MicroRNA Expression Profiling on Paired Primary and Lymph Node Metastatic Breast Cancer Revealed Distinct microRNA Profile Associated With LNM.
ABSTRACT: Breast cancer (BC) is the foremost cause of cancer-related deaths in women. BC patients are oftentimes presented with lymph node metastasis (LNM), which increases their risk of recurrence. Compelling data have recently implicated microRNAs in promoting BC metastasis. Therefore, the identification of microRNA (miRNA)-based molecular signature associated with LNM could provide an opportunity for a more personalized treatment for BC patients with high risk of LNM. In current study, we performed comprehensive miRNA profiling in matched primary breast and LNM and identified 40 miRNAs, which were differentially expressed in LNM compared to primary tumors. The expression of 14 miRNAs (Up: hsa-miR-155-5p, hsa-miR-150-5p, hsa-miR-146a-5p, hsa-miR-142-5p and down: hsa-miR-200a-3p, hsa-miR-200b-3p, hsa-miR-200c-3p, hsa-miR-205-5p, hsa-miR-210-3p, hsa-miR-214-3p, hsa-miR-141-3p, hsa-miR-127-3p, hsa-miR-125a-5p, and hsa-let-7c-5p) was subsequently validated in a second cohort of 32 breast and 32 matched LNM tumor tissues. Mechanistically, forced expression of hsa-miR-205-5p, or hsa-miR-214-3p epigenetically inhibited MDA-MB-231 cell proliferation, colony formation, and cell migration. Global gene expression profiling on MDA-MB-231 cells overexpressing hsa-miR-205-5p, or hsa-miR-214-3p in combination with in silico target prediction and ingenuity pathway analyses identified multiple bona fide targets for hsa-miR-205-5p, hsa-miR-214-3p affecting cellular proliferation and migration. Interestingly, interrogation of the expression levels of hsa-miR-205 and hsa-miR-214 in the METABRIC breast cancer dataset revealed significantly poor overall survival in patients with downregulated expression of miR-205 [HR = 0.75 (0.61-0.91)], p = 0.003 and hsa-miR-214 [HR = 0.74 (0.59-0.93) p = 0.008]. Our data unraveled the miRNA-transcriptional landscape associated with LNM and provide novel insight on the role of several miRNAs in promoting BC LNM, and suggest their potential utilization in the clinical management of BC patients.
Project description:Breast cancer is the second-most common cancer and second-leading cause of cancer mortality in American women. The dysregulation of microRNAs (miRNAs) plays a key role in almost all cancers, including breast cancer. We comprehensively analyzed miRNA expression, global gene expression, and patient survival from the Cancer Genomes Atlas (TCGA) to identify clinically relevant miRNAs and their potential gene targets in breast tumors. In our analysis, we found that increased expression of 12 mature miRNAs-hsa-miR-320a, hsa-miR-361-5p, hsa-miR-103a-3p, hsa-miR-21-5p, hsa-miR-374b-5p, hsa-miR-140-3p, hsa-miR-25-3p, hsa-miR-651-5p, hsa-miR-200c-3p, hsa-miR-30a-5p, hsa-miR-30c-5p, and hsa-let-7i-5p -each predicted improved breast cancer survival. Of the 12 miRNAs, miR-320a, miR-361-5p, miR-21-5p, miR-103a-3p were selected for further analysis. By correlating global gene expression with miRNA expression and then employing miRNA target prediction analysis, we suggest that the four miRNAs may exert protective phenotypes by targeting breast oncogenes that contribute to patient survival. We propose that miR-320a targets the survival-associated genes RAD51, RRP1B, and TDG; miR-361-5p targets ARCN1; and miR-21-5p targets MSH2, RMND5A, STAG2, and UBE2D3. The results of our stringent bioinformatics approach for identifying clinically relevant miRNAs and their targets indicate that miR-320a, miR-361-5p, and miR-21-5p may contribute to breast cancer survival.
