急性髓系白血病铁死亡相关基因筛选及预后模型的建立

基于急性髓系白血病(Acute Myeloid Leukemia,AML)临床大数据及多组学数据库探讨铁死亡相关基因在AML中的作用,并建立铁死亡基因表达相关预后模型。整合TCGA数据库中151例AML患者和GTEx数据库中337例正常人外周血的临床和转录组数据。将Wilcoxon检验和单因素Cox分析结果取交集,筛选出预后相关差异表达基因(Differential Expression Genes, DEGs),使用Lasso回归建立基因标志物预后模型,利用受试者工作特征曲线(Receiver Operating Characteristic Curve,ROC曲线)评价预测价值,Kaplan-Meier法进行生存分析,对AML患者临床数据进行单因素和多因素Cox回归分析,使用差异基因表达分析等方法比较高、低风险患者间的组学差异,最后,利用BeatAML数据库对基因标志物进行验证。将差异基因表达分析和单因素分析结果取交集,得到13个预后相关DEGs。构建了8个基因标志物的预后评分模型,并将患者分为高、低风险两组;ROC曲线分析证实了模型良好的预测性能;生存分析提示高、低风险组患者的生存率具有显著差异;单因素分析显示年龄和风险评分与患者整体生存显著相关,多因素分析显示,年龄和风险评分是独立预后指标。在2个风险组之间筛选出384个DEGs,GO富集分析结果显示,富集的基因大多与中性粒细胞和白细胞的趋化与迁移等免疫相关分子和通路显著相关,KEGG富集通路主要与TNF信号通路、细胞因子与细胞因子受体相互作用相关。BeatAML数据库验证结果显示,5个基因与预后显著相关。铁死亡相关基因在AML中显著表达,且高风险患者预后较差,该研究对AML铁死亡相关潜在生物标志物的发现和应用奠定了一定的基础。     

Splicing and alternative splicing in rice and humans

Rice is a monocot gramineous crop, and one of the most important staple foods. Rice is considered a model species for most gramineous crops. Extensive research on rice has provided critical guidance for other crops, such as maize and wheat. In recent years, climate change and exacerbated soil degradation have resulted in a variety of abiotic stresses, such as greenhouse effects, lower temperatures, drought, floods, soil salinization and heavy metal pollution. As such, there is an extremely high demand for additional research, in order to address these negative factors. Studies have shown that the alternative splicing of many genes in rice is affected by stress conditions, suggesting that manipulation of the alternative splicing of specific genes may be an effective approach for rice to adapt to abiotic stress. With the advancement of microarrays, and more recently, next generation sequencing technology, several studies have shown that more than half of the genes in the rice genome undergo alternative splicing. This mini-review summarizes the latest progress in the research of splicing and alternative splicing in rice, compared to splicing in humans. Furthermore, we discuss how additional studies may change the landscape of investigation of rice functional genomics and genetically improved rice. [BMB Reports 2013; 46(9): 439-447]     

Cytogenetic analysis in 139 Tunisian patients with de novo acute myeloid leukemia

This paper presents the results of a cytogenetic analysis in 139 Tunisian patients with de novo acute myeloid leukemia (AML), including 27 children aged 1-15 years and 112 adults. Mean age was 32 (range 1-75) and the M/F ratio was 1.43. Of our patients, 45% had apparently normal karyotypes. Acquired chromosome aberrations were found in 77 (55% ) patients. t(8;21) was identified in 27 patients (19%); t(15;17) in 13 patients (9%); deletion 7q or monosomy 7 in seven patients (5%); +8 in seven patients (5%); abnormal 16 in four patients (3%); 11q23 rearrangements in two patients (2%) and del(5q), in one patient (1%). The remaining 16 patients had miscellaneous clonal abnormalities. Specific translocations associated with the FAB type were found: t(8;21) with AML2 and t(15;17) with AML3. We concluded that our study in a Tunisian population confirmed the relation between some specific abnormalities and the FAB classification. We found a higher incidence for t(8;21) than usually described.     

