Hsa-miR-1246, hsa-miR-320a and hsa-miR-196b-5p inhibitors can reduce the cytotoxicity of Ebola virus glycoprotein in vitro

Ebola virus(EBOV)causes a highly lethal hemorrhagic fever syndrome in humans and has been associated with mortality rates of up to 91%in Zaire,the most lethal strain.Though the viral envelope glycoprotein(GP)mediates widespread inflammation and cellular damage,these changes have mainly focused on alterations at the protein level,the role of microRNAs(miRNAs)in the molecular pathogenesis underlying this lethal disease is not fully understood.Here,we report that the miRNAs hsa-miR-1246,hsa-miR-320a and hsa-miR-196b-5p were induced in human umbilical vein endothelial cells(HUVECs)following expression of EBOV GP.Among the proteins encoded by predicted targets of these miRNAs,the adhesion-related molecules tissue factor pathway inhibitor(TFPI),dystroglycan1(DAG1)and the caspase 8 and FADD-like apoptosis regulator(CFLAR)were significantly downregulated in EBOV GP-expressing HUVECs.Moreover,inhibition of hsa-miR-1246,hsa-miR-320a and hsa-miR-196b-5p,or overexpression of TFPI,DAG1 and CFLAR rescued the cell viability that was induced by EBOV GP.Our results provide a novel molecular basis for EBOV pathogenesis and may contribute to the development of strategies to protect against future EBOV pandemics.     

Expression Analyses of MicroRNAs in Hamster Lung Tissues Infected by SARS-CoV-2

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an infectious disease with multiple severe symptoms, such as fever over 37.5°C, cough, dyspnea, and pneumonia. In our research, microRNAs (miRNAs) binding to the genome sequences of severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory-related coronavirus (MERS-CoV), and SARS-CoV-2 were identified by bioinformatic tools. Five miRNAs (hsa-miR-15a-5p, hsa-miR-15b-5p, hsa-miR-195-5p, hsa-miR-16-5p, and hsa-miR-196a-1-3p) were found to commonly bind to SARS-CoV, MERS-CoV, and SARS-CoV-2. We also identified miRNAs that bind to receptor proteins, such as ACE2, ADAM17, and TMPRSS2, which are important for understanding the infection mechanism of SARS-CoV-2. The expression patterns of those miRNAs were examined in hamster lung samples infected by SARS-CoV-2. Five miRNAs (hsa-miR-15b-5p, hsa-miR-195-5p, hsa-miR-221-3p, hsa-miR-140-3p, and hsa-miR-422a) showed differential expression patterns in lung tissues before and after infection. Especially, hsa-miR-15b-5p and hsa-miR-195-5p showed a large difference in expression, indicating that they may potentially be diagnostic biomarkers for SARS-CoV-2 infection.     

Circ-U2AF1 promotes human glioma via derepressing neuro-oncological ventral antigen 2 by sponging hsa-miR-7-5p

The prognosis for human glioma, a malignant tumor of the central nervous system, is poor due to its rapid growth, genetic heterogeneity, and inadequate understanding of its underlying molecular mechanisms. Circular RNAs composed of exonic sequences, represent an understudied form of noncoding RNAs (ncRNAs) that was discovered more than a decade ago, function as microRNA sponges. We aimed to assess the relationship between circ-U2AF1 (CircRNA ID: hsa_circ_0061868) and hsa-mir-7-5p and examine their effects on proliferation, apoptosis, and the metastatic phenotype of glioma cells regulated by neuro-oncological ventral antigen 2 (NOVA2). We found that the expression levels of circ-U2AF1 and NOVA2 were upregulated, while hsa-miR-7-5p was downregulated in human glioma tissues and glioma cell lines. Our data and bioinformatic analysis indicated the association of these molecules with glioma grade, a positive correlation between circ-U2AF1 and NOVA2 expression levels and a negative correlation of hsa-miR-7-5p with both circ-U2AF1 and NOVA2, respectively. In addition, silencing of circ-U2AF1 expression resulted in increased hsa-miR-7-5p expression and decreased NOVA2 expression both in vitro and in vivo. Luciferase assay confirmed hsa-miR-7-5p as a direct target of circ-U2AF1 and NOVA2 as a direct target of hsa-miR-7-5p. Functionally, silencing of circ-U2AF1 inhibits glioma development by repressing NOVA2 via upregulating hsa-miR-7-5p both in vitro and in vivo. Thus, we assumed that circ-U2AF1 promotes glioma malignancy via derepressing NOVA2 by sponging hsa-miR-7-5p. Taken together, we suggest that circ-U2AF1 can be a prognostic biomarker and the circ-U2AF1/hsa-miR-7-5p/NOVA2 regulatory pathway may be a novel therapeutic target for treating gliomas.     

