HLA-DR antigen-negative acute myeloid leukemia.

Human leukocyte antigen (HLA) Class II antigens are variably expressed on acute myeloid leukemia (AML) blasts. The biological and clinical significance of HLA Class II antigen expression by AML cells is not known. Therefore, we sought to characterize cases of AML without detectable HLA-DR expression. Samples from 248 consecutive adult AML patients were immunophenotyped by multiparameter flow cytometry at diagnosis. HLA-DR antigens were not detected on AML cells from 43 patients, including 20 with acute promyelocytic leukemia (APL), and 23 with other subtypes of AML. All APL cases had t(15;17), but there were no characteristic chromosome abnormalities in non-APL cases. No direct expression of other antigens was identified in HLA-DR-negative APL and non-APL cases. Interestingly, cells from three HLA-DR-negative non-APL patients had similar morphology to that of the hypogranular variant of APL. This morphology, however, was not present in any HLA-DR-positive AML cases. Treatment response was similar in the 23 HLA-DR-negative non-APL and the 205 HLA-DR-positive patients. Finally, relapse was infrequently associated with changes in HLA-DR antigen expression, as the HLA-DR antigen was lost at relapse in only 4% of HLA-DR-positive cases, and was gained at relapse in only 17% of HLA-DR-negative cases. We conclude that HLA-DR-negative AML includes approximately equal numbers of APL and non-APL cases, and that the morphology of HLA-DR-negative non-APL cases can mimic the hypogranular variant of APL. The diagnosis of APL cannot be based on morphology and lack of HLA-DR antigen expression; rather, it requires cytogenetic or molecular confirmation.     

Acute promyelocytic leukemia: where did we start,where are we now,and the future

Historically, acute promyelocytic leukemia (APL) was considered to be one of the most fatal forms of acute leukemia with poor outcomes before the introduction of the vitamin A derivative all-trans retinoic acid (ATRA). With considerable advances in therapy, including the introduction of ATRA initially as a single agent and then in combination with anthracyclines, and more recently by development of arsenic trioxide (ATO)-containing regimens, APL is now characterized by complete remission rates of 90% and cure rates of ∼80%, even higher among low-risk patients. Furthermore, with ATRA–ATO combinations, chemotherapy may safely be omitted in low-risk patients. The disease is now considered to be the most curable subtype of acute myeloid leukemia (AML) in adults. Nevertheless, APL remains associated with a significant incidence of early death related to the characteristic bleeding diathesis. Early death, rather than resistant disease so common in all other subtypes of AML, has emerged as the major cause of treatment failure.     

Induction of differentiation and apoptosis- a possible strategy in the treatment of adult acute myelogenous leukemia

A differentiation block with accumulation of immature myeloid cells characterizes acute myelogenous leukemia (AML). However, native AML cells often show some morphological signs of differentiation that allow a classification into different subsets, and further differentiation may be induced by exposure to various soluble mediators, e.g., all trans-retinoic acid (ATRA) and several cytokines. Combination therapy with ATRA and chemotherapy should now be regarded as the standard treatment for the acute promyelocytic leukemia variant of AML. Several agents can induce leukemic cell differentiation for other AML subtypes, although these effects differ between patients. Differentiation may then be associated with induction of apoptosis, and differentiation-inducing therapy may therefore become useful in combination with intensive chemotherapy to increase the susceptibility of AML blasts to drug-induced apoptosis. However, it should be emphasized that differentiation and apoptosis can occur as separate events with different regulation in AML cells, and future studies in AML should therefore focus on: A) the identification of new agents with more predictable effects on differentiation and apoptosis; B) the use of clinical and laboratory parameters to define new subsets of AML patients in which differentiation/apoptosis induction has a predictable and beneficial effect, and C) further characterization of how AML blast sensitivity to drug-induced apoptosis is modulated by differentiation induction.     