Methylenetetrahydrofolate reductase gene polymorphisms contribute to acute myeloid leukemia and chronic myeloid leukemia susceptibilities: Evidence from meta-analyses

PurposeThe expression of methylenetetrahydrofolate reductase (MTHFR) is associated with acute myeloid leukemia (AML) and chronic myeloid leukemia (CML). Most studies have linked the common functional C677T and A1298C polymorphisms of the MTHFR gene and susceptibility to AML and CML, but the results were not consistent. The aim of the present study was to derive a more precise estimation of the relationship.MethodsMeta-analyses assessing the association of MTHFR C677T and A1298C variations with AML and CML were conducted. Eligible articles were identified from the PubMed and EMBASE databases. All statistical analyses were conducted using Review Manager Software.Results10 and 10 studies were included in the meta-analysis about the role of C677T polymorphism on the AML and CML risks, respectively; 6 and 4 studies were included about the role of A1298C polymorphism on the AML and CML risks, respectively. Overall, both the C677T and A1298C polymorphisms were significantly associated with CML risk under the recessive model (P = 0.04, OR = 1.35, 95% CI = 1.02–1.79 for C677T and P = 0.003, OR = 2.17, 95% CI = 1.29–3.63 for A1298C). In addition, the risk of CML was higher in 1298CC genotype carriers than in 1298AA genotype carriers (P = 0.004, OR = 2.17, 95% = 1.28–3.69). Conversely, the overall data failed to indicate a significant association of C677T or A1298C polymorphisms with AML risk under any model.ConclusionsThe findings provide evidence that C677T and A1298C polymorphisms are risk factors for CML risk.     

Identification of novel prognostic alternative splicing signature in papillary renal cell carcinoma

Papillary renal cell carcinoma (pRCC) is a heterogeneous disease containing multifocal or solitary tumors with an aggressive phenotype. Increasing evidence has indicated the involvement of aberrant splicing variants in renal cell cancer, while systematic profiling of aberrant alternative splicing (AS) in pRCC was lacking and largely unknown. In the current study, comprehensive profiling of AS events were performed based on the integration of pRCC cohort from the Cancer Genome Atlas database and SpliceSeq software. With rigorous screening and univariate Cox analysis, a total of 2077 prognoses AS events from 1642 parent genes were identified. Then, stepwise least absolute shrinkage and selection operator method-penalized Cox regression analyses with 10-fold cross-validation followed by multivariate Cox regression were used to construct the prognostic AS signatures within each AS type. And a final 21 AS event-based signature was proposed which showed potent prognostic capability in stratifying patients into low- and high-risk subgroups (P < .0001). Furthermore, time-dependent receiver operating characteristics curves confirmed that the final AS signature was effective and robust in predicting overall survival for pRCC patients with the area under the curve above 0.9 from 1 to 5 years. In addition, splicing correlation network was built to uncover the potential regulatory pattern among prognostic splicing factors and candidate AS events. Besides, gene set enrichment analysis revealed the involvement of these candidates AS events in tumor-related pathways including extracellular matrix organization, oxidative phosphorylation, and P53 signaling pathways. Taken together, our results could contribute to elucidating the underlying mechanism of AS in the oncogenesis process and broaden the novel field of prognostic and clinical application of molecule-targeted approaches in pRCC.     

The molecular signature of oncofusion proteins in acute myeloid leukemia

Acute myeloid leukemia (AML) associated translocations often cause gene fusions that encode oncofusion proteins. Although many of the breakpoints involved in chromosomal translocations have been cloned, in most cases the role of the chimeric proteins in tumorigenesis is not elucidated. Here we will discuss the fusion proteins of the 4 most common translocations associated with AML as well as the common molecular mechanisms that these four and other fusion proteins utilize to transform progenitor cells. Intriguingly, although the individual partners within the fusion proteins represent a wide variety of cellular functions, at the molecular level many commodities can be found.     

