基于生物信息学分析SCHIP1在急性髓系白血病中的表达及其临床意义

通过数据挖掘和生物信息学分析手段,探讨施旺膜蛋白相互作用因子1(schwannomin interacting protein 1,SCHIP1)基因在急性髓系白血病患者中的表达情况及其临床意义。首先,对Oncomine数据库中收录的所有急性髓系白血病(acute myloid leukaemia,AML)数据集进行荟萃分析,筛选出目标基因SCHIP1并进一步分析其在AML病人中的表达变化。随后,从GEO数据库中下载含生存信息的AML数据集源文件,分析SCHIP1表达对疾病的预后作用。另外,利用TCGA数据库对SCHIP1的表达情况进行亚组分析及与FLT3基因突变、PML/RARα融合基因和RAS活化等高危因素进行相关性分析。最后,利用GEPIA2工具验证SCHIP1的表达情况、预后意义及与FLT3、PML基因表达的相关性。结果发现Oncomine数据库中收录了44个AML数据集,总计共3534个样本数据。其中5个数据集共1188个样本包含“Cancer vs.Normal”的mRNA表达数据,对其进行荟萃分析显示SCHIP1位于显著高表达分子的第17位。生存分析显示,SCHIP1表达量与AML患者总体生存率呈负相关。亚组分析显示SCHIP1在M0/M1/M2中较M3/M6中表达更高,但与年龄、性别和种族无关。另外,相关性研究分析显示SCHIP1与FLT3基因突变弱相关,但与PML/RARα融合基因和RAS活化等高危因素无显著相关性。这些结果表明SCHIP1在急性髓系白血病中高表达,且其高表达与患者的生存预后呈显著负相关。因此,SCHIP1可作为疾病的预后生物标志物,并有望成为AML的精准治疗靶点。     

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。     

Daxx在急性白血病的表达及意义

目的研究Daxx在急性白血病(AL)中的表达及其与AL的分型、临床特征、疗效及预后的关系.方法应用免疫组织化学方法检测88例初治AL骨髓细胞Daxx蛋白的表达情况,分析其与FAB分型、临床特征、疗效及预后的关系.结果急性髓细胞白血病(AML)骨髓细胞中Daxx的表达显著高于正常对照组(P<0.05)与急性淋巴细胞白血病(ALL)组(P<0.05).在AML亚型中,Daxx在M3中的表达显著高于M1,M2,M3,M4和M5.采用逐步回归法分析,校正其它参数,Daxx的表达与初诊时的白细胞数及骨髓原始细胞数目呈正相关,与疗效呈负相关. 结论 Daxx在急性髓细胞白血病中广泛表达,其表达的紊乱可能在AML的发病中起作用,还与AML的某些临床特征、疗效及预后密切相关.     

急性白血病的P16基因表达分析

应用免疫组织化学方法．对61例急性白血病患者P16基因表达进行检测和分析。结果表明：AML和ALL患者的P16表达均显著低于对照组（P＜0.001）,ALLP16表达又显著低于AML（P＜0．05）。AL亚型中P16表达率以M2最高,L3最低。比较耐药组和药敏组,P16表达两组间有显著差异（P＜0．05）,但初诊组与复发组、缓解组与未缓解组比较未见统计学差异（P＞0．05）。上述结果提示：P16基因表达异常在急性白血病的发生、发展中起重要作用,但与白血病患者的治疗和预后无明显关系。     