Project description:Cytochrome P450 2E1 (CYP2E1) is an important drug metabolizing enzyme for processing numerous xenobiotics in the liver, including acetaminophen and ethanol. Previous studies have shown that microRNAs (miRNAs) can suppress CYP2E1 expression by binding to the 3'-untranslated region (3'-UTR) of its transcript. However, a systematic analysis of CYP2E1 regulation by miRNAs has not been described. Here, we applied in silico, in vivo, and in vitro approaches to investigate miRNAs involved in the regulation of CYP2E1. Initially, potential miRNA binding sites in the CYP2E1 mRNA transcript were identified and screened using in silico methods. Next, inverse correlations were found in human liver samples between the expression of CYP2E1 mRNA and the levels of two miRNA species, hsa-miR-214-3p and hsa-miR-942-5p. In a HepG2-derived CYP2E1 over-expression cell model, hsa-miR-214-3p exhibited strong suppression of CYP2E1 expression by targeting the coding region of its mRNA transcript, but hsa-miR-942-5p did not inhibit CYP2E1 levels. Electrophoretic mobility shift assays confirmed that hsa-miR-214-3p recruited other cellular protein factors to form stable complexes with specific sequences present in the CYP2E1 mRNA open reading frame. Transfection of HepaRG cells with hsa-miR-214-3p mimics inhibited expression of the endogenous CYP2E1 gene. Further, hsa-miR-214-3p mimics partially blocked ethanol-dependent increases in CYP2E1 mRNA and protein levels in HepG2 cells and they reduced the release of alanine aminotransferase from CYP2E1-overexpressing HepG2 cells exposed to acetaminophen. These results substantiate the suppressing effect of hsa-miR-214-3p on CYP2E1 expression.
Project description:Breast cancer (BC) is the most common cancer type and the second cause of cancer-related death among women. Therefore, better understanding of breast cancer tumor biology and the identification of novel biomarkers is essential for the early diagnosis and for better disease stratification and management choices. Herein we developed a novel approach which relies on the isolation of circulating microRNAs through an enrichment step using speed-vacuum concentration which resulted in 5-fold increase in microRNA abundance. Global miRNA microarray expression profiling performed on individual samples from 23 BC and 9 normals identified 18 up-regulated miRNAs in BC patients (p(corr)?<?0.05). Nine miRNAs (hsa-miR-4270, hsa-miR-1225-5p, hsa-miR-188-5p, hsa-miR-1202, hsa-miR-4281, hsa-miR-1207-5p, hsa-miR-642b-3p, hsa-miR-1290, and hsa-miR-3141) were subsequently validated using qRT-PCR in a cohort of 46 BC and 14 controls. The expression of those microRNAs was overall higher in patients with stage I, II, and III, compared to stage IV, with potential utilization for early detection. The expression of this microRNA panel was slightly higher in the HER2 and TN compared to patients with luminal subtype. Therefore, we developed a novel approach which led to the identification of a novel microRNA panel which was upregulated in BC patients with potential utilization in disease diagnosis and stratification.
Project description:MicroRNAs (miRNAs) regulate gene expression post-transcriptionally. In this way they might influence whether a cell is sensitive or resistant to a certain drug. So far, only a limited number of relatively small scale studies comprising few cell lines and/or drugs have been performed. To obtain a broader view on miRNAs and their association with drug response, we investigated the expression levels of 411 miRNAs in relation to drug sensitivity in 36 breast cancer cell lines. For this purpose IC50 values of a drug screen involving 34 drugs were associated with miRNA expression data of the same breast cancer cell lines. Since molecular subtype of the breast cancer cell lines is considered a confounding factor in drug association studies, multivariate analysis taking subtype into account was performed on significant miRNA-drug associations which retained 13 associations. These associations consisted of 11 different miRNAs and eight different drugs (among which Paclitaxel, Docetaxel and Veliparib). The taxanes, Paclitaxel and Docetaxel, were the only drugs having miRNAs in common: hsa-miR-187-5p and hsa-miR-106a-3p indicative of drug resistance while Paclitaxel sensitivity alone associated with hsa-miR-556-5p. Tivantinib was associated with hsa-let-7d-5p and hsa-miR-18a-5p for sensitivity and hsa-miR-637 for resistance. Drug sensitivity was associated with hsa-let-7a-5p for Bortezomib, hsa-miR-135a-3p for JNJ-707 and hsa-miR-185-3p for Panobinostat. Drug resistance was associated with hsa-miR-182-5p for Veliparib and hsa-miR-629-5p for Tipifarnib. Pathway analysis for significant miRNAs was performed to reveal biological roles, aiding to find a potential mechanistic link for the observed associations with drug response. By doing so hsa-miR-187-5p was linked to the cell cycle G2-M checkpoint in line with this checkpoint being the target of taxanes. In conclusion, our study shows that miRNAs could potentially serve as biomarkers for intrinsic drug resistance and that pathway analyses can provide additional information in this context.
