequent cytogenetic abnormalities associated with acute myelogenous leukemia (AML). It is seen in approximately 12 to 15% of AML cases and is present in about 40% of AML cases with a French-American-British classified M2 phenotype. We have generated a murine model of the t(8;21) translocation by retroviral expression of AML1-ETO in purified hematopoietic stem cells (HSC). Animals reconstituted with AML1-ETO-expressing cells recapitulate the hematopoietic developmental abnormalities seen in the bone marrow of human patients with the t(8;21) translocation. Primitive myeloblasts were increased to approximately 10% of bone marrow by 10 months posttransplant. Consistent with this observation was a 50-fold increase in myeloid colony-forming cells in vitro. Accumulation of late-stage metamyelocytes was also observed in bone marrow along with an increase in immature eosinophilic myelocytes that showed abnormal basophilic granulation. HSC numbers in the bone marrow of 10-month-posttransplant animals were 29-fold greater than in transplant-matched control mice, suggesting that AML1-ETO expression overrides the normal genetic control of HSC pool size. In summary, AMLI-ETO-expressing animals recapitulate many (and perhaps all) of the developmental abnormalities seen in human patients with the t(8;21) translocation, although the animals do not develop leukemia or disseminated disease in peripheral tissues like the liver or spleen. This suggests that the principal contribution of AML1-ETO to acute myeloid leukemia is the inhibition of multiple developmental pathways.     

Lytic activity against primary AML cells is stimulated in vitro by an autologous whole cell vaccine expressing IL-2 and CD80

Despite being of the myeloid lineage, acute myeloid leukaemia (AML) blasts are of low immunogenicity, probably because they lack the costimulatory molecule CD80 and secrete immunosuppressive factors. We have previously shown that in vitro stimulation of autologous peripheral blood mononuclear cells (PBMCs) with primary AML cells modified to express CD80 and IL-2 promotes proliferation, secretion of Th1 cytokines and expansion of activated CD8⁺ T cells. In this study, we show that allogeneic effector cells (from a healthy donor or AML patients) when stimulated with IL-2/CD80 modified AML blasts were able to induce the lysis of unmodified AML blasts. Effector cells stimulated with IL-2/CD80AML blasts had higher lytic activity than cells stimulated with AML cells expressing CD80 or IL-2 alone. Similarly, AML patient PBMCs primed with autologous IL-2/CD80 AML cells had a higher frequency of IFN-γ secreting cells and show cytotoxicity against autologous, unmodified blasts. Crucially, the response appears to be leukaemia specific, since stimulated patient PBMCs show higher frequencies of IFN-γ secreting effector cells in response to AML blasts than to remission bone marrow cells from the same patients. Although studied in a small number of heterogeneous patient samples, the data are encouraging and support the continuing development of vaccination for poor prognosis AML patients with autologous cells genetically modified to express IL-2/CD80.     

Cytogenetic studies in bone marrow transplant recipients

Chromosome studies were performed in 24 patients who underwent allogeneic bone marrow transplantation (BMT) for severe aplastic anaemia (8), chronic myeloid leukemia (5 in chronic, 2 in accelerated phase and 1 in lymphoid blast crisis), acute myeloid leukemia (6), acute lymphoblastic leukemia in relapse (1) and Hodgkin's disease (1). Donor-cell type engraftment was demonstrated in 21 patients: in all 17 sex-mismatched transplants and - as demonstrated by reconstitution with Ph-negative cell populations - in 4 CML patients with a sex-matched donor. Recipient-type mitoses were seen in the bone marrow of 5 cases (1 SAA, 3 CML, 1 AML) after transplantation. They were only observed on one occasion in patients with SAA (4 of 25 on day 33) and AML (44 of 50 on day 14). Despite the continued demonstration of some Ph-positive mitoses in 3 patients with CML up to day 28, 323 and 451 after BMT, respectively, all surviving CML patients are still in complete haematological and clinical remission. So far the significance of these cytogenetically abnormal persisting host cells remains unknown.     

