Differential miRNA expression in childhood acute lymphoblastic leukemia and association with clinical and biological features

The present study aimed to analyze the expression profile of the microRNAs previously described as associated with childhood ALL, miR-92a, miR-100, miR-125a-5p, miR-128a, miR-181b, miR-196b and let-7e, and their association with biological/prognostic features in 128 consecutive samples of childhood acute lymphoblastic leukemia (ALL) by quantitative real-time PCR. A significant association was observed between higher expression levels of miR-196b and T-ALL, miR-100 and patients with low white blood cell count at diagnosis and t(12;21) positive ALL. These findings suggest a potential activity of these microRNAs in pediatric ALL biology.     

Distinctive microRNA signature is associated with the diagnosis and prognosis of acute leukemia

MicroRNAs (miRNAs) are of great importance in pathogenesis, diagnosis and prognosis of acute leukemia (AL). We studied five AL-related miRNAs to confirm the significance of these miRNAs in AL. Samples tested included acute myeloid leukemia (AML), 107 cases; acute lymphoblastic leukemia (ALL), 40 cases. Five AL-related miRNAs: miR-128, let-7b, miR-223, miR-181a and miR-155 expression were detected by qRT-PCR. Analysis showed that miRNA-128 expression was significantly higher in ALL (P?相似文献   

Methylation-mediated repression of microRNA-143 enhances MLL-AF4 oncogene expression

Fusion proteins containing the amino terminus of mixed lineage leukemia (MLL) are common in acute lymphoblastic leukemia (ALL) due to translocations. The MLL-AF4 fusion protein is generated by the translocation t(4;11)(q21;q23), and t(4;11)-positive ALL patients (MLL-AF4 ALL), have a notoriously poorer prognosis compared with patients with other MLL-associated leukemias. The detailed role of this fusion protein in leukemogenesis is not well understood. MicroRNAs (miRNAs) targeting the AF4 3' untranslated regions may modulate MLL-AF4 fusion protein levels, raising the question of whether regulation of these miRNAs are involved in the progression of MLL-AF4 ALL. In this study, we show that miR-143 was identified as a regulator of MLL-AF4 expression in MLL-AF4 ALL samples. Restoration of miR-143 in MLL-AF4-positive RS4;11 and MV4-11 cells induced apoptosis, negatively contributing to leukemia cell growth by reducing MLL-AF4 fusion protein levels. Furthermore, miR-143 was epigenetically repressed by promoter hypermethylation in MLL-AF4-positive primary blasts and cell lines, but not in normal bone marrow cells and MLL-AF4-negative primary blasts, which was directly associated with expression of the MLL-AF4 oncogene. This is the first study to show that miR-143 functions as a tumor suppressor in MLL-AF4 B-cell ALL. These data reveal the therapeutic promise of upregulating miR-143 expression for MLL-AF4 B-cell ALL.     

Circulating microRNA-196a/b are novel biomarkers associated with metastatic gastric cancer

miR-196a and/or miR-196b, involved in cancer initiation and progression, are frequently upregulated in tumour tissues. However, the clinical significance of these microRNAs in gastric cancer (GC) remains to be clarified. In the current study, we investigated the potential utility of circulating miR-196a/b as novel biomarkers for early detection and/or metastatic prognosis of GC. The quantitative real time-polymerase chain reaction data revealed markedly higher pre-operative circulating miR-196a and miR-196b levels in GC patients than healthy controls. Receiver-operating characteristics curve analysis showed that circulating miR-196a, miR-196b and combined miR-196a and miR-196b (miR-196a/b) are more effective than carcinoembryonic antigen or carbohydrate antigen 19-9 alone in distinguishing GC patients from healthy controls, with higher sensitivity and specificity. Circulating miR-196a exhibited higher diagnostic capacity than combined miR-196a/b or miR-196b alone, highlighting its potential as an effective plasma biomarker for GC. In clinicopathological analysis, elevated circulating miR-196a/b levels were highly correlated with metastatic potential or more advanced stages of disease and poorer survival. In addition, the expression levels of circulating miR-196a/b were reduced after surgical resection in GC patients. Taken together, we propose that circulating miR-196a/b serve as a more sensitive and specific novel biomarker than carbohydrate antigen 19-9 for GC monitor, diagnosis and prognosis.     