Project description:PURPOSE:To identify potential molecular targets for lung cancer intervention and diagnosis, we analyzed the differential miRNA expression of peripheral blood between lung cancer patients and healthy controls. METHODS:Three pairs of cases' and controls' peripheral blood samples were evaluated for miRNA expression by microarray. 12 miRNAs were selected for RT-PCR validation and target genes prediction. In addition, 4 miRNAs were selected for future validation by RT-PCR in a large sample of 145 cases and 55 frequency-matched healthy controls. RESULTS:A total of 338 differentially expressed miRNAs were screened and identified by microarray. According to the fold changes, the top ten upregulated miRNAs were hsa-miR-124-3p, hsa-miR-379-5p, hsa-miR-3655, hsa-miR-450b-5p, hsa-miR-29a-5p, hsa-miR-200a-3p, hsa-miR-542-3p, hsa-miR-138-5p, hsa-miR-219a-2-3p, and hsa-miR-4701-3p, and the top ten downregulated miRNAs were hsa-miR-34c-5p, hsa-miR-135a-5p, hsa-miR-132-3p, hsa-miR-3178, hsa-miR-4449, hsa-miR-4999-3p, hsa-miR-1246, hsa-miR-4424, hsa-miR-1252-5p, and hsa-miR-24-2-5p. RT-PCR verification of the 12 miRNAs revealed that 5 of 8 upregulated miRNAs, 2 of 4 downregulated miRNAs showed a significant difference between the cases and controls (P < .05). A large number of target genes and their functional set showed overlapping among the 453 predicted target genes of the 12 miRNAs (P < .01). RT-PCR in the large sample confirmed the significant differential expression level of hsa-miR-29a-5p, hsa-miR-135a-5p, hsa-miR-542-3p, and hsa-miR-4491 between cases and controls (P < .05), and three of these microRNA, except hsa-miR-29a-5p, were significant after Bonferroni correction for adjustment of multiple comparisons. CONCLUSION:There was a significant difference in miRNAs expression in the peripheral blood between lung cancer patients and healthy controls, and 4 miRNAs were validated by a large-size sample.
Project description:BACKGROUND:MicroRNAs (miRNAs) play an important role in the development and progression of breast cancer (BC). The purpose of the present study was to identify plasma miRNAs enabling early diagnosis of BC. MATERIALS AND METHODS:Expression levels of seven plasma miRNAs (miR-23a-3p, miR-29b-2-5p, miR-130a-5p, miR-144-3p, miR-148a-3p, miR-152-3p, and miR-182-5p) in 106 patients with newly diagnosed BC and 96 healthy participants were analyzed by qRT-PCR. We also evaluated the relationship between the expression levels of these miRNAs and clinicopathological features of patients with BC. RESULTS:Compared with healthy controls, we found that miR-23a-3p (p = .025), miR-130a-5p (p = .006), miR-144-3p (p = .040), miR-148a-3p (p = .023), and miR-152-3p (p = .019) were downregulated in the plasma of patients with BC. MiR-130a-5p, miR-144-3p, and miR-152-3p were downexpressed in BC tissues as well as plasma. The expression of the miR-23a-3p, miR-144-3p, and miR-152-3p was related to ER positive and PR positive. Besides, miR-23a-3p, miR-144-3p, and miR-152-3p did show the significant difference in the staging compromised to the control, especially in stage I-II. Moreover, we also found that miR-144-3p and miR-148a-3p were associated with lymph node invasion. CONCLUSIONS:The expression levels of the miR-23a-3p, miR-130a-5p, miR-144-3p, miR-148a-3p, and miR-152-3p were lower in patients with BC compared to healthy controls and were associated with ex hormone receptor, clinical stage, and lymph node metastasis, indicating the diagnostic potential of these miRNAs in BC.
Project description:Novel noninvasive biomarkers with high sensitivity and specificity for the diagnosis of breast cancer (BC) are urgently needed in clinics. The aim of this study was to explore whether miRNAs from the miR-106a-363 cluster can be detected in the circulation of BC patients and whether these miRNAs can serve as potential diagnostic biomarkers.The expression of 12 miRNAs from the miR-106a-363 cluster was evaluated using qRT-PCR in 400 plasma samples (from 200 BC patients and 200 healthy controls (HCs)) and 406 serum samples (from 204 BC patients and 202 HCs) via a three-phase study. The identified miRNAs were further examined in tissues (32 paired breast tissues), plasma exosomes (from 32 BC patients and 32 HCs), and serum exosomes (from 32 BC patients and 32 HCs).Upregulated levels of four plasma miRNAs (miR-106a-3p, miR-106a-5p, miR-20b-5p, and miR-92a-2-5p) and four serum miRNAs (miR-106a-5p, miR-19b-3p, miR-20b-5p, and miR-92a-3p) were identified and validated in BC. A plasma 4-miRNA panel and a serum 4-miRNA panel were constructed to discriminate BC patients from HCs. The areas under the receiver-operating characteristic curves of the plasma panel were 0.880, 0.902, and 0.858, and those of the serum panel were 0.910, 0.974, and 0.949 for the training, testing, and external validation phases, respectively. Two overlapping miRNAs (miR-106a-5p and miR-20b-5p) were consistently upregulated in BC tissues. Except for the expression of the plasma-derived exosomal miR-20b-5p, the expression patterns of exosomal miRNAs were concordant between plasma and serum, indicating the potential use of exosomal miRNAs as biomarkers.We identified four plasma miRNAs and four serum miRNAs from the miR-106a-363 cluster as promising novel biomarkers for the diagnosis of BC.