流式细胞术动态监测微小残留病灶在急性髓系白血病中的意义*

目的:探讨急性髓系白血病(AML)在治疗过程中应用多参数流式细胞术(MFC)动态微小残留病灶(MRD)的意义。方法:选择2015年1月至2017年2月在我院血液科收治的60例AML患者,在诱导治疗第8天,第21天,每次巩固治疗前1天,结束化疗随访过程中,检测骨髓形态学变化和应用MFC监测患者骨髓MRD,并动态随访。定义未发现异常表型细胞(MRD10~(-4))为阴性,其余为阳性。结果:平均随访时间为11个月(3到16个月),早期MRD(诱导化疗第8天)阴性的患者占58.33%,MRD阳性的患者占41.67%。MRD阴性的患者在诱导治疗接受第21天100%达到CR,MRD阳性的患者80%达到CR。第一次巩固治疗结束后,MRD阴性的患者预后明显较MRD阳性的患者好。结论:动态监测MRD对预测AML患者对治疗的反应和预测复发有重要意义。     

Marrow culture studies in adult acute myeloid leukemia at diagnosis and during complete remission

We used the human placental conditioned medium stimulated single layer agar culture technique to study the in vitro growth of marrow cells from 62 adult patients with acute myeloid leukemia (AML). Bone marrow cells were cultured from 50 patients at the time of initial diagnosis, 19 patients in early remission and 20 patients during their full complete remission. Marrow cultures from untreated patients exhibited heterogeneous growth patterns ranging from complete growth failure to excessive microcluster formation. We classified the growth patterns into 4 groups: (1) Gr I: normal growth, (2) Gr II: no growth, (3) Gr III: decreased growth, (4) Gr IV: excessive growth of microclusters. At presentation, none had Gr I growth; Gr II growth was observed in 23; Gr III in 14 and Gr IV in 13. A predominance of no growth were seen in M1 and M3 subtypes, while Gr IV growth was more commonly observed in M2 or M4 subtype. We were unable to correlate the culture findings with age or white cell count. The present results not only indicated that AML at diagnosis was characterized by abnormal granulopoiesis but also demonstrated that leukemic progenitor cells were heterogeneous with different capacities to express their proliferating potential in vitro. Except few with decreased growth, the growth characteristics generally returned to normal with successful remission induction. Both Gr II and Gr IV growth patterns were not observed either in early remission or during full complete remission.     

Adoptive transfer of PR1 cytotoxic T lymphocytes associated with reduced leukemia burden in a mouse acute myeloid leukemia xenograft model

Background aimsTumor antigen-specific cytotoxic T lymphocytes (CTL) have been used in the treatment of human cancer, including leukemia. Several studies have established PR1 peptide, an HLA-A2.1-restricted peptide derived from proteinase 3 (P3), as a human leukemia-associated antigen. PR1-specific CTL elicited in vitro from healthy donors have been shown to lyse P3-expressing AML cells from patients. We investigated whether PR1-CTL can be adoptively transferred into NOD/SCID mice to eliminate human leukemia cells.MethodsPR1-CTL were generated in bulk culture from peripheral blood mononuclear cells (PBMC) stimulated with autologous dendritic cells. Human acute myeloid leukemia (AML) patient samples were injected and engrafted in murine bone marrow at 2 weeks post-transfer.ResultsFollowing adoptive transfer, bone marrow aspirate from mice that received AML alone had 72–88% blasts in a hypercellular marrow, whereas mice that received AML plus PR1-CTL co-infusion had normal hematopoietic elements and only 3–18% blasts in a hypocellular marrow. The PR1-CTL persisted in the bone marrow and liver and maintained a CD45RA^? CD28⁺ effector phenotype.ConclusionsWe found that adoptive transfer of PR1-CTL generated in vitro is associated with reduced AML cells in NOD/SCID mice. PR1-CTL can migrate to the sites of disease and maintain their capacity to kill the AML cells. The surface phenotype of PR1-CTL was consistent with their trafficking pattern in both vascular and end-organ tissues.     

Expression of PD‐L1 on regulatory B cells in patients with acute myeloid leukaemia and its effect on prognosis

Programmed death‐ligand 1 (PD‐L1) is involved in immunosuppression in variety of tumours. Regulatory B cells (Bregs) are critical immune regulatory cells, and it has been demonstrated that the number of regulatory B cells in patients with acute myeloid leukaemia (AML) is much higher than that in healthy donors (HDs), which is linked to a poor prognosis. This study aimed to determine whether increased expression of PD‐L1, including in Bregs, is associated with a worse prognosis in individuals with AML. The proportion of Bregs, PD‐L1 expression in Bregs and PD‐1 expression in T cells were determined using flow cytometry using patient samples from 21 newly diagnosed AML patients at different stages of treatment and 25 HDs. We confirmed PD‐L1 expression in Bregs, and PD‐1 expression in CD3⁺CD4⁺T cells in bone marrow and peripheral blood samples from AML patients was higher than that in samples from HDs. The complete remission (CR) and progression‐free survival (PFS) of Bregs with high PD‐L1 expression were significantly decreased following induction chemotherapy. PD‐L1 expression is indeed increased in Bregs from individuals with AML, and high PD‐L1 expression is related to a poor prognosis.     