LncRNA RP1-86C11.7 exacerbates the glioma progression and oncogenicity by hsa-miR-144-3p/TFRC signaling.

Glioblastoma (GBM) remains the most common and malignant tumor of the human central nervous system. Increasing evidence has highlighted that tumor cells with high transferrin receptor (TFRC) expression show advantages in growth. Long noncoding RNAs (lncRNAs) are related to glioma progression by mediating microRNAs (miRNAs). However, the underlying mechanism among TFRC, miRNA and lncRNA in GBM is limited. In the current study, we identified a new lncRNA-induced signaling mechanism that regulates the TFRC levels in GBM. The TFRC level was higher in glioma cell lines, and elevated TFRC expression promoted the proliferation and survival of glioma cells. Further study showed that hsa-miR-144a-3p bound to the 3′-UTR of TFRC mRNA and inhibited its expression, preventing the malignant properties of glioma cells, such as proliferation and survival. We also found that the lncRNA RP1-86C11.7 sponges hsa-miR-144-3p to suppress its protective role in glioma. RP1-86C11.7 overexpression in glioma cells elevated TFRC expression, increased the intracellular free iron level, and deteriorated oncogenicity, with a significant reduction in hsa-miR-144-3p. By contrast, silencing RP1-86C11.7 upregulated the hsa-miR-144-3p level, resulting in decreased TFRC expression and repressed glioma progression. However, the effect of silencing RP1-86C11.7 was reversed with simultaneous hsa-miR-144-3p inhibitor treatment: the TFRC level, intracellular iron level and proliferation in glioma cells increased. Mechanistically, our data indicated that RP1-86C11.7 exacerbates the malignant behavior of glioma through the hsa-miR-144-3p/TFRC axis. RP1-86C11.7 may be a potential biomarker or target to treat glioma in the future.     

LncRNA RP1-86C11.7 exacerbates the glioma progression and oncogenicity by hsa-miR-144-3p/TFRC signaling.

Glioblastoma (GBM) remains the most common and malignant tumor of the human central nervous system. Increasing evidence has highlighted that tumor cells with high transferrin receptor (TFRC) expression show advantages in growth. Long noncoding RNAs (lncRNAs) are related to glioma progression by mediating microRNAs (miRNAs). However, the underlying mechanism among TFRC, miRNA and lncRNA in GBM is limited. In the current study, we identified a new lncRNA-induced signaling mechanism that regulates the TFRC levels in GBM. The TFRC level was higher in glioma cell lines, and elevated TFRC expression promoted the proliferation and survival of glioma cells. Further study showed that hsa-miR-144a-3p bound to the 3′-UTR of TFRC mRNA and inhibited its expression, preventing the malignant properties of glioma cells, such as proliferation and survival. We also found that the lncRNA RP1-86C11.7 sponges hsa-miR-144-3p to suppress its protective role in glioma. RP1-86C11.7 overexpression in glioma cells elevated TFRC expression, increased the intracellular free iron level, and deteriorated oncogenicity, with a significant reduction in hsa-miR-144-3p. By contrast, silencing RP1-86C11.7 upregulated the hsa-miR-144-3p level, resulting in decreased TFRC expression and repressed glioma progression. However, the effect of silencing RP1-86C11.7 was reversed with simultaneous hsa-miR-144-3p inhibitor treatment: the TFRC level, intracellular iron level and proliferation in glioma cells increased. Mechanistically, our data indicated that RP1-86C11.7 exacerbates the malignant behavior of glioma through the hsa-miR-144-3p/TFRC axis. RP1-86C11.7 may be a potential biomarker or target to treat glioma in the future.     