miR-181a/b significantly enhances drug sensitivity in chronic lymphocytic leukemia cells via targeting multiple anti-apoptosis genes

MicroRNAs (miRNAs) have been shown to play critical roles in regulating the progress of leukemia. We performed miRNA expression profile in six Chinese patients with chronic lymphocytic leukemia (CLL), and in peripheral B cells from pooled 30 healthy donors, using a platform containing 866 human miRNAs. The most frequent changes in miRNAs in CLL cells included downregulation of miR-126, miR-572, miR-494, miR-923, miR-638, miR-130a, miR-181a and miR-181b and up-regulation of miR-29a, miR-660, miR-20a, miR-106b, miR-142-5p, miR-101, miR-30b, miR-34a, miR-let-7f, miR-21 and miR-155. Among the miRNAs down-regulated in CLL cells, we showed that miR-181a/b expression levels were significantly lower in poor prognostic subgroups defined by unmutated immunoglobulin heavy chain variable status and p53 aberrations. Furthermore, under-expression of miR-181a and miR-181b was associated with shorter overall survival and treatment-free survival in CLL patients. We further evaluated fludarabine-induced apoptosis after transfection of primary CLL cells from 40 patients with miR-15a, miR-16-1, miR-34a, miR-181a and miR-181b mimics. Transfection of miR-34a, miR-181a and miR-181b mimics into CLL cells from p53 wild-type patients led to significant increase in apoptosis compared with miRNA control. However, enforced expression of these miRNAs had no effect on B-CLL cells from p53-attenuated patients. We further demonstrated that miR-181a and miR-181b inhibiting BCL-2, MCL-1 and X-linked inhibitor of apoptosis protein by direct binding to 3'UTR. Thus, these results suggest that miR-181a/b may play important roles in the pathogenesis of CLL and may provide a possible therapeutic avenue and a sensitive indicator of the activity of the p53 axis in CLL.     

Low expression of microRNA-204 (miR-204) is associated with poor clinical outcome of acute myeloid leukemia (AML) patients

Background

Acute myeloid leukemia (AML) is a heterogeneous neoplasm of the bone marrow with poor prognosis. In clinical practice new prognostic factors are still needed. MicroRNAs (miRs), small endogenous noncoding RNAs, play an essential role in the development and progression of acute leukemia. The aim of the study was to evaluate miR-204 expression in patients with AML at diagnosis and after induction chemotherapy, in comparison to healthy controls. We also investigated, if miR-204 expression correlates with clinical features of AML patients.

Methods

miR-204 expression has been analyzed using RT-PCR in 95 bone marrow specimens from newly diagnosed AML patients in comparison to 20 healthy subject.

Results

We showed down-regulated miR-204 expression in AML patients, which was associated with shorter patients’ survival. Higher expression of miR-204 in patients after induction therapy was correlated with complete remission achieving.

Conclusions

We showed low miR-204 expression in AML and found it to be an independent prognostic factor in this patient population.     

miR-181通过Wnt/β-catenin信号通路调控对B-ALL细胞增殖及凋亡的影响

目的:探究miR-181在急性B淋巴细胞白血病Nalm-6细胞中表达及其过表达对细胞增殖及凋亡的影响。方法:利用实时荧光定量PCR检测miR-181在急性B淋巴细胞白血病患者骨髓单个核细胞中的表达水平,利用慢病毒转染技术使miR-181过表达,利用CCK-8实验检测过表达miR-181对人急性 B 淋巴细胞白血病细胞Nalm-6增殖的影响,利用流式细胞仪检测过表达miR-181对Nalm-6细胞凋亡的影响,利用蛋白印迹实验检测Wnt/β-catenin信号通路中关键蛋白的表达水平。结果:实时荧光定量PCR结果显示,与正常人相比,急性B淋巴细胞白血病患者骨髓单个核细胞中miR-181的表达水平显著下调;CCK-8检测细胞增殖结果显示,过表达miR-181显著抑制了人急性 B 淋巴细胞白血病细胞Nalm-6的增殖;流式细胞仪检测细胞凋亡结果显示,过表达miR-181显著促进了Nalm-6细胞的凋亡;Western blotting 结果显示,miR-181过表达,Wnt1、β-catenin、cyclin D1 和c-myc蛋白的表达均显著下调。结论:急性B淋巴细胞白血病患者骨髓单个核细胞中miR-181的表达水平下调,过表达miR-181抑制Nalm-6细胞增殖、促进Nalm-6细胞凋亡,其机制可能与Wnt/β-catenin信号转导通路的受抑有关。     