A Novel Application of Furazolidone: Anti-Leukemic Activity in Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is the most common malignant myeloid disorder of progenitor cells in myeloid hematopoiesis and exemplifies a genetically heterogeneous disease. The patients with AML also show a heterogeneous response to therapy. Although all-trans retinoic acid (ATRA) has been successfully introduced to treat acute promyelocytic leukemia (APL), it is rather ineffective in non-APL AML. In our present study, 1200 off-patent marketed drugs and natural compounds that have been approved by the Food and Drug Administration (FDA) were screened for anti-leukemia activity using the retrovirus transduction/transformation assay (RTTA). Furazolidone (FZD) was shown to inhibit bone marrow transformation mediated by several leukemia fusion proteins, including AML1-ETO. Furazolidone has been used in the treatment of certain bacterial and protozoan infections in human and animals for more than sixty years. We investigated the anti-leukemic activity of FZD in a series of AML cells. FZD displayed potent antiproliferative properties at submicromolar concentrations and induced apoptosis in AML cell lines. Importantly, FZD treatment of certain AML cells induced myeloid cell differentiation by morphology and flow cytometry for CD11b expression. Furthermore, FZD treatment resulted in increased stability of tumor suppressor p53 protein in AML cells. Our in vitro results suggest furazolidone as a novel therapeutic strategy in AML patients.     

High expression of lactotriaosylceramide, a differentiation-associated glycosphingolipid, in the bone marrow of acute myeloid leukemia patients

Glycosphingolipids (GSLs) are information-bearing biomolecules that play critical roles in embryonic development, signal transduction and carcinogenesis. Previous studies indicate that certain GSLs are associated with differentiation in acute myeloid leukemia (AML) cells. In this study, we collected bone marrow samples from healthy donors and AML patients and analyzed the GSL expression profiles comprehensively using electrospray ionization linear ion-trap mass spectrometry. The results showed that AML patients had higher expression of the GSL lactotriaosylceramide (Lc3), GM3 and neolactotetraosylceramide (nLc4) in their bone marrow than did the healthy donors (P < 0.05), especially the M1 subtype of AML. To further explore the molecular mechanisms of Lc3, we examined the expression of the Lc3 synthase β1,3-N-acetylglucosaminyltransferase5 (β3Gn-T5) and found that the bone marrow samples of AML patients had 16-fold higher expression of β3Gn-T5 than those of healthy donors (P < 0.05). Our results suggest that AML-associated GSLs Lc3, GM3 and nLc4 are possibly involved in initiation and differentiation of AML.     

Identification of Novel Genomic Aberrations in AML-M5 in a Level of Array CGH

To assess the possible existence of unbalanced chromosomal abnormalities and delineate the characterization of copy number alterations (CNAs) of acute myeloid leukemia-M5 (AML-M5), R-banding karyotype, oligonucelotide array CGH and FISH were performed in 24 patients with AML-M5. A total of 117 CNAs with size ranging from 0.004 to 146.263 Mb was recognized in 12 of 24 cases, involving all chromosomes other than chromosome 1, 4, X and Y. Cryptic CNAs with size less than 5 Mb accounted for 59.8% of all the CNAs. 12 recurrent chromosomal alterations were mapped. Seven out of them were described in the previous AML studies and five were new candidate AML-M5 associated CNAs, including gains of 3q26.2-qter and 13q31.3 as well as losses of 2q24.2, 8p12 and 14q32. Amplication of 3q26.2-qter was the sole large recurrent chromosomal anomaly and the pathogenic mechanism in AML-M5 was possibly different from the classical recurrent 3q21q26 abnormality in AML. As a tumor suppressor gene, FOXN3, was singled out from the small recurrent CNA of 14q32, however, it is proved that deletion of FOXN3 is a common marker of myeloid leukemia rather than a specific marker for AML-M5 subtype. Moreover, the concurrent amplication of MLL and deletion of CDKN2A were noted and it might be associated with AML-M5. The number of CNA did not show a significant association with clinico-biological parameters and CR number of the 22 patients received chemotherapy. This study provided the evidence that array CGH served as a complementary platform for routine cytogenetic analysis to identify those cryptic alterations in the patients with AML-M5. As a subtype of AML, AML-M5 carries both common recurrent CNAs and unique CNAs, which may harbor novel oncogenes or tumor suppressor genes. Clarifying the role of these genes will contribute to the understanding of leukemogenic network of AML-M5.