CD47 在急性白血病干细胞中的表达及其对临床疗效及预后的影响

目的:探讨CD47在急性白血病患者骨髓白血病细胞的表达及其临床意义。方法:选择2013年5月-2015年5月在我院确诊的急性白血病患者101例作为研究对象,其中急性淋巴细胞白血病50例(ALL组),急性髓系白血病51例(AML组)。另选取同期在我院接受体检的健康志愿者39例作为对照组。采用流式细胞仪检测白血病细胞表面CD47的表达情况,并分析CD47表达与急性白血病患者临床疗效及复发情况的关系。结果:急性白血病患者白血病细胞CD47的阳性表达率明显高于健康对照组,差异具有统计学意义(P0.05);而ALL组与AML组患者白血病细胞CD47的阳性表达率比较差异无统计学意义(P0.05);CD47阴性表达的急性白血病患者CR率显著高于阳性表达者,差异具有统计学意义(P0.05);ALL组和AML组CD47阴性表达患者CR率显著高于CD47阳性表达患者,差异具有统计学意义(P0.05),但两组之间比较,差异无统计学意义(P0.05);CD47阳性表达的急性白血病患者复发率显著高于阴性表达患者,差异具有统计学意义(P0.05);ALL组和AML组CD47表达阳性患者复发率明显高于阴性患者,差异具有统计学意义(P0.05),但两组之间比较差异无统计学意义(P0.05)。结论:急性白血病患者白血病细胞表面CD47的表达异常升高,且与白血病患者的疗效和预后有关,CD47可能作为一种急性白血病的诊断及疗效和预后的辅助评估指标。     

急性髓系白血病患者外周血FOXO1、PBX3的表达及对预后的预测价值

摘要 目的：探讨急性髓系白血病（AML）患者外周血叉头框转录因子O1（FOXO1）、B细胞白血病同源盒基因3（PBX3）的表达及对预后的预测价值。方法：前瞻性选取2017年8月至2021年8月收治的60例急性髓系白血病患者作为研究对象。所有患者均随访1年并按照随访结果分为预后良好组（36例）和预后不良组（24例）。采用Pearson检验进行相关性分析;采用logistics回归模型分析AML患者预后的独立危险因素;采用ROC曲线分析FOXO1、PBX3对AML患者预后的预测价值。结果：预后良好组和预后不良组年龄、性别、BMI比较无显著差异（P＞0.05）,而在血小板、中性粒细胞比值、FOXO1、PBX3、白细胞计数存在显著差异（P<0.05）;FOXO1、PBX3与血小板、中性粒细胞比值、白细胞计显著正相关（P<0.05）;多因素分析结果显示,血小板、中性粒细胞比值、FOXO1、PBX3、白细胞计数是影响急性髓系白血病患者预后的独立危险因素（P<0.05）;血清FOXO1、PBX3预测AML患者预后的灵敏度和特异性达到91.31 %和92.31 %。结论：急性髓系白血病患者外周血FOXO1、PBX3的表达上调可作为预测评估其预后发展的可靠血清标志物。     

EVI1基因在急性髓系白血病中对PTEN表达呈负相关

探讨急性髓系白血病中EVI1基因对PTEN表达的影响及二者的关系。采用RT-PCR方法检测263例AML病人(初诊与慢粒急髓变组239例,复发组13例,缓解组11例)骨髓单个核细胞中EVI1和PTEN m RNA的表达水平,对照组12例为正常人及缺铁性贫血病人。EVI1和PTEN m RNA在AML病人骨髓单个核细胞中阳性表达率分别为7.5%和61.1%。初诊组患者EVI1 m RNA表达水平与缓解组相比显著升高(p0.000 1),对照组患者中并无EVI1表达;PTEN m RNA表达量较缓解组及正常对照组显著下降(p值分别为p=0.010 7,p0.000 1);缓解组患者EVI1 m RNA表达量降低,PTEN m RNA表达量升高,与初诊组及对照组比较有统计学差异。AML各亚型间EVI1、PTEN m RNA的表达量无统计学意义。AML初诊组中EVI1、PTEN m RNA表达呈负相关,R=0.611 4,p=0.000 3。在AML中EVI1和PTEN基因的表达呈负相关,为EVI1阳性的AML患者寻找治疗靶点提供依据。     