Acute promyelocytic leukemia

Opinion statement The treatment of acute promyelocytic leukemia (APL) is different from other subtypes of acute myelocytic leukemia (AML). All trans-retinoic acid (ATRA) is an essential component of the standard remission induction for all newly diagnosed APL patients. Remission induction with ATRA and chemotherapy given concurrently appears to be associated with fewer relapses. With further consolidation chemotherapy without high-dose cytosine arabinoside, the disease-free survival rate can reach 70% to 80%, and many of these patients are cured, more so than in any other AML subtype. APL is especially sensitive to anthracyclines, which should be included in the chemotherapy cycles at a higher dose than in other AML subtypes. Maintenance with low-dose chemotherapy (oral daily 6-mercaptopurine with weekly methotrexate) or ATRA further improves the long-term outcome. New approaches are also available for relapsing patients, although the optimal treatment is unknown. Patients who did not receive oral ATRA in first relapse can be treated with this agent, as can first relapsing patients who have been off the drug for more than 1 year. Because of poor remission rates, ATRA should not be used in patients with second or subsequent relapses (whether ATRA was given in the past), in patients with relapses early after ATRA discontinuation, or in patients relapsing while on ATRA therapy. Arsenic trioxide can also be used, especially in patients resistant to ATRA. Because arsenic trioxide is still experimental and not yet widely available, patients who are unlikely to respond to ATRA or who unsuccessfully undergo ATRA reinduction should be treated with chemotherapy. Patients in second or subsequent remission induced with ATRA or chemotherapy should receive consolidation chemotherapy. When arsenic trioxide is used for reinduction, the drug should be continued for several cycles; however, adding consolidation chemotherapy might improve the results. Because it is unknown whether APL in second or subsequent remission is curable with salvage therapy, allogeneic hematopoietic stem cell transplantation is often considered for patients with a human leukocyte antigen (HL-A)-identical sibling and autologous transplantation when a donor does not exist. However, compared with the new treatments, the role of transplantation for relapse is unclear. In first remission, there is no role for transplantation. A new liposomal formulation of intravenous ATRA is being investigated and seems effective in late first relapses, and it may be able to induce and maintain first remissions in selected patients without chemotherapy.     

In vivo use of all-trans retinoic acid prior to induction chemotherapy improves complete remission rate and increases rhodamine 123 uptake in patients with de novo acute myeloid leukemia

All-trans retinoic acid (ATRA) is used in the treatment of acute promyelocytic leukemia. Because ATRA has effects (increase in apoptosis, suppression of bcl-2), it has also been used for the treatment of other French-American-British (FAB) subtypes of acute myelogenous leukemia (AML). To find out the in vivo and in vitro effects of ATRA in AML, we analyzed 37 patients with de novo AML. Twenty-seven patients received ATRA before remission-induction (RI) treatment (ATRA group). Results were compared to a control group (10 patients) that received induction without ATRA during the same time period. Bone marrow or peripheral blood samples were collected from all patients on d 0 and 4. The immunphenotype, myeloperoxidase (MPO), reaction and the efflux uptake of rhodamine 123 (Rh123) were analyzed on myeloblasts in these samples. In the myeloblasts from patients treated with ATRA, the uptake of Rh123 was increased significantly (p=0.026) from d 0 to d 4, and all other parameters remained unaltered. ATRA administration increased the complete remission (CR) rate (88%, 22/25 vs 55%, 5/9) significantly (p=0.042). Logistic regression analysis revealed that ATRA administration was the important factor in CR, among other potential factors including age, white blood count, bcl-2 expression, and the uptake and efflux of Rh123 (p=0.05). Estimated disease-free survival and overall survival were similar between these two groups (43% vs 37.5% and 51.2% vs 37.5%, respectively). In conclusion, ATRA treatment prior to RI treatment may improve the CR rate in patients with de novo AML, which seems to be related to its beneficial effect on multidrug resistance.     

Traitement des leucémies aiguës promyélocytaires de l’adulte

Acute promyelocytic leukaemia (APL) accounts for approximately 8% of all acute myeloid leukemias (AML) in adults. APL is characterized by a t(15;17) chromosomal translocation, a strong tendency for bleeding at diagnosis, and a sensitivity to those agents that induce a differentiation of leukaemia blasts such as all-trans retinoic acid (ATRA) or arsenic. Combined ATRA and chemotherapy have significantly improved patient outcome, with a rate of relapse that has been reduced to approximately 15%. Today, arsenic is being used successfully to treat relapsed patients; its high efficacy allows undertaking haematopoietic stem cell transplantation and achieving second full remission. Other new agents also exhibit high efficacy in APL, which makes this type of AML, considered fearsome for long, a subset for which there are a lot of therapeutic options and satisfactory therapeutic outcome.     