A genetic variant in microRNA-196a2 is associated with increased cancer risk: a meta-analysis

MicroRNAs (miRNAs) are small non-coding RNA molecules that function as negative regulators of gene expression. Common genetic variants (single nucleotide polymorphisms, SNPs) in miRNA genes may alter their expression or maturation resulting in varied functional consequences. Until now, several studies had evaluated the association between the polymorphisms in the hsa-miR-196a2 rs11614913 and cancer risk in diverse populations and in multiple types of cancer, with contradictory outcomes. Therefore, here we performed a meta-analysis to address the association between this polymorphism and cancer risk. A total of nine studies involving 6,540 cases and 7,562 controls were retrieved based on PubMed. Our analysis demonstrated that hsa-miR-196a2 rs11614913 CC genotype significantly increased the cancer risk in homozygote comparison model compared to TT genotype (OR = 1.18; 95% CI, 1.01–1.68). Moreover, significant association of this polymorphism with breast cancer was found based on homozygote comparison model (OR = 1.30; 95% CI, 1.01–1.26) and dominant model (OR = 1.11; 95% CI, 1.01–1.23). In addition, hsa-miR-196a2 rs11614913 CC genotype was significantly associated with cancer risk in Chinese and Indian (OR = 1.21; 95% CI, 1.05–1.40), but not in Caucasians (OR = 1.03; 95% CI, 0.89–1.19). Taken together, our results indicate that the polymorphism of hsa-miR-196a2 rs11614913 is associated with cancer susceptibility, especially with breast cancer and in Chinese and Indian populations.     

Hsa-miR-34a-5p reverses multidrug resistance in gastric cancer cells by targeting the 3′-UTR of SIRT1 and inhibiting its expression

Multidrug resistance (MDR) is a major obstacle to chemotherapy, which leads to ineffective chemotherapy, an important treatment strategy for gastric cancer (GC). The abnormality of microRNAs (miRNAs) is critical to the occurrence and progression of MDR in various tumors. In this study, hsa-miR-34a-5p was found to be decreased in multidrug resistant GC cells SGC-7901/5-Fluorouracil (SGC-7901/5-Fu) compared to the parental SGC-7901 cells. Overexpression of hsa-miR-34a-5p in SGC-7901/5-Fu cells promoted apoptosis and decreased migration and invasiveness after chemotherapy. In addition, overexpression of hsa-miR-34a-5p suppressed the growth of drug-resistant tumor in vivo. The mechanism of the effects of hsa-miR-34a-5p could include the regulation of the expression of Sirtuin 1 (SIRT1), P-glycoprotein (P-gp) or Multidrug resistance-related protein 1 (MRP1) through direct binding to the 3′-untranslated region (UTR) of SIRT1. Functional gain-and-loss experiments indicated that hsa-miR-34a-5p enhances the chemotherapy sensitivity of MDR GC cells by inhibiting SIRT1, P-gp and MRP1. In conclusion, hsa-miR-34a-5p can reverse the MDR of GC cells by inhibiting the expression of SIRT1, P-gp or MRP1.     