急性髓系白血病免疫表型特征及遗传学特征分析

目的:研究急性髓系白血病免疫表型特征以及遗传学特征。方法:选取2011年1月到2014年5月我院收治的急性髓系白血病患者169例,采用流式细胞术和相关的单克隆抗体来分析所有患者的骨髓免疫表型,采用染G染色体显带技术分析患者的核型,根据淋系抗原(lym Ag)的表达将患者分为lym Ag+组和lym Ag-组。结果:抗原CD13、CD33、CD117以及MPO等髓系抗原最常在急性髓系白血病患者中表达,其中CD117在M3型病例中表达为85.7%(24/28),而CD34、HLA-DR双阴性、较强的自发荧光、CD13、CD33和MPO对M3型的诊断也具有一定的价值;其中47.9%(81/169)的患者伴随着淋系抗原表达,以CD7和CD56为主;60.4%(102/169)的患者伴随着核型异常;而伴随着t(8:21)的M2患者中的CD15、CD19和CD56的表达显著增强,而t(15:17)均发生于M3型患者中;而lym Ag+组患者CD34的阳性患者为77.8%(63/81)显著高于lym Ag-组的47.7%(42/88),两组比较差异具有统计学意义(P0.05)。结论:免疫表型对急性髓系白血病的诊断具有重要的意义,且免疫表型和异常核型存在密切的联系。     

小鼠急性髓系白血病模型的构建

MLL基因的异常重排会引发急性淋巴系(ALL)和急性髓系白血病(AML)。该文详细阐述了利用逆转录病毒载体MLL-AF9构建小鼠AML模型的方法。该研究比较了免疫磁珠法与5-氟尿嘧啶(5-FU)方法富集骨髓细胞的效率,以及不同时间点收获的病毒对骨髓Lin-细胞感染效率的影响。通过流式检测发现, 5-氟尿嘧啶富集的骨髓Lin–细能够被48 h收获的病毒高效感染。受体鼠在移植了MLL-AF9感染的骨髓Lin–细胞60天后,外周血、骨髓、脾脏组织中均有大量的白血病细胞浸润, RT-qPCR也验证了白血病靶基因的表达上调,表明小鼠AML模型的成功构建。这项研究为从事白血病研究的科研人员提供了一种有效的小鼠急性髓系白血病模型,为研究白血病发病机理与研发白血病治疗药物提供有用的工具。     

过表达TRAF6对人急性髓系白血病细胞自噬活性的影响

该文旨在探讨过表达肿瘤坏死因子受体相关因子6(tumor necrosis factor receptorassociated factor 6,TRAF6)对人急性髓系白血病(acute myeloid leukemia,AML)细胞自噬活性的影响。利用基因表达数据库GEO分析TRAF6在AML患者白血病细胞中的mRNA表达水平。通过癌症基因组图谱TCGA分析TRAF6表达与AML患者临床预后的关系。将TRAF6重组质粒载体转染人AML细胞系(KG-1a和THP-1),采用自噬激活剂雷帕霉素(Rapamycin)和自噬相关抑制剂3-甲基腺嘌呤(3-methyladenine,3-MA)、巴弗洛霉素A1(bafilomycin A1,Baf-A1)分别处理AML细胞。荧光定量PCR、蛋白免疫印迹技术检测过表达TRAF6后白血病细胞自噬标志物(LC3和p62)mRNA和蛋白水平;免疫荧光方法检测LC3绿色荧光斑点结构(puncta);流式细胞术检测细胞凋亡率;CCK-8实验检测AML细胞的体外增殖能力。结果显示,AML患者白血病细胞高表达TRAF6(P<0.01);TRAF6高表达的白血病患者总体生存率和无事件生存率均较TRAF6低表达组显著降低(P=0.01)。TRAF6重组质粒转染能够显著增加两株AML细胞系中TRAF6的mRNA和蛋白水平(P<0.05)。Rapamycin处理能够激活AML细胞系自噬水平,过表达TRAF6后AML细胞LC3 mRNA和LC3II蛋白水平表达上调(P<0.05)、p62 mRNA和蛋白水平下调(P<0.05)以及LC3 puncta聚集增多。用Baf-A1处理以阻断过表达TRAF6的白血病细胞系中的自噬流后,LC3II蛋白表达水平显著提高(P<0.05)。3-MA处理过表达TRAF6的白血病细胞后,LC3II蛋白表达减少、p62蛋白表达增加(P<0.05)。此外,过表达TRAF6降低白血病细胞凋亡率和促进细胞的体外增殖(P<0.001),而过表达TRAF6后联合3-MA处理则可逆转TRAF6对白血病细胞的抗凋亡和促增殖作用(P<0.001)。以上研究结果提示,过表达TRAF6能够增强AML细胞的自噬活性,促进AML细胞的生长。     