Epidermal growth factor receptor expression in acute myelogenous leukaemia is associated with clinical prognosis

The epidermal growth factor receptor (EGFR) family belongs to type I receptor tyrosine kinases. Overexpression or mutation of EGFR/ErbB1 gene has been detected in a large number of human solid tumours. According to some previous report, this gene is not expressed in hematological malignancies. However, two recent clinical case reports showed that erlotinib caused complete remission of acute myeloid leukaemia (AML)-M1 in patients who had both AML-M1 and non-small-cell lung cancer. These results are supported by preclinical studies in which EGFR tyrosine kinase inhibitors have anti-proliferative effects on AML. These findings prompted us to determine whether EGFR is expressed in human AML, through a large-scale screening of both leukaemic cell lines and clinical samples. Our results show that EGFR is expressed by about 33% of human AML (containing M1 to M7 subtypes) and by some human leukaemia cell lines (K562, MEG-01, CEM and SKO-007). Its expression is not limited to certain AML types but has been detected in many leukaemic cells. In addition, EGFR expression was intimately associated with the poor clinical outcomes. Finally, we find that only EGFR-positive leukaemic cells respond to antibody-dependent cellular cytotoxicity of cetuximab, the monoclonal antibodies against EGFR.     

Treatment results of childhood acute myeloid leukemia in Serbia

We present the results of a retrospective study of acute myeloid leukemia (AML) treatment in Serbia in the period 2000–2012. Treatment was performed in four centers, two of which were located in Belgrade, one in Nis, and one in Novi Sad. Children affected by non-acute promyelocytic leukemia (non-APL) subtypes received treatment regimens derived from protocols of Berlin Frankfurt Munster AML study group, whereas children with APL were treated according to the AIEOP/GIMEMA ATRA plus Idarubicin “AIDA” protocol. Total number of patients was 106, out of whom 48 were girls (45.3 %) and 58 boys (54.7 %); median age at diagnosis was 9.0 years (range 1 month–17.9 years). In total, 82.1 % of patients achieved complete remission after induction treatment. Twelve patients (11.3 %) died during induction, before achieving complete remission; there were nine deaths during remission (10.5 %) and 20 patients relapsed (23.2 %). Median time of follow-up was 54 months. Two patients (1.9 %) were lost to follow-up. Event-free survival was 50.3 % and overall survival was 58.9 %.     

The granulocyte-associated transcription factor Krüppel-like factor 5 is silenced by hypermethylation in acute myeloid leukemia

Krüppel-like factor 5 (KLF5) has been implicated as a tumor suppressor in various solid tumors such as breast and prostate, and recent studies have demonstrated a role for this protein in neutrophil differentiation of acute promyelocytic leukemia cells in response to ATRA. Here, we show that KLF5 expression increases during primary granulocyte differentiation and that expression of KLF5 is a requirement for granulocyte differentiation of 32D cells. In AML, we show that KLF5 mRNA expression levels are reduced in multiple French-American-British subtypes compared to normal controls, and also in leukemic stem cells relative to normal hematopoietic stem cells. We demonstrate that in selected AML cases, reduced expression is associated with hypermethylation of the KLF5 locus in the proximal promoter and/or intron 1, suggesting that this may represent a Class II genetic lesion in the development of AML.     