Risk of adult acute and chronic myeloid leukemia with cigarette smoking and cessation

BackgroundCigarette smoking is an established risk factor for adult myeloid leukemia, particularly acute myeloid leukemia (AML), but less is known about the nature of this association and effects of smoking cessation on risk.MethodsIn a large population-based case–control study of myeloid leukemia that included 414 AML and 185 chronic myeloid leukemia (CML) cases and 692 controls ages 20–79 years, we evaluated risk associated with cigarette smoking and smoking cessation using unconditional logistic regression methods and cubic spline modeling.ResultsAML and CML risk increased with increasing cigarette smoking intensity in men and women. A monotonic decrease in AML risk was observed with increasing time since quitting, whereas for CML, the risk reduction was more gradual. For both AML and CML, among long-term quitters (≥30 years), risk was comparable to non-smokers.ConclusionsOur study confirms the increased risk of myeloid leukemia with cigarette smoking and provides encouraging evidence of risk attenuation following cessation.     

Identification and targeting leukemia stem cells: The path to the cure for acute myeloid leukemia

Accumulating evidence support the notion that acute myeloid leukemia(AML) is organized in a hierarchical system, originating from a special proportion of leukemia stem cells(LSC). Similar to their normal counterpart, hematopoietic stem cells(HSC), LSC possess selfrenewal capacity and are responsible for the continued growth and proliferation of the bulk of leukemia cells in the blood and bone marrow. It is believed that LSC are also the root cause for the treatment failure and relapse of AML because LSC are often resistant to chemotherapy. In the past decade, we have made significant advancement in identification and understanding the molecular biology of LSC, but it remains a daunting task to specifically targeting LSC, while sparing normalHSC. In this review, we will first provide a historical overview of the discovery of LSC, followed by a summary of identification and separation of LSC by either cell surface markers or functional assays. Next, the review will focus on the current, various strategies for eradicating LSC. Finally, we will highlight future directions and challenges ahead of our ultimate goal for the cure of AML by targeting LSC.     

RNAi干预急性髓系白血病对p38信号通路的影响

目的 探讨RNAi技术治疗急性髓系白血病（acute myeloid leukemia,AML）对p38信号通路的影响。方法 以NC（HL-60细胞）和HK（含siRNA-FLT3的阴性对照的HL-60细胞）、F1（实验组,含siRNA-FLT3的HL-60细胞,本实验室已实验证明对FLT3有干扰作用）三种细胞株为研究对象,分为药物组（信号通道抑制作用组）和对照组（未加信号通道抑制剂）。通过RT-PCR和Western blot检测对照组和药物组p38 mRNA水平和蛋白表达的变化,MTT和FCM分别测定细胞活性和细胞凋亡率的变化。结果 药物组、对照组组内比较：以NC做参照,F1能诱导p38 mRNA水平和蛋白表达下降、从而抑制细胞的增殖,促进细胞凋亡,差异有统计学意义（均P 0.05）;与对照组相比,药物组p38 mRNA水平和蛋白表达下降,细胞活性下降,细胞凋亡率增加,差异有统计学意义（均P<0.05）;药物组的F1与药物组的NC、药物组的F1与对照组的F1分别比较,差异有统计学意义（均P s<0.05）。结论 p38信号通路是RNAi技术干扰HL-60细胞FLT3基因表达引起细胞凋亡的途径之一,SB203580（P38 MAPK抑制剂）对其有协同增效作用。     