Project description:Sperm contain microRNAs (miRNAs), which may have roles in epigenetic control. Regarding phylogenetic relationships among various swine breeds, Yorkshire and Landrace, are considered phenotypically and genetically very similar, but distinctly different from Duroc. The objective of the present study was to compare abundance of boar sperm miRNAs in these three breeds. Overall, 252 prioritized miRNAs were investigated using real-time PCR; relative expression of miRNAs in sperm was similar in Yorkshire and Landrace boars, but significantly different compared to Duroc. Seventeen miRNAs (hsa-miR-196a-5p, hsa-miR-514a-3p, hsa-miR-938, hsa-miR-372-3p, hsa-miR-558, hsa-miR-579-3p, hsa-miR-595, hsa-miR-648, hsa-miR-524-3p, hsa-miR-512-3p, hsa-miR-429, hsa-miR-639, hsa-miR-551a, hsa-miR-624-5p, hsa-miR-585-3p, hsa-miR-508-3p and hsa-miR-626) were down-regulated (P?<?0.05; fold regulation ?-2) in Yorkshire and Landrace sperm, compared to Duroc sperm. Furthermore, three miRNAs (hsa-miR-9-5p, hsa-miR-150-5p, and hsa-miR-99a-5p) were significantly up-regulated in Yorkshire and Landrace sperm compared to Duroc sperm, However, 240 miRNAs were not significantly different (within?+?2 fold) between Yorkshire and Landrace sperm. We concluded that miRNAs in sperm were not significantly different between Yorkshire and Landrace boars, but there were significant differences between those two breeds and Duroc boars. Furthermore, integrated target genes for selected down-regulated miRNAs (identified via an in-silico method) appeared to participate in spermatogenesis and sperm functions.
Project description:To improve the clinical decision-making regarding further treatment management and follow-up scheduling for patients with muscle-invasive bladder cancer (MIBC) after radical cystectomy (RC), a better prediction accuracy of prognosis for these patients is urgently needed. The objective of this study was to evaluate the validity of differentially expressed microRNAs (miRNAs) based on a previous study as prognostic markers for overall survival (OS) after RC in models combined with clinicopathological data. The expression of six miRNAs (miR-100-5p, miR-130b-3p, miR-141-3p, miR-199a-3p, miR-205-5p, and miR-214-3p) was measured by RT-qPCR in formalin-fixed, paraffin-embedded tissue samples from 156 MIBC patients who received RC in three urological centers. Samples from 2000 to 2013 were used according to their tissue availability, with follow-up until June 2016. The patient cohort was randomly divided into a training (n = 100) and test set (n = 56). Seventy-three samples from adjacent normal tissue were used as controls. Kaplan-Meier, univariate and multivariate Cox regression, and decision curve analyses were carried out to assess the association of clinicopathological variables and miRNAs to OS. Both increased (miR-130b-3p and miR-141-3p) and reduced (miR-100-5p, miR-199a-3p, and miR-214-3p) miRNA expressions were found in MIBC samples in comparison to nonmalignant tissue samples (P < 0.0001). miR-199a-3p and miR-214-3p were independent markers of OS in Cox regression models with the significant clinicopathological variables age, tumor status, and lymph node status. The prediction model with the clinicopathological variables was improved by these two miRNAs in both sets. The predictive benefit was confirmed by decision curve analysis. In conclusion, the inclusion of both miRNAs into models based on clinical data for the outcome prediction of MIBC patients after RC could be a valuable approach to improve prognostic accuracy.
Project description:The new epidemic Middle East Respiratory Syndrome (MERS) is caused by a type of human coronavirus called MERS-CoV which has global fatality rate of about 30%. We are investigating potential antiviral therapeutics against MERS-CoV by using host microRNAs (miRNAs) which may downregulate viral gene expression to quell viral replication. We computationally predicted potential 13 cellular miRNAs from 11 potential hairpin sequences of MERS-CoV genome. Our study provided an interesting hypothesis that those miRNAs, that is, hsa-miR-628-5p, hsa-miR-6804-3p, hsa-miR-4289, hsa-miR-208a-3p, hsa-miR-510-3p, hsa-miR-18a-3p, hsa-miR-329-3p, hsa-miR-548ax, hsa-miR-3934-5p, hsa-miR-4474-5p, hsa-miR-7974, hsa-miR-6865-5p, and hsa-miR-342-3p, would be antiviral therapeutics against MERS-CoV infection.