Recent advantages of serum microRNAs (miRNAs) open a new realm of possibilities for noninvasive diagnosis and prognosis of bladder cancer (BC). The aim of our study was to identify serum miRNA expression signatures in patients with BC and establish new models for the diagnosis of BC and recurrence prediction. We performed genome-wide serum miRNA analysis by Miseq sequencing followed by evaluations in the training and validation sets with reverse transcription quantitative real-time PCR assays from serum samples of 250 patients with BC and 240 controls. A six-miRNA panel (miR-152, miR-148b-3p, miR-3187-3p, miR-15b-5p, miR-27a-3p and miR-30a-5p) for the diagnosis of BC was finally developed by multivariate logistic regression model with an area under the receiver operating characteristic curve of 0.899. The corresponding sensitivities of this panel for Ta, T1 and T2-T4 were 90.00, 84.85 and 89.36%, significantly higher than those of urine cytology, which were 13.33, 30.30 and 44.68%, respectively (all at p < 0.001). In addition, Kaplan-Meier analysis showed that patients with nonmuscle-invasive BC (NMIBC) with high miR-152 level and low miR-3187-3p level had worse recurrence-free survival (p 5 0.023 and 0.043, respectively). In multivariate Cox regression analysis, miR-152 was independently associated with tumor recurrence of NMIBC (p 5 0.028). Our results suggested that a serum miRNA signature may have considerable clinical value in diagnosing BC. Furthermore, expression level of serum miR-152 could provide information on the recurrence risk of NMIBC.Bladder cancer (BC) is one of the leading causes of cancerrelated death worldwide, with an estimated 72,570 new cases and 15,210 deaths in 2013 in the United States. 1 About 70% of patients are initially diagnosed with nonmuscle-invasive BC (NMIBC), but as many as 50-70% of these tumors will recur and roughly 10-20% will progress to muscle-invasive BC (MIBC). 2 Cancer screening and early diagnosis have major importance in improving survival of patients with BC. Currently, urine cytology is most commonly used as the noninvasive test for the detection of BC. However, this test is of limited value owing to its poor sensitivity, especially for low-grade lesions. [3][4][5][6][7] Cystoscopy-guided biopsy for histological evaluation can offer high diagnostic accuracy, but it is invasive and inconvenient, which limits its use for general cancer screening. Therefore, noninvasive and more sensitive molecular biomarkers remain needed to complete and improve on current strategies for the detection of BC.MicroRNAs (miRNAs) are an abundant class of noncoding small RNAs ( 22 necleotides in length) that regulate gene expression by binding to the 3 0 untranslated region of target mRNAs. 8,9 miRNAs participate in almost all of the known hallmarks of tumorigenesis, including cell proliferation, differentiation, apoptosis, invasion and metastasis. [10][11][12][13][14][15] The previous studies have shown that miRNAs exist stably in human serum and have potential role in the diagnosis a...

MicroRNA-210 (miR-210), the master hypoxamir, plays pleiotropic roles in certain cancers; however, its role in the development of human colorectal cancer remains unclear. Herein, we report that miR-210 is frequently up-regulated in colorectal cancer tissues, with high miR-210 expression significantly correlating with large tumor size, lymph node metastasis, advanced clinical stage and poor prognosis. Functionally, miR-210 overexpression promotes the migration and invasion of colorectal cancer cells. Furthermore, miR-210 can be induced by hypoxia and mediates the hypoxia-induced metastasis of colorectal cancer cells. In addition, vacuole membrane protein 1 (VMP1) is identified as the direct and functional target of miR-210. Thus, miR-210 is a useful biomarker for hypoxic tumor cells and a prognostic factor that plays an essential role in colorectal cancer metastasis.