Interleukin-17 in acute myeloid leukemia

There are several reports that angiogenesis plays important roles in hematological malignancies including acute myeloid leukemia (AML). Human interleukin-17 (IL-17) is a proinflammatory cytokine produced by activated CD4 T cells. IL-17 plays a potential role in T cell mediated angiogenesis. The role of IL-17 in pathologic angiogenesis has not been evaluated yet. The aim of the study was to determine plasma level of IL-17 in patients with AML. IL-17 levels were measured by ELISA in plasma samples taken from 68 adult patients with AML before chemotherapy was administered. In addition 20 out of 68 patients were reanalysed after achieving complete remission (CR). Ten samples from healthy volunteers were evaluated as the control. In this study we have demonstrated that serum level of IL-17 is not elevated in AML patients. These results suggest that angiogenesis in AML is not mediated by CD4 T cells. To our knowledge this is the first report about IL-17 serum level in acute leukemias. We are currently evaluating IL-17 levels in others haematological malignancies.     

Targeting leukemia stem cells: The new goal of therapy in adult acute myeloid leukemia

The most popular view of hematopoietic cell lineage organization is that of complex reactive or adaptative systems. Leukemia contains a subpopulation of cells that display characteristics of stem cells. These cells maintain tumor growth. The properties of leukemia stem cells indicate that current conventional chemotherapy, directed against the bulk of the tumor, will not be effective. Leukemia stem cells are quiescent and do not respond to cell cycle-specific cytotoxic agents used to treat leukemia and thus contribute to treatment failure. New strategies are required that specifically target this malignant stem cell population.     

Identification of novel CD33 antigen-specific peptides for the generation of cytotoxic T lymphocytes against acute myeloid leukemia

Identification of immunogenic peptides for the generation of cytotoxic T lymphocytes (CTLs) may lead to the development of novel cellular therapies to treat disease relapse in acute myeloid leukemia (AML) patients. The objective of these studies was to evaluate the ability of unique HLA-A2.1-specific nonameric peptides derived from CD33 antigen to generate AML-specific CTLs ex vivo. We present data here on the identification of an immunogeneic HLA-A2.1-specific CD33(65-73) peptide (AIISGDSPV) that was capable of inducing CTLs targeted to AML cells. The CD33-CTLs displayed HLA-A2.1-restricted cytotoxicity against both mononuclear cells from AML patients and the AML cell line. The peptide- as well as AML cell-specificity of CD33-CTLs was demonstrated and the secretion of IFN-gamma by the CTLs was detected in response to CD33(65-73) peptide stimulation. The cultures contained a distinct CD33(65-73) peptide-tetramer(+)/CD8(+) population. Alteration of the native CD33(65-73) peptide at the first amino acid residue from alanine (A) to tyrosine (Y) enhanced the HLA-A2.1 affinity/stability of the modified CD33 peptide (YIISGDSPV) and induced CTLs with increased cytotoxicity against AML cells. These data therefore demonstrate the potential of using immunogenic HLA-A2.1-specific CD33 peptides in developing a cellular immunotherapy for the treatment of AML patients.     

Potential CD34 signaling through phosphorylated-BAD in chemotherapy-resistant acute myeloid leukemia

Abstract

Acute myeloid leukemia (AML) constitutively express growth factors and cytokines for survival. Chemotherapy alters these signals to induce cell death. However, drug resistance in AML remains a major hindrance to successful treatment and early warning is unavailable. Modulation of signaling pathways during chemotherapy may provide a window to detect response and predict treatment outcome. Blood samples collected from AML patients before and at day-3 of induction therapy were compared for changes in expression of CD117, CD34, pro-inflammatory cytokines and mediators of Akt and MAPK pathways, using multi-color flow cytometry. Nine patients were diagnosed as drug-resistant and seven sensitive to chemotherapy. Twelve were paired. Average percentages of CD34 (66.8?±?11.7% vs. 26.2?±?5.8%, p?=?0.033) and pBAD (66.9?±?8.2% vs. 28.9?±?8.2%, p?=?0.016) were significantly increased in chemo-resistant (N?=?9) compared to chemo-sensitive (N?=?5) samples. Percentages of CD34 were strongly correlated with pBAD (R?=?0.785; p?=?0.001; N?=?14) and pFKHR (R?=?0.755; p?=?0.002; N?=?14) at day-3 induction. Chemo-sensitive cases expressed significantly higher percentages of IL-18Rα (71.9?±?9.6% vs. 29.8?±?5.8%, p?=?0.016). Though not significantly different in the outcome, IL-1β was strongly associated with activated Akt-S473, IL-6 with phosphorylated JNK and FKHR while TNF-α appeared to trigger Bim, in treated samples. These preliminary results suggested AML cells resistant to chemotherapy increased expression of CD34 and may signal through pBAD while cells sensitive to chemotherapy-induced IL18Rα expression. These were observed early during induction therapy. Identifying CD34 is interesting as it is a convenient marker to monitor drug-resistance in AML patients. Inhibition of CD34 and pBAD signaling may be important in treating drug-resistant AML.     