All-Trans Retinoic Acid (ATRA) and the Regulation of Adhesion Molecules in Acute Myeloid Leukemia

A review of recent information on the expression and the ATRA-driven modulation of cell surface adhesion molecules of acute myelogenous leukemia blast cells is presented. Cytofluorometric studies on fresh blast cells have demonstrated that CD11a, CD11b, CD11c, CD15, CD45RO and CD54 expression is significantly lower in acute promyelocytic leukemia (APL) than in acute myeloid leukemia of other subtypes (AML). In vitro treatment with ATRA dramatically modifies the adhesion phenotype of APL blast cells, promoting a consistently striking up-regulation of CD11b, CD11c, CD 15, CD65, CD54 and CD38. Which is, in general, poorly demonstrable in AML. The behaviour of CD15s is variable and fully independent from CD15 and CD65 in induction experiments, suggesting a differential enzymatic regulation within the selectin ligand system. ATRA is capable, in both APL and AML, of producing a switch from the high- (RA) to the low- (RO) molecular weight isoform of CD45. Moreover, treatment with this retinoid exerts a negative regulation of the membrane expression of CD49e, CD58 and CD11a in APL as well as in AML. Of particular interest is the fact that the negative effect on CD11a expression generates an asynchronous phenotype in APL (CD11a-, CD11b+, CD 15+), undetectable on normal maturing myeloid cells.

In the last part of this review the possible implications of adhesion molecule modulation in the pathogenesis of ATRA syndrome are discussed.     

Sensitivity of MLL-rearranged AML cells to all-trans retinoic acid is associated with the level of H3K4me2 in the RARα promoter region

All-trans retinoic acid (ATRA) is well established as differentiation therapy for acute promyelocytic leukemia (APL) in which the PML–RARα (promyelocytic leukemia-retinoic acid receptor α) fusion protein causes blockade of the retinoic acid (RA) pathway; however, in types of acute myeloid leukemia (AML) other than APL, the mechanism of RA pathway inactivation is not fully understood. This study revealed the potential mechanism of high ATRA sensitivity of mixed-lineage leukemia (MLL)-AF9-positive AML compared with MLL-AF4/5q31-positive AML. Treatment with ATRA induced significant myeloid differentiation accompanied by upregulation of RARα, C/EBPα, C/EBPɛ and PU.1 in MLL-AF9-positive but not in MLL-AF4/5q31-positive cells. Combining ATRA with cytarabine had a synergistic antileukemic effect in MLL-AF9-positive cells in vitro. The level of dimethyl histone H3 lysine 4 (H3K4me2) in the RARα gene-promoter region, PU.1 upstream regulatory region (URE) and RUNX1+24/+25 intronic enhancer was higher in MLL-AF9-positive cells than in MLL-AF4-positive cells, and inhibiting lysine-specific demethylase 1, which acts as a histone demethylase inhibitor, reactivated ATRA sensitivity in MLL-AF4-positive cells. These findings suggest that the level of H3K4me2 in the RARα gene-promoter region, PU.1 URE and RUNX1 intronic enhancer is determined by the MLL-fusion partner. Our findings provide insight into the mechanisms of ATRA sensitivity in AML and novel treatment strategies for ATRA-resistant AML.     

Infectious complications in patients with acute promyelocytic leukaemia treated with the AIDA regimen.

Infections represent a frequent complication of chemotherapy used for acute myeloid leukaemia (AML) and are associated with important toxicity frequently leading to treatment discontinuation. Acute promyelocytic leukaemia (APL) is a unique AML subset requiring tailored therapy including all-trans retinoic acid and anthracycline-based chemotherapy. We analysed in this study the incidence and type of infections complicating the clinical course of 89 consecutive APL patients receiving the AIDA protocol at a single institution. A total of 179 febrile episodes were registered during induction and consolidation, 52% of which were of unknown origin. Infections were clinically and microbiologically documented in 10.6 and 37.4% of cases, respectively. Coagulase-negative staphylococci represented the major cause of septicaemia (28%) and were more frequently isolated during induction, whereas viridans group streptococci, the second pathogen most frequently isolated from blood (27%), represented the principal pathogen detected during consolidation and were significantly associated with mucositis. Gram-negative bacteria accounted for 33.3% of all blood isolates. Fungal infections were only occasionally observed. Bloodstream infections in APL patients were compared with those documented in 271 consecutive patients affected by other subtypes of AML. The incidence of total septicaemia episodes, of staphylococcal bacteraemias and of fungaemias was significantly higher in patients with other AMLs. Empirical antibiotic therapy with ceftriaxone plus amikacin was effective in 73% of APL cases, most of the remaining cases being successfully managed by the addition of teicoplanin. One single death apparently related to infectious complication was recorded. Overall, infections led to antileukaemic treatment withdrawal in six patients, five of whom currently remain in haematologic remission for 13-106 months. These results indicate that a particular pattern of infections is observed in APL patients receiving ATRA plus anthracycline-based chemotherapy and that these appear to be effectively counteracted by standard management.     