Serum microRNAs (miRNAs) have become a highlighted research hotspot, especially for their great potential as a novel promising non-invasive biomarker in cancer diagnosis. The most frequently used approach for serum miRNAs detection is quantitative real time polymerase chain reaction (qPCR). In order to obtain reliable qPCR data of miRNAs expression, the use of reference genes as endogenous control is undoubtly necessary. However, no systematic evaluation and validation of reference genes for normalizing qPCR analysis of serum miRNAs has been reported in colorectal adenocarcinoma. We firstly profiled pooled serum of colorectal adenocarcinoma, colorectal adenoma and healthy controls and selected a list of 13 miRNAs as candidate reference genes. U6 snRNA (U6) and above-mentioned 13 miRNAs were included in further confirmation by qPCR. As a result, 5 miRNAs (miR-151a-3p, miR-4446-3p, miR-221-3p, miR-93-5p and miR-3184-3p) were not detected in all samples and 2 miRNAs (miR-197-3p and miR-26a-5p) were relatively low with median Cq more than 35, and were excluded from further stability analysis. Then variable stability of other 6 miRNAs (miR-103b, miR-484, miR-16-5p, miR-3615, miR-18a-3p and miR-191-5p) and U6 were evaluated using two algorithms: geNorm and NormFinder which both identified miR-191-5p as the most stably expressed reference gene and selected miR-191-5p and U6 as the most stable pair of reference genes. After validating in an independent large cohorts and selecting miR-92a-3p as target miRNA to evaluate the effect of reference gene, we propose that combination of miR-191-5p and U6 could be used as reference genes for serum microRNAs qPCR data in colorectal adenocarcinoma, colorectal adenoma and healthy controls.

Deregulation of microRNAs is a frequent event in the tumorigenesis and tumor progression. The aim of this study was to investigate the clinical significance and potential role of miR-24-3p expression in colorectal cancer (CRC). The expression level of miR-24-3p was determined in 95 CRC patients who underwent radical resection by quantitative real-time PCR. The associations between miR-24-3p expression and clinicopathological parameters were analyzed. In vitro function assays including cell proliferation, cell migration and invasion were further explored. We found that miR-24-3p was reduced in CRC tissues compared with their corresponding non-cancerous tissues (P < 0.001) and significantly correlated with local invasion (P = 0.002), lymph node metastasis (P = 0.0007) and clinical stage (P < 0.001). Moreover, Kaplan-Meier survival analysis showed that patients with low miR-24-3p level had a significantly poorer prognosis than those with high miR-24-3p level (P < 0.001). Multivariate analysis revealed that miR-24-3p (HR 2.767; 95 % CI 1.203-6.364; P = 0.017) and clinical TNM stage (HR 0.456; 95 % CI 0.212-0.978; P = 0.044) could be independent prognostic indicators for overall survival rates of CRC patients. In addition, functional assays showed that over-expression of miR-24-3p suppressed CRC cell proliferation, cell migration and invasion. miR-24-3p functions as a tumor suppressor in CRC. Down-regulation of miR-24-3p contributes to the development and progression of CRC and may have a potential role in prognosis and therapy.

Zinc finger protein 217 (ZNF217) is essential for cell proliferation and has been implicated in tumorigenesis. However, its expression and exact roles in colorectal cancer (CRC) remain unclear. In this study, we demonstrated that ZNF217 expression was aberrantly upregulated in CRC tissues and associated with poor overall survival of CRC patients. In addition, we found that ZNF217 was a putative target of microRNA (miR)-203 using bioinformatics analysis and confirmed that using luciferase reporter assay. Moreover, in vitro knockdown of ZNF217 or enforced expression of miR-203 attenuated CRC cell proliferation, invasion and migration. Furthermore, combined treatment of ZNF217 siRNA and miR-203 exhibited synergistic inhibitory effects. Taken together, our results provide new evidences that ZNF217 has an oncogenic role in CRC and is regulated by miR-203, and open up the possibility of ZNF217- and miR-203-targeted therapy for CRC.