Dihydromyricetin sensitizes human acute myeloid leukemia cells to retinoic acid-induced myeloid differentiation by activating STAT1

The success of all-trans retinoic acid (ATRA) in differentiation therapy for patients with acute promyelocytic leukemia (APL) highly encourages researches to apply a new combination therapy based on ATRA. Therefore, research strategies to further sensitize cells to retinoids are urgently needed. In this study, we showed that Dihydromyricetin (DMY), a 2,3-dihydroflavonol compound, exhibited a strong synergy with ATRA to promote APL NB4 cell differentiation. We observed that DMY sensitized the NB4 cells to ATRA-induced cell growth inhibition, CD11b expression, NBT reduction and myeloid regulator expression. PML-RARα might not be essential for DMY-enhanced differentiation when combined with ATRA, while the enhanced differentiation was dependent on the activation of p38-STAT1 signaling pathway. Taken together, our study is the first to evaluate the synergy of DMY and ATRA in NB4 cell differentiation and to assess new opportunities for the combination of DMY and ATRA as a promising approach for future differentiation therapy.     

Over-expression of FoxM1 is associated with adverse prognosis and FLT3-ITD in acute myeloid leukemia

Forkhead box M1 (FoxM1) drives cell cycle progression and the prevention of growth arrest and is over-expressed in many human malignancies. However, the characteristics of FoxM1 in acute myeloid leukemia (AML) are not clearly understood. We investigated the expression level of FoxM1 and analyzed the correlation of FoxM1 expression with AML patient characteristics and prognoses. Changes in FoxM1 expression were detected after MV4–11 cells, which have an internal tandem duplication (ITD) of the fms-like tyrosine kinase 3 gene (FLT3-ITD), and control THP1 cells (encoding wild-type FLT3) were treated with the FLT3 receptor tyrosine kinase inhibitor AC220 (quizartinib) or FLT3 ligand (FL). Finally, we determined the apoptosis rates after the addition of the FoxM1 inhibitor thiostrepton (TST) to AML cells with or without FLT3-ITD. The expression of FoxM1 in AML patients was correlated with the presence of FLT3-ITD, genetic groups, and possibly overall survival. Inhibition of FLT3-ITD by AC220 down-regulated FoxM1 expression in MV4–11 cells, and stimulation of FLT3 by FL up-regulated FoxM1 expression in MV4–11 and THP1 cells. TST induced the apoptosis of MV4–11 and THP1 cells in a dose-dependent manner. Thus, FoxM1 is a potential prognostic marker and a promising therapeutic target in AML.     

Visualization of alternative splicing in vivo

The analysis of sequences required for alternative splicing of mRNA has predominantly been performed using cell culture systems. However, the phenotype of cultured cells is almost invariably different from that of cells in the intact animal. It is therefore possible that there are significant differences in the regulation of specific splicing reactions in vivo compared to in cell culture. Here, we describe methods for the visualization and analysis of alternative splicing in vivo using transgenic mice. These methods allow for the analysis of the temporal and tissue-specific regulation of alternative splicing both visually and quantitatively.