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.     

RUNX1 mutation and elevated FLT3 gene expression cooperates to induce inferior prognosis in cytogenetically normal acute myeloid leukemia patients

BackgroundAcute myeloid leukemia (AML) is a bone marrow malignancy having multiple molecular pathways driving its progress. In recent years, the main causes of AML considered all over the world are genetic variations in cancerous cells. The RUNX1 and FLT3 genes are necessary for the normal hematopoiesis and differentiation process of hematopoietic stem cells into mature blood cells, therefore they are the most common targets for point mutations resulting in AML.MethodsWe screened 32 CN-AML patients for FLT3-ITD (by Allele-specific PCR) and RUNX1 mutations (by Sanger sequencing). The FLT3 mRNA expression was assessed in all AML patients and its subgroups.ResultsEight patients (25%) carried RUNX1 mutation (K83E) while three patients (9.37%) were found to have internal tandem duplications in FLT3 gene. The RUNX1 mutation data were correlated with clinical parameters and FLT3 gene expression profile. The RUNX1 mutations were observed to be significantly prevalent in older males. Moreover, RUNX1 and FLT3-mutated patients had lower complete remission rate, event-free survival rate, and lower overall survival rate than patients with wild-type RUNX1 and FLT3 gene. The RUNX1 and FLT3 mutant patients with up-regulated FLT3 gene expression showed even worse prognosis. Bradford Assay showed that protein concentration was down-regulated in RUNX1 and FLT3 mutants in comparison to RUNX1 and FLT3 wild-type groups.ConclusionThis study constitutes the first report from Pakistan reporting significant molecular mutation analysis of RUNX1 and FLT3 genes including FLT3 expression evaluation with follow-up. This provides an insight that aforementioned mutations are markers of poor prognosis but the study with a large AML cohort will be useful to further investigate their role in disease biology of AML.     

Molecular mechanisms that produce secondary MDS/AML by RUNX1/AML1 point mutations

RUNX1/AML1 point mutations have been identified in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) patients. A heterozygous germline mutation of the RUNX1 gene causes a familial platelet disorder with a predisposition to AML. RUNX1 mutations have also been detected with high frequency in minimally differentiated AML M0 subtypes and myelodysplastic/myeloproliferative neoplasms. Here we propose a new disease category of myelodysplastic neoplasms (MDN) consisting of MDS refractory anemia with excess blasts and AML with myelodysplasia-related changes, including therapy-related cases. RUNX1 mutations have been detected in about 20% of patients with "MDN". Among the MDN cases, histories of radiation exposure, therapy-related myeloid neoplasms after successful treatment for acute promyelocytic leukemia, and leukemic transformation of myeloproliferative neoplasms have been reported to have a strong association with RUNX1 mutations. The mutations occur in a normal, a receptive, or a disease-committed hematopoietic stem cell. It is suspected that the "MDN" phenotypes are defined by the RUNX1 mutations in addition to some other abnormalities.     

