Diagnostic and prognostic value of circulating miR-21 for cancer: A systematic review and meta-analysis

Background

MicroRNAs (miRNAs) have been reported to be aberrantly expressed in patients with cancer. Many studies have shown that circulating miRNAs could play potential roles as diagnostic and prognostic biomarkers of cancers. The aim of this meta-analysis is to summarize the role of circulating miR-21 as a biomarker in patients with a variety of carcinomas.

Material and methods

Eligible studies were identified and assessed for quality through multiple search strategies. For diagnostic meta-analysis, the sensitivity, specificity, and other measures of miR-21 in the diagnosis of cancer were pooled using bivariate random-effects approach models. For prognostic meta-analysis, pooled hazard ratios (HRs) of circulating miR-21 for survival were calculated.

Results

A total of 36 studies dealing with various carcinomas were included for the systemic review. Among them, 23 studies were finally enrolled in the global meta-analysis (17 studies for diagnosis and 6 studies for prognosis). For diagnostic meta-analysis, the overall pooled results for sensitivity, specificity, positive likelihood ratio (LRP), negative likelihood ratios (LRN) and diagnostic odds ratio (DOR) were 75.7% (95% CI: 67.1%–82.6%), 79.3% (95% CI: 74.2%–83.5%), 3.65 (95% CI: 2.83–4.70), 0.31 (95% CI: 0.22–0.43), and 11.88 (95% CI: 6.99–20.19), respectively. For prognostic meta-analysis, the pooled HR of higher miR-21 expression in circulation was 2.37 (95% CI: 1.83–3.06, P < 0.001), which could significantly predict poorer survival in general carcinomas. Importantly, subgroup analysis suggested that higher expression of miR-21 correlated with worse overall survival (OS) significantly in carcinomas of digestion system (HR, 5.77 [95% CI: 2.65–12.52]).

Conclusions

Our findings suggest that circulating miR-21 may not suitable to be a diagnostic biomarker, but it has a prognostic value in patients with cancer.     

Plasma miR-601 and miR-760 Are Novel Biomarkers for the Early Detection of Colorectal Cancer

Background

Colorectal cancer (CRC) is a major cause of death worldwide. Sensitive, non-invasive diagnostic screen methods are urgently needed to improve its survival rates. Stable circulating microRNA offers unique opportunities for the early diagnosis of several diseases, including cancers. Our aim has been to find new plasma miRNAs that can be used as biomarkers for the detection of CRC.

Methodology/Principal Findings

According to the results of miRNA profiling performed on pooling plasma samples form 10 CRC patients or 10 healthy controls, a panel of miRNAs (hsa-miR-10a, -19a, -22*, -24, -92a, 125a-5p, -141, -150, -188-3p, -192, -210, -221, -224*, -376a, -425*, -495, -572, -601, -720, -760 and hsa-let-7a, -7e) were deregulated in CRC plasma with fold changes >5. After large scale validation by qRT-PCR performed on another 191 independent individuals (90 CRC, 43 advanced adenoma and 58 healthy participants), we found that the levels of plasma miR-601 and miR-760 were significantly decreased in colorectal neoplasia (carcinomas and advanced adenomas) compared with healthy controls. ROC curve analysis showed that plasma miR-601 and miR-760 were of significant diagnostic value for advanced neoplasia. These two miRNAs together yield an AUC of 0.792 with 83.3% sensitivity and 69.1% specificity for separating CRC from normal controls, and yield an AUC of 0.683 with 72.1% sensitivity and 62.1% specificity in discriminating advanced adenomas from normal controls.

Conclusions/Significance

Plasma miR-601 and miR-760 can potentially serve as promising non-invasive biomarkers for the early detection of CRC.     

Downregulation of miR-122 in the rodent and human hepatocellular carcinomas

MicroRNAs (miRs) are conserved small non-coding RNAs that negatively regulate gene expression. The miR profiles are markedly altered in cancers and some of them have a causal role in tumorigenesis. Here, we report changes in miR expression profile in hepatocellular carcinomas (HCCs) developed in male Fisher rats-fed folic acid, methionine, and choline-deficient (FMD) diet. Comparison of the miR profile by microarray analysis showed altered expression of some miRs in hepatomas compared to the livers from age-matched rats on the normal diet. While let-7a, miR-21, miR-23, miR-130, miR-190, and miR-17-92 family of genes was upregulated, miR-122, an abundant liver-specific miR, was downregulated in the tumors. The decrease in hepatic miR-122 was a tumor-specific event because it did not occur in the rats switched to the folate and methyl-adequate diet after 36 weeks on deficient diet, which did not lead to hepatocarcinogenesis. miR-122 was also silent in a transplanted rat hepatoma. Extrapolation of this study to human primary HCCs revealed that miR-122 expression was significantly (P = 0.013) reduced in 10 out of 20 tumors compared to the pair-matched control tissues. These findings suggest that the downregulation of miR-122 is associated with hepatocarcinogenesis and could be a potential biomarker for liver cancers.     