早幼粒白血病-维甲酸受体α融合蛋白对RIAM基因转录的负调控作用探讨

目的:研究异常转录因子早幼粒白血病-维甲酸受体α(promyelocytic leukemia-retinoic acid receptor alpha,PML-RARα)融合蛋白对RIAM基因的转录调控机制.方法:利用表达谱数据库(GSE1159)比较RIAM基因在急性髓系白血病(acute myeloid leukemia,AML)各亚型中的表达情况,以及RIAM基因和PML-RARα融合蛋白之间的相关性.采用蛋白质印迹法和实时荧光定量-PCR法分别检测PML-RARα融合蛋白和RIAM基因在急性早幼粒细胞白血病(acute promyelocytic leukemia,APL)模式细胞株PR9及APL患者来源的细胞株NB4中的表达情况,以及在全反式维甲酸(all-trans retinoic acid,ATRA)处理前后的RIAM mRNA的表达情况.利用染色质免疫沉淀技术检测细胞内PML-RARα融合蛋白在RIAM基因启动子附近的结合情况.结果:RIAM基因在APL(即M3型AML)中的表达水平明显低于其他AML亚型(M0、M1、M2、M4、M5和M7型)和正常造血细胞(P＜0.05).随着PML-RARα融合蛋白的表达,RIAM基因的表达水平明显降低.ATRA能激活RIAM基因的表达,且PML-RARα融合蛋白的表达能增强ATRA对RIAM基因表达的激活效应.PML-RARα融合蛋白结合在RIAM基因的启动子区域.结论:RIAM基因是PML-RARα融合蛋白的靶基因,PML-RARα融合蛋白通过结合到RIAM基因的启动子区域对其转录进行负调控.     

Myelodysplastic syndrome after acute promyelocytic leukemia: the European APL group experience.

With improved treatment of acute promyelocytic leukemia (APL) by all trans retinoic acid (ATRA) combined to anthracycline-aracytin chemotherapy (CT), a larger number of those patients may be at risk of late complications. Recently, the Rome group reported five cases of myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML, non-APL) occurring during the course of 77 APL patients (6.5%) in complete remission (CR). From 1991 to 1998, we treated 677 newly diagnosed cases of APL, and 617 of them achieved CR with ATRA combined to CT (n=579) or CT alone (n=38); 246 of them received subsequent maintenance CT with 6 mercaptopurine and methotrexate. With a median follow-up of 51 months, 6 patients (0.97%) developed MDS, 13-74 months after the diagnosis of APL. In all six cases, t(15;17) and PML-RARalpha rearrangement were absent at the time of MDS diagnosis, and karyotype mainly showed complex cytogenetic abnormalities involving chromosomes 5 and/or 7, typical of MDS observed after treatment with alkylating agents, although none of the six patients had received such agents for the treatment of APL. Our findings suggest that MDS can indeed be a long-term complication in APL, although probably at lower incidence than that previously reported.     

All-trans retinoic acid and hematopoietic growth factors regulating the growth and differentiation of blast progenitors in acute promyelocytic leukemia