The diagnostic and prognostic values of microRNA-196a in cancer

MicroRNA-196a (miR-196a) was previously reported to be up-regulated in cancers, and it has the diagnostic and prognostic values in cancers. Whereas, the conclusion was still unclear according to the published data. To assess such roles of miR-196a in cancers, the present study was conducted based on published data and online cancer-related databases. To identify the relevant published data, we searched articles in databases and then the relevant data were extracted to evaluate the correlation between miR-196a expression and diagnosis, prognosis for cancer patients. The pooled results showed that miR-196a was a valuable diagnostic biomarker in cancer (area under curve (AUC) = 0.87, 95% CI: 0.84–0.90; sensitivity (SEN) = 0.73, 95% CI: 0.64–0.81; specificity (SPE) = 0.90, 95% CI: 0.81–0.95), which was consistent with the data from databases (breast cancer: miR-196a-3p: AUC = 0.77, 95% CI: 0.74–0.79; miR-196a-5p: AUC = 0.71, 95% CI: 0.66–0.75; pancreatic cancer: miR-196a-3p: AUC = 0.80, 95% CI: 0.73–0.87; miR-196a-5p: AUC = 0.61, 95% CI: 0.51–0.71). In addition, the pooled result revealed that elevated miR-196a expression in tumor tissues (HR = 2.54, 95% CI: 1.79–3.61, P_{Heterogeneity}=0.000, I² = 75.8%) or serum/plasma (HR = 4.06, 95% CI: 2.67–6.18, P_{Heterogeneity}=0.668, I² = 0%) of patients was an unfavorable survival biomarker, which was consistent with the data from databases (adrenocortical carcinoma: HR = 5.70; esophageal carcinoma: HR = 1.93; brain lower grade glioma: HR = 2.91; {"type":"entrez-geo","attrs":{"text":"GSE40267","term_id":"40267"}}GSE40267: HR = 2.47, 95% CI: 1.2–5.07; TCGA: HR = 1.82, 95% CI: 1.21–2.74; {"type":"entrez-geo","attrs":{"text":"GSE19783","term_id":"19783"}}GSE19783: HR = 4.24, 95% CI: 1–18.06). In short, our results demonstrated that miR-196a in tumor tissue or serum/plasma could be used as a prognostic and diagnostic values for cancers.     

Comprehensive analysis of the functional microRNA–mRNA regulatory network identifies miRNA signatures associated with glioma malignant progression

Glioma is the most common and fatal primary brain tumour with poor prognosis; however, the functional roles of miRNAs in glioma malignant progression are insufficiently understood. Here, we used an integrated approach to identify miRNA functional targets during glioma malignant progression by combining the paired expression profiles of miRNAs and mRNAs across 160 Chinese glioma patients, and further constructed the functional miRNA–mRNA regulatory network. As a result, most tumour-suppressive miRNAs in glioma progression were newly discovered, whose functions were widely involved in gliomagenesis. Moreover, three miRNA signatures, with different combinations of hub miRNAs (regulations≥30) were constructed, which could independently predict the survival of patients with all gliomas, high-grade glioma and glioblastoma. Our network-based method increased the ability to identify the prognostic biomarkers, when compared with the traditional method and random conditions. Hsa-miR-524-5p and hsa-miR-628-5p, shared by these three signatures, acted as protective factors and their expression decreased gradually during glioma progression. Functional analysis of these miRNA signatures highlighted their critical roles in cell cycle and cell proliferation in glioblastoma malignant progression, especially hsa-miR-524-5p and hsa-miR-628-5p exhibited dominant regulatory activities. Therefore, network-based biomarkers are expected to be more effective and provide deep insights into the molecular mechanism of glioma malignant progression.     

Overexpressed miR-196a accelerates osteogenic differentiation in osteoporotic mice via GNAS-dependent Hedgehog signaling pathway

Osteoporosis (OP), a common metabolic bone disease, is accompanied by reduced bone mass, bone mineral density (BMD), as well as microstructure destruction of bone. Previously, microRNA-196a-2 (miR-196a-2) and miR-196a-3p were reported for its involvement in BMD. Herein, this study set out to identify the functional relevance of miR-196a in osteogenic differentiation in osteoporotic mice and explore the associated mechanism by establishing an OP mouse model. Guanine nucleotide binding protein, alpha stimulating (GNAS) was verified as a target gene of miR-196a, which was decreased in OP mice. Furthermore, the bone marrow stromal cells (BMSCs) were then extracted from OP mice and treated with miR-196 mimic/inhibitor or small interfering RNA against GNAS to investigate miR-196a interaction with GNAS and the Hedgehog signaling pathway. BMSCs in OP mice transfected with miR-196a mimic or si-GNAS displayed the elevated expression of Smo, ALP, Runx2, and OPN, as well as bone gla protein and tartrate-resistant acid phosphatase, elevated ALP vitality and bone formation ability as well as reduced expression of GNAS and PTCH. Taken conjointly, overexpression of miR-196a repressed GNAS expression by activating the Hedgehog signaling pathway, thus promoting osteogenic differentiation in mice with OP.     