Clinical significance of miR-140-5p and miR-193b expression in patients with breast cancer and relationship to IGFBP5

The functional role of IGFBP5 in breast cancer is complicated. Experimental and bioinformatics studies have shown that IGFBP5 is targeted by miR-140-5p and miR-193b, although this has not yet been proven in clinical samples. The aim of this study was to evaluate the expression of miR-140-5p and miR-193b in breast cancer and adjacent normal tissue and assess its correlation with IGFBP5 and the clinicopathological characteristics of the tumors. IGFBP5 protein expression was analyzed immunohistochemically and IGFBP5, miR-140 and miR-193b mRNA expression levels were analyzed with real-time RT-PCR. Tumor tissue had higher miR-140-5p expression than adjacent normal tissue (p = 0.015). Samples with no immunohistochemical staining for IGFBP5 showed increased miR-140-5p expression (p = 0.009). miR-140-5p expression was elevated in invasive ductal carcinomas (p = 0.002), whereas basal-like tumors had decreased expression of miR-140-5p compared to other tumors (p = 0.008). Lymph node-positive samples showed an approximately 13-fold increase in miR-140-5p expression compared to lymph node-negative tissue (p = 0.049). These findings suggest that miR-140-5p, but not miR-193b, could be an important determinant of IGFBP5 expression and clinical phenotype in breast cancer patients. Further studies are needed to clarify the expressional regulation of IGFBP5 by miR-140-5p.     

miR-122 Regulates Tumorigenesis in Hepatocellular Carcinoma by Targeting AKT3

MicroRNAs (miRNAs) have been implicated in the orchestration of diverse cellular processes including differentiation, proliferation, and apoptosis and are believed to play pivotal roles as oncogenes and tumor suppressors. miR-122, a liver specific miRNA, is significantly down-regulated in most hepatocellular carcinomas (HCCs) but its role in tumorigenesis remains poorly understood. Here we identify AKT3 as a novel and direct target of miR-122. Restoration of miR-122 expression in HCC cell lines decreases AKT3 levels, inhibits cell migration and proliferation, and induces apoptosis. These anti-tumor phenotypes can be rescued by reconstitution of AKT3 expression indicating the essential role of AKT3 in miR-122 mediated HCC transformation. In vivo, restoration of miR-122 completely inhibited xenograft growth of HCC tumor in mice. Our data strongly suggest that miR-122 is a tumor suppressor that targets AKT3 to regulate tumorigenesis in HCCs and a potential therapeutic candidate for liver cancer.     

胃癌中显著下调的miR-433的作用靶点研究

目的为了筛选胃癌中miRNAs的表达标记,验证胃癌相关miRNAs的作用靶点,建立一种新的诊断和治疗胃癌的方法。方法运用基因芯片技术检测3个正常胃组织标本,24个胃癌组织标本,胃癌细胞SGC7901和正常胃黏膜细胞GES-1中328个miRNAs的表达情况。用以上方法检测出在胃癌组织和SGC7901中,miR-433的表达水平显著下调。为了确保结果的准确性,采用实时荧光定量PCR对其进行验证。并用基因克隆和Western印迹方法分析miR-433的作用靶点。结果共有26个miRNAs在胃癌标本（包括24个胃癌组织和SGC7901）中异常表达。其中19个miRNAs下调,7个miRNAs上调。实时荧光定量PCR检测出miR-433在胃癌标本中的表达水平显著下调,该结果和基因芯片检测结果一致。另外,在本实验中发现miR-433与Grb2（growth factor receptor—bound protein 2）的表达呈负相关。结论胃癌相关miRNAs已进行了初步筛选。其中,miR-433可能是胃癌中的标记性miRNAs之一,Grb2是其作用靶点。这为建立新的以miRNAs为基础的诊断和治疗胃癌的方法提供了相关信息。     