Acquired TET2 mutation in one patient with familial platelet disorder with predisposition to AML led to the development of pre‐leukaemic clone resulting in T2‐ALL and AML‐M0

Familial platelet disorder with predisposition to acute myeloid leukaemia (FPD/AML) is characterized by germline RUNX1 mutations, thrombocytopaenia, platelet dysfunction and a risk of developing acute myeloid and in rare cases lymphoid T leukaemia. Here, we focus on a case of a man with a familial history of RUNX1^R174Q mutation who developed at the age of 42 years a T2‐ALL and, 2 years after remission, an AML‐M0. Both AML‐M0 and T2‐ALL blast populations demonstrated a loss of 1p36.32‐23 and 17q11.2 regions as well as other small deletions, clonal rearrangements of both TCRγ and TCRδ and a presence of 18 variants at a frequency of more than 40%. Additional variants were identified only in T2‐ALL or in AML‐M0 evoking the existence of a common original clone, which gave rise to subclonal populations. Next generation sequencing (NGS) performed on peripheral blood‐derived CD34⁺ cells 5 years prior to T2‐ALL development revealed only the missense TET2^P1962T mutation at a frequency of 1%, which increases to more than 40% in fully transformed leukaemic T2‐ALL and AML‐M0 clones. This result suggests that TET2^P1962T mutation in association with germline RUNX1^R174Q mutation leads to amplification of a haematopoietic clone susceptible to acquire other transforming alterations.     

Proteogenomics approaches for studying cancer biology and their potential in the identification of acute myeloid leukemia biomarkers

Introduction: Mass spectrometry (MS)-based proteomics has become an indispensable tool for the characterization of the proteome and its post-translational modifications (PTM). In addition to standard protein sequence databases, proteogenomics strategies search the spectral data against the theoretical spectra obtained from customized protein sequence databases. Up to date, there are no published proteogenomics studies on acute myeloid leukemia (AML) samples.

Areas covered: Proteogenomics involves the understanding of genomic and proteomic data. The intersection of both datatypes requires advanced bioinformatics skills. A standard proteogenomics workflow that could be used for the study of AML samples is described. The generation of customized protein sequence databases as well as bioinformatics tools and pipelines commonly used in proteogenomics are discussed in detail.

Expert commentary: Drawing on evidence from recent cancer proteogenomics studies and taking into account the public availability of AML genomic data, the interpretation of present and future MS-based AML proteomic data using AML-specific protein sequence databases could discover new biological mechanisms and targets in AML. However, proteogenomics workflows including bioinformatics guidelines can be challenging for the wide AML research community. It is expected that further automation and simplification of the bioinformatics procedures might attract AML investigators to adopt the proteogenomics strategy.     

血清miR-203、miR-217表达与急性髓系白血病患者预后的关系

摘要 目的：探究血清miR-203、miR-217表达与急性髓系白血病(AML)患者预后的关系。方法：选择2010年4月至2014年4月我院诊治的101例AML患者作为AML组,AML组根据治疗效果进一步分为完全缓解组和复发组,选择同期在我院体检的101例健康者作为健康组。采用荧光定量PCR检测各组的血清miR-203、miR-217表达水平,分析血清miR-203、miR-217表达水平与患者临床病理特征的关系,采用Kaplan-Meier法分析不同血清miR-203、miR-217表达水平AML患者的预后。结果：与健康组相比,AML组的血清miR-203、miR-217表达水平明显更低（P<0.05）。与完全缓解组相比,复发组的血清miR-203、miR-217表达水平明显更低（P<0.05）。血清miR-203表达水平与AML患者白细胞计数相关（P<0.05）,而血清miR-217表达水平与AML患者血小板计数相关（P<0.05）。血清miR-203相对高表达和miR-217相对高表达的AML患者5年生存率分别高于血清miR-203相对低表达和miR-217相对低表达患者（Log Rank ^miR-203 =17.870,Log Rank ^miR-217 =28.926,均P=0.000）。结论：血清miR-203、miR-217的表达水平与AML密切相关,检测血清miR-203、miR-217表达水平可能有助于评估AML患者的预后。