p53/miR-30a-5p/ SOX4 feedback loop mediates cellular proliferation,apoptosis, and migration of non-small-cell lung cancer

Many microRNAs (miRNAs) play vital roles in the tumorigenesis and development of cancers. In this study, we aimed to identify the differentially expressed miRNAs and their specific mechanisms in non-small-cell lung cancer (NSCLC). Based on data from the GSE56036 database, miR-30a-5p expression was identified to be downregulated in NSCLC. Further investigations showed that overexpression of miR-30a-5p inhibited cell proliferation, migration, and promoted apoptosis in NSCLC. Increase of miR-30a-5p level could induce the increase of Bax protein level and decrease of Bcl-2 protein level. In addition, chromatin immunoprecipitation assays showed that miR-30a-5p expression was induced by binding of p53 to the promoter of MIR30A. Bioinformatics prediction indicated that miR-30a-5p targets SOX4, and western blot analysis indicated that overexpression of the miRNA decreases the SOX4 protein expression level, which in turn regulated the level of p53. Thus, this study provides evidence for the existence of a p53/miR-30a-5p/SOX4 feedback loop, which likely plays a key role in the regulation of proliferation, apoptosis, and migration in NSCLC, highlighting a new therapeutic target.     

脑胶质瘤组织中p16基因启动子区CpG岛甲基化检测及其临床意义研究

目的：研究脑胶质瘤中p16基因启动子区甲基化情况及其临床意义。方法：用甲基化特异性PCR技术检测42例脑胶质瘤组织和癌旁正常脑组织中p16基因启动子甲基化,并分析该基因启动子甲基化与临床病理特征之间的关系。结果：脑胶质瘤组织中p16基因异常甲基化率（38.27%）显著高于癌旁正常脑组织中p16基因的异常甲基化率（8.8%,P=0.000）。发生甲基化的肿瘤组织或者正常脑组织中p16基因mRNA和蛋白表达显著降低。此外,p16基因异常甲基化和肿瘤病理分级有相关性（P=0.007）,而与患者性别、年龄及肿瘤类型等临床特征无关（P=0.669,0.869和0.944）。结论：p16基因启动子区CpG岛高甲基化与p16表达下调相关,推测p16启动子区CpG岛高甲基化是导致p16基因在脑胶质瘤中表达下调的重要因素,有望成为脑胶质瘤早期辅助诊断的分子标志物之一。     

Dynamic expression of viral and cellular microRNAs in infectious mononucleosis caused by primary Epstein-Barr virus infection in children

Background

Epstein-Barr virus (EBV) was the first virus identified to encode microRNAs (miRNAs). Both of viral and human cellular miRNAs are important in EBV infection. However, the dynamic expression profile of miRNAs during primary EBV infection was unknown. This study aimed to investigate the dynamic expression profile of viral and cellular miRNAs in infectious mononucleosis (IM) caused by primary EBV infection.

Methods

The levels of viral and cellular miRNAs were measured in fifteen pediatric IM patients at three different time-points. Fifteen healthy children who were seropositive for EBV were enrolled in the control group. Relative expression levels of miRNAs were detected by quantitative real-time PCR (qPCR) assay.