miR-9: From function to therapeutic potential in cancer

Malignant neoplasms are regarded as the main cause of death around the world; hence, many research studies were conducted to further perceive molecular mechanisms, treatment, and cancer prognosis. Cancer is known as a major factor for health-related problems in the world. The main challenges associated with these diseases are prompt diagnosis, disease remission classification and treatment status forecast. Therefore, progressing in such areas by developing new and optimized methods with the help of minimally invasive biological markers such as circular microRNAs (miRNAs) can be considered important. miRNA interactions with target genes have specified their role in development, apoptosis, differentiation, and proliferation and also, confirm direct miRNA function in cancer. Different miRNAs expression levels in various types of malignant neoplasms have been observed to be associated with prognosis of various carcinomas. miR-9 seems to implement opposite practices in different tissues or under various cancer incidences by influencing different genes. Aberrant miR-9 levels have been observed in many cancer types. Therefore, we intended to investigate the precise role of miR-9 in patients with malignant neoplasms. To this end, in this study, we attempted to examine different studies to clarify the overall role of miR-9 as a prognostic marker in several human tumors. The presented data in this study can help us to find the novel therapeutic avenues for treatment of human cancers.     

Identification of miR-7 as an oncogene in renal cell carcinoma

MicroRNA-7 (miR-7) has been described as a tumor suppressor in several human cancers, but the results of a study to identify miRNAs associated with metastatic capability in breast cancer suggested that miR-7 may be characterized as an oncogene. The present study was to determine the expression and function of miR-7 in renal cell carcinoma. Quantitative real-time polymerase chain reaction was used to validate the expressions of miR-7 in 48 paired renal cell carcinomas (RCC) and normal tissues, based on the preliminary sequencing results of miRNAs. Furthermore, the impacts of miR-7 on cell migration, proliferation and apoptosis were analyzed using wound scratch assay, MTT and flow cytometry, respectively. The results demonstrated that miR-7 was up-regulated in RCC compared with normal tissues (p = 0.001). Down-regulation of miR-7 with synthesized inhibitor inhibited cell migration in vitro, suppressed cell proliferation and induced renal cancer cell apoptosis, prompting that miR-7 could be characterized as an oncogene in RCC. The present study was the first to reveal that miR-7 was up-regulated in RCC and it played an important role in RCC by affecting cellular migration, proliferation and apoptosis. Further researches should be conducted to explore the roles and target genes of miR-7 in RCC and other cancers.     

miR-21 downregulates the tumor suppressor P12 CDK2AP1 and stimulates cell proliferation and invasion

The present study was undertaken to investigate the regulation of P12(CDK2AP1) by miRNAs. A conserved target site for miR-21 within the CDK2AP1-3'-UTR at nt 349-370 was predicted by bioinformatics software and an inverse correlation of miR-21 and CDK2AP1 protein was observed. Highly specific amplification and quantification of miR-21 was achieved using real-time RT-PCR. Transfection of HaCaT cells with pre-miR-21 significantly suppressed a luciferase reporter including the CDK2AP1-3'-UTR, whereas transfection of Tca8113 with anti-miR-21 increased activity of this reporter. This was abolished when a construct mutated at the miR-21/nt 349-370 target site was used instead. Anti-miR-21-transfected Tca8113 cells showed an increase of CDK2AP1 protein and reduced proliferation and invasion. Resected primary tumors and tumor-free surgical margins of 18 patients with head and neck squamous cell carcinomas demonstrated an inverse correlation between miR-21 and P12(CDK2AP1). This study shows that P12(CDK2AP1) is downregulated by miR-21 and that miR-21 promotes proliferation and invasion in cultured cells.     

The clinical significance of miR-335, miR-124, miR-218 and miR-484 downregulation in gastric cancer

Gastric cancer (GC) is one of the leading types of malignancy worldwide, particularly in Asian populations. Although the exact molecular mechanism of GC development remains unknown, microRNA (miRNA) has recently been shown to be involved. The current study aims to investigate the expression levels of bioinformatically ranked miRNAs in gastric tissues. Using bioinformatics tools, we prioritized miRNAs thought to be implicated in GC. Furthermore, polyA-qPCR was used to validate bioinformatics findings in 40 GC, 31 normal gastric tissue (NG) and 45 gastric dysplasia (GD) samples. As identified by bioinformatics analysis, miR-335 was shown to be the top-ranked miRNA implicated in GC. Moreover, a significant downregulation of miR-335, miR-124, miR-218 and miR-484 was found in GC and GD compared to NG samples. We found bioinformatics to be an efficient approach to finding candidate miRNAs relevant to GC development. Finally, the findings show that downregulation of miRNAs such as miR-124 and miR-218 in gastric tissue can be a significant indicator for neoplastic transformation.     