Increased Risk of Thai Childhood Acute Lymphoblastic Leukemia with the MiR196a2 T>C Polymorphism

Objectives: This study assessed associations of the miR196a2 (rs11614913) T>C polymorphism withsusceptibility to childhood acute lymphoblastic leukemia (ALL) and clinical outcomes. Materials and Methods: Blood DNA samples from 104 childhood ALL patients and 180 healthy children were studied for the miR-196a2 (rs11614913) polymorphism using a polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP) approach. Results: The frequency of the miR-196a2 (rs11614913) T allele in controls was 0.51 compared with 0.33 in ALL cases. In this study, CC, TC heterozygote and CC/TC genotypes were significantly associated with increase childhood ALL susceptibility compared with the TT wild type (OR =4.321, 95% CI = 2.091-8.930 p=0.000, OR = 2.248, 95% CI =1.103-4.579, p=0.024, OR = 2.921, 95% CI = 1.504-5.673 p=0.001, respectively). However, the miR-196a2 (rs11614913) T>C polymorphism was not associated with demographic data or clinico-pathological data in ALL cases. Conclusion: CC, TC and CC+TC genotypes of miR-196a2 (rs11614913) was significantly associated with increased susceptibility in Thai childhood ALL but not with clinical variables.     

MicroRNA-145和KLF5在多发性骨髓瘤骨髓液中的表达及其临床意义

目的:分析微小RNA-145(microRNA-145,miR-145）和Krüpple 样因子5(Krüepple-like factor 5,KLF5)在多发性骨髓瘤（multiple myeloma,MM）骨髓液中的表达及其临床意义。方法:收集2015年1月至2016年3月住院MM诊治患者65例（疾病组）和体检健康正常者36例（对照组）作为研究对象,采用实时荧光定量PCR（qRT-PCR）法检测骨髓液中miR-145 mRNA和KLF5 mRNA表达水平;采用Western Blot检测KLF5蛋白表达水平;分析miR-145与KLF5 mRNA在MM髓液中表达的相关性。结果:miR-145 mRNA在MM骨髓液中的表达水平显著低于对照组（P<0.05）;KLF5 mRNA及蛋白表达水平显著高于对照组（P<0.05）;与I期、Ⅱ期比较,Ⅲ期MM患者骨髓液中miR-145表达水平显著降低（P<0.05）,KLF5 mRNA表达水平显著升高（P<0.05）;与IgG 型和IgA 型相比,轻链型与非分泌型MM患者骨髓液中miR-145相对表达量明显降低（P<0.05）;KLF5 mRNA在不同免疫分型中的表达水平比较,差异无统计学意义（P>0.05）;miR-145和KLF5的表达呈负相关（r=-0.452,P<0.05）;死亡MM患者骨髓液中miR-145表达水平显著低于存活患者（P<0.05）,KLF5 mRNA表达水平显著高于存活患者（P<0.05）。结论:miR-145在MM患者骨髓液中低表达,KLF5高表达,二者表达可能与MM的发生和病情发展密切相关。     