Results

EBV-miR-BHRF1-1, 1-2-3P, miR-BART13-1, 19-3p, 11-3P, 12–1, and 16–1 in IM patients of early phase were significantly higher than in healthy children. Most cellular miRNAs of B cells, such as hsa-miR-155-5p, ?34a-5p, ?18b-5p, ?181a-5p, and ?142-5p were up-regulated; while most of cellular miRNAs of CD8?+?T cells, such as hsa-miR-223, ?29c-3p, ?181a, ?200a-3p, miR-155-5p, ?146a, and ?142-5p were down-regulated in IM patients. With disease progression, nearly all of EBV-miRNAs decreased, especially miR-BHRF1, but at a slower rate than EBV DNA loads. Most of the cellular miRNAs of B cells, including hsa-miR-134-5p, ?18b-5p, ?34a-5p, and -196a-5p increased with time. However, most of the cellular miRNAs of CD8?+?T cells, including hsa-let-7a-5p, ?142-3p, ?142-5p, and ?155-5p decreased with time. Additionally, hsa-miR-155-5p of B cells and hsa-miR-18b-5p of CD8+ T cells exhibited a positive correlation with miR-BHRF1-2-5P and miR-BART2-5P (0.96?≤?r?≤?0.99, P?<?0.05). Finally, hsa-miR-181a-5p of B cells had positive correlation with miR-BART4-3p, 4-5P, 16–1, and 22 (0.97?≤?r?≤?0.99, P?<?0.05).

Conclusions

Our study is the first to describe the expression profile of viral and cellular miRNAs in IM caused by primary EBV infection. These results might be the basis of investigating the pathogenic mechanism of EBV-related diseases and bring new insights into their diagnosis and treatment.

     

Hsa_circ_0110757 upregulates ITGA1 to facilitate temozolomide resistance in glioma by suppressing hsa-miR-1298-5p

Temozolomide (TMZ) is the internationally recognized and preferred drug for glioma chemotherapy treatment. However, TMZ resistance in glioma appears after long-term use and is an urgent problem that needs to be solved. Circular RNAs (circRNAs) are noncoding RNAs and play an important role in the pathogenesis and progression of tumors. Hsa_circ_0110757 was identified in TMZ-resistant glioma cells by high-throughput sequencing analysis and was derived from reverse splicing of myeloid cell leukemia-1 (Mcl-1) exons. The role of hsa_circ_0110757 in TMZ-resistant glioma was evaluated both in vitro and in vivo. It was found that hsa_circ_0110757 and ITGA1 are more highly expressed in TMZ-resistant glioma than in TMZ-sensitive glioma. The overexpression of hsa_circ_0110757 in glioma patients treated with TMZ was obviously associated with tumor invasion. This study indicates that hsa_circ_0110757 inhibits glioma cell apoptosis by sponging hsa-miR-1298-5p to promote ITGA1 expression. Thus, hsa_circ_0110757/hsa-miR-1298-5p/ITGA could be a potential therapeutic target for reversing the resistance of glioma to TMZ.Subject terms: Chemotherapy, Tumour-suppressor proteins     

microRNA-199a-3p,DNMT3A,and aberrant DNA methylation in testicular cancer

It was previously demonstrated that miR-199a was downregulated in testicular germ cell tumor (TGCT), probably due to hypermethylation of its promoter. Further study found that re-expression of miR-199a in testicular cancer cells (NT2) led to suppression of cell growth, cancer migration, invasion and metastasis. More detailed analyses showed that these properties of miR-199a could be assigned to miR-199a-5p, one of its two derivatives. The biological role of the other derivative, miR-199a-3p in TGCT, remains largely uncharacterized. In this report, we identified DNA (cytosine-5)-methyltransferase 3A (DNMT3A), the de novo methyltransferase, as a direct target of miR-199a-3p using a 3′-UTR reporter assay. Transient expression of miR-199a-3p in NT2 cells led to decrease, while knocking down of miR-199a-3p in a normal human testicular cell line (HT) led to elevation, of DNMT3A2 (DNMT3A gene isoform 2) mRNA and protein levels. In clinical samples, DNMT3A2 was significantly overexpressed in malignant testicular tumor, and the expression of DNMT3A2 was inversely correlated with the expression of miR-199a-3p. However, DNMT3A did not affect miR-199a expression in NT2 cells. Further characterization of miR-199a-3p revealed that it negatively regulated DNA methylation, partly through targeting DNMT3A. Overexpression of miR-199a-3p restored the expression of APC and MGMT tumor-suppressor genes in NT2 cells by affecting DNA methylation of their promoter regions. Our studies demonstrated the deregulation of miR-199a-3p expression in TGCT may provide novel mechanistic insights into TGCT carcinogenesis and suggested a potentially therapeutic use of synthetic miR-199a-3p oligonucleotides as effective hypomethylating compounds in the treatment of TGCT.     