Prognostic Value of miR-21 in Various Cancers: An Updating Meta-Analysis

Background

Recently, more and more studies investigated the value of microRNA (miRNA) as a diagnostic or prognostic biomarker in various cancers. MiR-21 was found dysregulated in almost all types of cancers. While the prognostic role of miR-21 in many cancers has been studied, the results were not consistent.

Methods

We performed a meta-analysis to investigate the correlation between miR-21 and survival of general cancers by calculating pooled hazard ratios (HR) and 95% confidence intervals (CI).

Results

The pooled results of 63 published studies showed that elevated miR-21 was a predictor for poor survival of general carcinomas, with pooled HR of 1.91 (95%CI: 1.66–2.19) for OS, 1.42 (95% CI: 1.16–1.74) for DFS and 2.2 (95% CI: 1.64–2.96) for RFS/CSS. MiR-21 was also a prognostic biomarker in the patients who received adjuvant therapy, with pooled HR of 2.4 (95%CI: 1.18–4.9) for OS.

Conclusions

Our results showed that miR-21 could act as a significant biomarker in the prognosis of various cancers. Further studies are warranted before the application of the useful biomarker in the clinical.     

Differentially expressed miR-20, miR-21, miR-100, miR-125a and miR-146a as a potential biomarker for prostate cancer

Prostate cancer is the leading cause of death among men worldwide. Deregulation of microRNAs has been reported in many cancers. Expression of microRNAs miR-20a-5p, miR-21-5p, miR-100-5p, miR-125a-5p and miR-146a-5p in tissue blocks of histologically confirmed prostate cancer patients compared with BPH patients, to identify potential microRNA biomarker for prostate cancer. MicroRNA was isolated and expression was quantified by qRT-PCR using Taqman Advanced microRNA assay kits. The interactions between the microRNA:target mRNA were predicted by using bioinformatics tools such as miRwalk and miRTargetlink. The experimentally validated targets were analysed using gprofiler to identify their molecular function, biological process and related pathways. The expression analysis revealed that miR-21 and miR-100 were significantly down-regulated whereas miR-125a was up-regulated in prostate cancer patients. Comparative analysis of the expression levels with tumor grading reveal that miR-100 was significantly down-regulated (p?<?0.05) in high grade tumor, indicating that miR-100 associated with prostate cancer. ROC analysis revealed that combined analysis of down-regulated miRNAs (miR-21 and miR-100) shown AUC of 0.72 (95% CI 0.65–0.79). The combined analysis of all five miRNAs showed AUC of 0.87 (95% CI 0.81–0.92). The targets prediction analysis revealed several validated targets including BCL2, ROCK1, EGFR, PTEN, MTOR, NAIF1 and VEGFA. Our results provide evidence that combined analysis of all the five miRNAs as a panel can significantly improve the prediction level of the presence of prostate cancer and may be used as a potential diagnostic biomarker.

     

Circulating microRNAs,miR-939, miR-595, miR-519d and miR-494, Identify Cirrhotic Patients with HCC

The performance of circulating biomarkers for the diagnosis of hepatocellular carcinoma (HCC) is sub-optimal. In this study we tested circulating microRNAs as biomarkers for HCC in cirrhotic patients by performing a two stage study: a discovery phase conducted by microarray and a validation phase performed by qRT-PCR in an independent series of 118 patients. Beside miRNAs emerged from the discovery phase, miR-21, miR-221, miR-519d were also tested in the validation setting on the basis of literary and tissue findings. Deregulated microRNAs were assayed in HCC-derived cells in the intracellular compartment, cell culture supernatant and exosomal fraction. Serum and tissue microRNA levels were compared in 14 patients surgically treated for HCC. From the discovery study, it emerged that seven circulating microRNAs were differentially expressed in cirrhotic patients with and without HCC. In the validation set, miR-939, miR-595 and miR-519d were shown to differentiate cirrhotic patients with and without HCC. MiR-939 and miR-595 are independent factors for HCC. ROC curves of miR-939, miR-595 and miR-519d displayed that AUC was higher than AFP. An exosomal secretion of miR-519d, miR-21, miR-221 and miR-1228 and a correlation between circulating and tissue levels of miR-519d, miR-494 and miR-21 were found in HCC patients. Therefore, we show that circulating microRNAs deserve attention as non-invasive biomarkers in the diagnostic setting of HCC and that exosomal secretion contributes to discharging a subset of microRNAs into the extracellular compartment.