Purpose: Piwi-interacting RNAs (piRNAs) are a novel class of small non-coding RNAs, which are not easily degraded but detectable in human body fluids. Recent studies have shown that aberrant piRNA expression is a signature feature across multiple tumor types. However, the expressions of piRNAs in serum of tumor patients and their potential clinical values remain largely unclear. Patients and methods: High-throughput sequencing was performed to investigate the serum piRNA profiles, followed by evaluations in serum samples of 220 colorectal cancer (CRC) patients and 220 healthy controls using reverse transcription quantitative real-time PCR (RT-qPCR). Biomarker panels including piRNA-based Panel I and carcinoembryonic antigen (CEA)-based Panel II, were developed by logistic regression model, and their diagnostic potentials were compared. Fagan’s nomogram was plotted to promote clinical application. Results: We identified five differentially expressed serum piRNAs (piR-001311, piR-004153, piR-017723, piR-017724 and piR-020365), which, when combined in the piRNA-based Panel I, outperformed the CEA-based Panel II ( P <0.001) and could detect CRC with an area under the receiver operating characteristic curve of 0.867. In addition, Kaplan–Meier analysis showed that patients with low serum piR-017724 level had worse overall survival (OS) and progression-free survival (PFS). In multivariate Cox regression analysis, serum piR-017724 was an independent prognostic factor for OS and PFS ( P <0.05). Conclusion: Our findings suggest serum piRNA expression signatures have potential for use as biomarkers for CRC detection and to predict prognosis at the time of diagnosis.

microRNA-210 (miR-210), the master hypoxamir, is overexpressed and generally exhibits oncogenic properties in most human solid tumours, including colorectal cancer (CRC). However, the status of circulating miR-210 in CRC is still unknown. This study aims to assess the clinical significance of circulating miR-210 in CRC. Using (reverse transcription quantitative PCR) RT-qPCR analysis, we compared the expression levels of circulating miR-210 in serum of 268 CRC patients and 102 healthy controls, and found that serum miR-210 was significantly higher in CRC than in healthy controls (P < 0.001). The area under the receiver operating characteristic curve (AUC) of circulating miR-210 to detect CRC was 0.821, with a sensitivity of 74.6% and a specificity of 73.5%. The AUC of circulating miR-210 showed significantly higher detection capability than that of carcinoembryogenic antigen (P < 0.05). Kaplan-Meier analysis demonstrated that increased serum miR-210 level correlated with reduced overall survival (OS) and disease-free survival (DFS) (P = 0.008 and P = 0.008 respectively). Cox analysis indicated circulating miR-210 was an independent prognostic factor for OS and DFS. Taken together, our data suggested that circulating miR-210 could be a potential non-invasive marker for diagnosis and prognosis of CRC.

Serum microRNAs (miRNAs) have been proposed as novel non-invasive biomarkers for the early detection of cancer. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is the most commonly used method for investigating miRNA expression levels, however, the interpretation of RT-qPCR results depends largely on normalization to an appropriate endogenous control. The present study involved 129 patients with non-muscle-invasive bladder cancer (NMIBC), 121 patients with muscle-invasive bladder cancer (MIBC) and 158 healthy controls. The aim of the present study was to determine the most stable reference genes for the investigations of serum miRNA in bladder cancer (BC). MiSeq sequencing was performed and the expression levels of 10 miRNAs and U6 were then measured using RT-qPCR. Following RT‑qPCR, five genes (hsa-miR-193a-5p, hsa-miR-16-5p, U6, hsa-miR-191-5p and hsa-let-7d-3p) were selected for stability analysis using geNorm and NormFinder software. These algorithms identified hsa-miR-193a-5p and hsa-miR-16-5p as the most stably expressed reference genes. The availability of hsa-miR-193a-5p and hsa-miR-16-5p was confirmed in an additional cohort. One-way analysis of variance indicated that no significant differences were present in the expression levels among the three groups. Furthermore, miR-148b-3p was selected as a target miRNA to determine the effect of hsa-miR-193a-5p and hsa-miR-16-5p on miRNA quantification. The combined use of hsa-miR-193a-5p and hsa-miR-16-5p enabled the detection of a significant upregulation of miR-148b-3p in the BC serum. The results of the present study demonstrated that normalization of miRNA data, using a combination of hsa-miR-193a-5p and hsa-miR-16-5p as reference genes, may produce reliable and accurate results for the detection of serum miRNAs in BC.