MicroRNA miR-21 overexpression in human breast cancer is associated with advanced clinical stage, lymph node metastasis and patient poor prognosis

To investigate the global expression profile of miRNAs in primary breast cancer (BC) and normal adjacent tumor tissues (NATs) and its potential relevance to clinicopathological characteristics and patient survival, the genome-wide expression profiling of miRNAs in BC was investigated using a microarray containing 435 mature human miRNA oligonucleotide probes. Nine miRNAs of hsa-miR-21, hsa-miR-365, hsa-miR-181b, hsa-let-7f, hsa-miR-155, hsa-miR-29b, hsa-miR-181d, hsa-miR-98, and hsa-miR-29c were observed to be up-regulated greater than twofold in BC compared with NAT, whereas seven miRNAs of hsa-miR-497, hsa-miR-31, hsa-miR-355, hsa-miR-320, rno-mir-140, hsa-miR-127 and hsa-miR-30a-3p were observed to be down-regulated greater than twofold. The most significantly up-regulated miRNAs, hsa-mir-21 (miR-21), was quantitatively analyzed by TaqMan real-time PCR in 113 BC tumors. Interestingly, among the 113 BC cases, high level expression of miR-21 was significantly correlated with advanced clinical stage (P = 0.006, Fisher's exact text), lymph node metastasis (P = 0.007, Fisher's exact text), and shortened survival of the patients (hazard ratio [HR]=5.476, P < 0.001). Multivariate Cox regression analysis revealed this prognostic impact (HR=4.133, P = 0.001) to be independent of disease stage (HR=2.226, P = 0.013) and histological grade (HR=3.681, P = 0.033). This study could identify the differentiated miRNAs expression profile in BC and reveal that miR-21 overexpression was correlated with specific breast cancer biopathologic features, such as advanced tumor stage, lymph node metastasis, and poor survival of the patients, indicating that miR-21 may serve as a molecular prognostic marker for BC and disease progression.     

Epigenetic inactivation of the hsa-miR-203 in haematological malignancies

miR-203 is a tumour suppressor microRNA (miRNA). We studied the methylation of hsa-miR-203 in 150 samples including acute myeloid leukaemia (AML), acute lymphoblastic leukaemia (ALL), chronic myeloid leukaemia (CML), chronic lymphocytic leukaemia (CLL) and non-Hodgkin's lymphoma (NHL) by methylation-specific PCR, and miRNA expression by stem-loop RT-qPCR. hsa-miR-203 promoter was unmethylated in normal controls but homozygously methylated in two AML and four lymphoma cell lines, in which 5-Aza-2'-deoxycytidine treatment led to promoter demethylation and miR-203 re-expression. Restoration of miR-203 expression in lymphoma cells inhibited cellular proliferation and increased cell death, suggesting an inherent tumour suppressor activity. In primary samples, hsa-miR-203 methylation was absent in CML but detected in 5.0% ALL, 10.0% AML, 42.0% CLL and 38.8% of NHL (including six [60.0%] natural killer-cell, nine [40.9%] B-cell and four [23.5%] T cell NHL). Moreover, hsa-miR-203 methylation was associated with hypermethylation of hsa-miR-34a, -124a and -196b in NHL but not CLL. In CLL, hsa-miR-203 methylation was associated with a higher presenting Hb level (P = 0.033). The projected 10 year overall survival of the CLL patients was 58.2%, which was impacted by Rai stage and high-risk karyotypes but not hsa-miR-203 methylation. hsa-miR-203 was more frequently methylated in lymphoid than myeloid malignancies (P = 0.002). In conclusion, miR-203, a tumour suppressor gene, was hypermethylated in a tumour-specific manner with gene silencing. hsa-miR-203 was more frequently hypermethylated in lymphoid than myeloid malignancies. In NHL, hsa-miR-203 methylation was associated with concomitant methylation of other tumour suppressor miRNAs. The frequent hsa-miR-203 methylation in lymphoid malignancies suggested a pathogenetic role of hsa-miR-203 methylation.