BCL6、KLF5、NCL基因在儿童急性淋巴细胞白血病中异常表达的特点

目的:研究转录因子基因BCL6、KLF5及核仁蛋白基因NCL在急性淋巴细胞白血病(acute lymphoblastic leukemia,ALL)患儿骨髓细胞中的表达情况及其在不同疾病状态下的表达特点。方法:选取北京儿童医院2004年1月至2005年12月住院ALL患儿100例,另取由于骨骼畸形而在北京儿童医院进行外科手术的5例非ALL患儿作对照;以基因芯片检测ALL患儿骨髓细胞中异常表达基因,GeXP多重基因表达分析系统检测BCL6、KLF5、NCL基因在另外选取的10例配对ALL患儿初诊及缓解期的表达变化。结果:基因芯片筛查发现,在100例各亚型ALL标本中,BCL6和KLF5mRNA表达均下调,NCLmRNA表达均上调。BCL6和KLF5mRNA在10例ALL初诊患儿骨髓细胞中表达较低,完全缓解后表达升高(0.380±0.16vs0.850±0.10,0.074±0.021vs0.228±0.049;均P<0.01);NCLmRNA在ALL初诊患儿骨髓细胞中表达较高,完全缓解后表达降低(0.234±0.054vs0.151±0.055,P<0.01)。在10例配对患者儿中,TEL-AML1阳性及E2A...     

miR-128在急性淋巴细胞白血病中的表达

目的 探讨miR-128在急性淋巴细胞白血病(ALL)中的表达及其意义.方法 采用实时荧光定量聚合酶链反应法对62例ALL患者和20例骨髓正常患者骨髓单个核细胞miR-128的相对表达量进行检测,并与患者的临床资料进行相关性分析.结果 miR-128在ALL患者中的相对表达量高于健康对照组(P＜0.05),但其在急性B淋巴细胞白血病(B-ALL)及急性T淋巴细胞白血病(T-ALL)中的表达水平差异无统计学意义(P＞0.05),miR-128的表达与是否存在bcr-abl融合基因及其mRNA的表达水平均无相关性(均P＞ 0.05).结论 miR-128在ALL患者中表达上调,可作为临床诊断ALL的潜在分子标志物,bcr-abl融合基因的表达对miR-128在ALL中的表达无明显影响.     

MicroRNA-27b,microRNA-101 and microRNA-128 inhibit angiogenesis by down-regulating vascular endothelial growth factor C expression in gastric cancers

Vascular Endothelial Growth Factor C (VEGF-C) has critical roles in angiogenesis in human cancers; however, the underlying mechanisms regulating VEGF-C expression remain largely unknown. In the present study, VEGF-C protein expression and the density of blood vessels or lymphatic vessels were determined by immunohistochemistry in 103 cases of gastric cancer tissues. Suppression of VEGF-C by miR-27b, miR-101 and miR-128 was investigated by luciferase assays, Western blot and ELISA. The miRNAs expression levels were detected in human gastric cancers by real-time quantitative PCR. Cell proliferation, migration and invasion assays were performed to assess the effect of miRNAs on gastric cancer cells and human umbilical vascular endothelial cells (HUVECs). Our data showed that high VEGF-C expression was significantly associated with increased tumor size, advanced TNM classification and clinical stage, higher microvessel density (MVD) and lymphatic density (LVD), as well as poor survival in patients with gastric cancer. Furthermore, VEGF-C was found to be a direct target gene of miR-27b, miR-101, and miR-128. The expression levels of the three miRNAs were inversely correlated with MVD. Overexpression of miR-27b, miR-101, or miR-128 suppressed migration, proliferation activity, and tube formation in HUVECs by repressing VEGF-C secretion in gastric cancer cells. We conclude that miR-27b, miR-101 and miR-128 inhibit angiogenesis by down-regulating VEGF-C expression in gastric cancers.     

儿童急性粒细胞白血病及其亚型的微小RNA表达

目的 分析儿童急性白血病(AL)的微小RNA(miRNA)表达谱,深化对AL的分型诊断乃至发病机制和预后的认识.方法 应用基因芯片技术分析93例AL患儿骨髓的miRNA表达谱,另选12例儿童特发性血小板减少性紫癜作为对照,应用实时定量聚合酶链反应(PCR)方法印证miRNA的异常表达.结果 粒细胞系的急性髓细胞白血病(AML)与急性淋巴细胞白血病的miRNA 表达谱明显不同,前者分别有17种和18种miRNA明显表达上调和下调,后者分别有21种和11种表达上调和下调.根据法、美、英协作组(FAB)形态学分型标准,在粒系AML的M1、M2和M3亚型中,miRNA表达各异,分别有miR-335、miR-126和miR-125b特异性高表达,基因芯片能区分这3种亚型.在M2和M3亚型中,AML1 -ETO阳性组和PML-RARα阳性组的miR-126和miR-125b表达水平,分别比阴性组明显升高(均P＜0.01).miR-335和miR-146在粒系AML表达上调,不同于在成人患者中研究报道的结果.结论 miRNA可能为AL的分类和诊断提供新的生物学标记.各型AL之间有不同的miRNA调控网络,可能存在与特异的miRNA表达相关的不同发病机制和预后.     