Although acute leukemia is generally thought to be characterized by maturation arrest, it has been shown that differentiation occurs in blast cells of acute myelogenous leukemia (AML) in vitro as well as in vivo, and that morphologically abnormal mature polymorphonuclear neutrophils (PMNs) often seen in patients with AML are possibly derived from spontaneously differentiating leukemic cells. Acute promyelocytic leukemia (APL) is an unique example in which these features of AML are evident in an almost complete form; administration of all-trans retinoic acid (ATRA) induces differentiation of neoplastic cells into mature neutrophils and successfully induce complete remission in most patients. However, PMNs appearing during ATRA treatment are morphologically abnormal, as indicated not only by the presence of Auer rods but also by neutrophil secondary-granule deficiency that is commonly seen in AML. Moreover, ATRA has heterogeneous effects on the growth of blast progenitors in APL in different patients, being inhibitory, stimulatory or ineffective, which might account in part for the leukemia relapse in patients treated with ATRA alone. Hematopoietic growth factors regulate the growth of blast progenitors in APL. Among them, granulocyte colony-stimulating factor (G-CSF) is unique in that it preferentially stimulates clonal growth, but not self-renewal, in many APL cases, and synergistically enhances the differentiation-inducing effect of ATRA when used in combination. Many other compounds also exert such synergistic effects with ATRA, for which a variety of mechanisms have been suggested. It is crucial to precisely elucidate the functions of these molecules governing the growth/differentiation balance of AML blast progenitors and the mechanisms underlying their deregulated differentiation program in order to achieve effective differentiation therapy for patients with AML, not restricted to APL.     

Genome-wide functions of PML-RARα in acute promyelocytic leukaemia

PML-RAR (retinoic acid receptor)α is the hallmark protein of acute promyelocytic leukaemia, a highly malignant subtype of acute myeloid leukaemia that accounts for approximately 10% of all AML cases. Recently, several studies have been set out to obtain a comprehensive genome-wide view of the molecular actions of this chimeric protein. In this review, we highlight the new insights that arose from these studies, in particular focussing on newly identified PML-RARα target genes, its interplay with RXR and deregulation of epigenetic modifications.     

Immunocytochemical markers in acute leukaemias diagnosis

The study included 1742 patients with acute myeloblastic leukaemias (AML) and acute lymphoblastic leukaemias (ALL), Kyiv city residents and patients from 20 regions of Ukraine. Bone marrow and blood smears were sent at diagnosis to Reference Center. The analysis was based on May-Grünvald-Giemza (MGG) stain and cytochemical reactions (MPO, acNSE, CAE, AP, PAS). Immunocytochemical techniques (APAAP, LSAB) and broad panel of monoclonal antibodies (MoAbs) against lineage specific and differentiation antigens of leukocytes were employed for immunophenotyping of leukemic blast cells directly in blood and bone marrow smears. Different types of AML were defined by the expression of the cell surface and cytoplasmic antigens. Immunocytochemical study was required especially in diagnosing of AML with minimal differentiation, acute megakaryoblastic leukaemia, acute erythroid leukaemia and acute leukaemias of ambiguous lineage. Acute lymphoblastic leukaemias was broadly classified into B-lineage and T-lineage ALL. According to the degree of B-lymphoid differentiation of the blast cells four subtypes of B-lineage ALL were established. T-lineage ALL observed in patients were also divided into four subtypes. Immunocytochemical examination was required to diagnose AL of ambiguous lineage with no clear evidence of lineage differentiation (acute undifferentiated leukaemia) or those with blasts that express markers of more than one lineage (mixed phenotype acute leukaemias).     

Clinical features, cytogenetics and outcome in acute lymphoblastic and myeloid leukaemia of infancy: report from the MRC Childhood Leukaemia working party.

The clinical features, cytogenetics and response to treatment have been examined in 180 infants (aged <1 year) with acute leukaemia; 118 with acute lymphoblastic leukaemia (ALL) and 62 with acute myeloid leukaemia (AML). Comparison of clinical features showed no difference in age or sex distribution between infants with ALL and AML but infants with ALL tended to have higher leucocyte counts at presentation. Cytogenetic abnormalities involving 11q23 were found in 66% of ALL and 35% of AML cases, the commonest, t(4;11) being found only in ALL. The other recognised 11q23 translocations were found in both types of leukaemia. Few patients had the common cytogenetic abnormalities associated with ALL in older children and few with AML had good risk abnormalities. Four year event-free survival 60% cf 30% (P = 0.001) and survival 65% cf 41% (P = 0.007) were significantly better in AML than ALL. These results were due to a lower risk of relapse 27% cf 62% at four years. Superior event-free survival was also seen in the subgroup of patients with 11q23 abnormalities and AML (55% cf 23%). The reasons for superior response in AML are unknown but may be related to the intensity of treatment, lineage of the leukaemia or other as yet unidentified factors.