NET1 Enhances Proliferation and Chemoresistance in Acute Lymphoblastic Leukemia Cells

Acute lymphoblastic leukemia (ALL) is the most prevalent of pediatric cancers. Neuroepithelial cell-transforming 1 (NET1) has been associated with malignancy in a number of cancers, but the role of NET1 in ALL development is unclear. In the present study, we investigated the effect of NET1 gene in ALL cell proliferation and chemoresistance. We analyzed GEO microarray data comparing bone marrow expression profiles of pediatric B-cell ALL samples and those of age-matched controls. MTT and colony formation assays were performed to analyze cell proliferation. ELISA assays, Western blot analyses, and TUNEL staining were used to detect chemoresistance. We confirmed that NET1 was targeted by miR-206 using Western blot and luciferase reporter assays. We identified NET1 gene as one of the most significantly elevated genes in pediatric B-ALL. MTT and colony formation assays demonstrated that NET1 overexpression increases B-ALL cell proliferation in Nalm-6 cells. ELISA assays, Western blot analyses, and TUNEL staining showed that NET1 contributes to ALL cell doxorubicin resistance, whereas NET1 inhibition reduces resistance. Using the TargetScan database, we found that several microRNAs (miRNAs) were predicted to target NET1, including microRNA-206 (miR- 206), which has been shown to regulate cancer development. To determine whether miR-206 targets NET1 in vitro, we transfected Nalm-6 cells with miR-206 or its inhibitor miR-206-in. Western blot assays showed that miR-206 inhibits NET1 expression and miR-206-in increases NET1 expression. Luciferase assays using wild-type or mutant 3 -untranslated region (3 -UTR) of NET1 confirmed these findings. We ultimately found that miR-206 inhibits B-ALL cell proliferation and chemoresistance induced by NET1. Taken together, our results provide the first evidence that NET1 enhances proliferation and chemoresistance in B-ALL cells and that miR-206 regulates these effects by targeting NET1. This study therefore not only contributes to a greater understanding of the molecular mechanisms underlying B-ALL progression but also opens the possibility for developing curative interventions.     

Role of microRNA dysregulation in childhood acute leukemias: Diagnostics,monitoring and therapeutics: A comprehensive review

MicroRNAs (miRNAs) are short noncoding RNAs that regulate the expression of genes by sequence-specific binding to mRNA to either promote or block its translation; they can also act as tumor suppressors (e.g., let-7b, miR-29a, miR-99, mir-100, miR-155, and miR-181) and/or oncogenes (e.g., miR-29a, miR-125b, miR-143-p3, mir-155, miR-181, miR-183, miR-196b, and miR-223) in childhood acute leukemia (AL). Differentially expressed miRNAs are important factors associated with the initiation and progression of AL. As shown in many studies, they can be used as noninvasive diagnostic and prognostic biomarkers, which are useful in monitoring early stages of AL development or during therapy (e.g., miR-125b, miR-146b, miR-181c, and miR-4786), accurate classification of different cellular or molecular AL subgroups (e.g., let-7b, miR-98, miR-100, miR-128b, and miR-223), and identification and development of new therapeutic agents (e.g., mir-10, miR-125b, miR-203, miR-210, miR-335). Specific miRNA patterns have also been described for commonly used AL therapy drugs (e.g., miR-125b and miR-223 for doxorubicin, miR-335 and miR-1208 for prednisolone, and miR-203 for imatinib), uncovering miRNAs that are associated with treatment response. In the current review, the role of miRNAs in the development, progression, and therapy monitoring of pediatric ALs will be presented and discussed.