miRNAs In Cancer Cells Research Articles

Quercetin shortened survival of radio-resistant B-1 cells in vitro and in vivo by restoring miR15a/16 expression.

Chronic lymphocytic leukemia (CLL) is a malignancy disease characterized by the expansion of CD5+ B-1 cells. The NZB mouse model of CLL shows similarities to human CLL, has age-associated increase in malignant B-1 clones and decreased expression of miR-15a/16. It was demonstrated that systemic lentiviral delivery of miR-15a/16 ameliorates disease manifestations in this mouse model. Nowadays, new therapeutic approaches have been focus on miRNA in cancer cells. Natural compounds like quercetin can modulate these miRNAs, consequently, suppress oncogenes or stimulate tumor suppressor genes by altering miRNA expressions. Here we investigate the effects of quercetin on miRNA15a/16 expression by radio-resistant B-1 cells. It has been described that a small percentage of B-1 cell survives to irradiation in vitro, and these cells show similarities to B-CLL cells. In these cells, the level of miR15a/16 is diminished and Bcl-2 is overexpressed. Quercetin treatment restore both, miR15a/16 and Bcl-2, to normal levels. Furthermore, transference of radioresistant B-1 cells to NOD/SCID mice causes an expansion of this population and also a migration to the liver. However, after quercetin treatment, even radioresistant B-1 cells are not able to expand or disseminate in vivo, and the levels of miR15a/16 and Bcl-2 are also normalized. Our data support the hypothesis that quercetin is an important adjuvant molecule that acts on miRNA15a/16 level and leads cells more permissive to apoptosis. This work could help to design new approaches to therapy in CLL patients.

SRSF1 regulates exosome microRNA enrichment in human cancer cells

BackgroundExosomes are extracellular vesicles containing a variety of biological molecules including microRNAs (miRNAs). We have recently demonstrated that certain miRNA species are selectively and highly enriched in pancreatic cancer exosomes with miR-1246 being the most abundant. Exosome miRNAs have been shown to mediate intercellular communication in the tumor microenvironment and promote cancer progression. Therefore, understanding how exosomes selectively enrich specific miRNAs to initiate exosome miRNA signaling in cancer cells is critical to advancing cancer exosome biology.ResultsThe aim of this study was to identify RNA binding proteins responsible for selective enrichment of exosome miRNAs in cancer cells. A biotin-labeled miR-1246 probe was used to capture RNA binding proteins (RBPs) from PANC-1 cells. Among the RBPs identified through proteomic analysis, SRSF1, EIF3B and TIA1 were highly associated with the miR-1246 probe. RNA immunoprecipitation (RIP) and electrophoretic mobility shift assay (EMSA) confirmed the binding of SRSF1 to miR-1246. Lentivirus shRNA knockdown of SRSF1 in pancreatic cancer cells selectively reduced exosome miRNA enrichment whereas GFP-SRSF1 overexpression enhanced the enrichment as analyzed by next generation small RNA sequencing and qRT-PCR. miRNA sequence motif analysis identified a common motif shared by 36/45 of SRSF1-associated exosome miRNAs. EMSA confirmed that shared motif decoys inhibit the binding of SRSF1 to the miR-1246 sequence.ConclusionsWe conclude that SRSF1 mediates selective exosome miRNA enrichment in pancreatic cancer cells by binding to a commonly shared miRNA sequence motif.4wJAidug7kxVDqDNvfU8znVideo Graphical abstract

Abstract 3700: Role of salivary miRNAs and their target genes in oral cancer patients as a potential diagnostic tool

Abstract Oral squamous cell carcinoma (OSCC) is a major cancer type worldwide, ranked in among top ten cancers. However, in South Asia, OSCC is at third position in commonly occurring cancer. Pakistan is ranked at tenth level in world but second in South Asia in OSCC. High ratio of cancer suggests that it is a major reason of death around the globe and is mainly because of the late-stage diagnosis. Metastasis of OSCC depends on many environmental and genetic factors including different genes, proteins and non-coding RNAs i.e. miRNAs. In cancer cells, miRNAs express themselves differentially. This differential expression of miRNAs in cancer cells can be explored and is worth considering as a diagnostic biomarker. On account of these facts, this study was designed to aim at detection of miRNAs and their target genes in OSCC through functional and expression analysis. We already have reported a list of miRNAs and their differentially expressed target genes in OSCC along with their functional and network analysis, in an in silico study. To evaluate the role of miRNAs and their target genes in OSCC patients of Pakistan, we measured the expression of four miRNAs; miR-300, miR-27a-3p, miR-340-5p, miR-205-3p with their corresponding target genes; ALB, CDH1, CACNA1, ARID5B, CDKN1A, BCL2, ABL1 in saliva samples of patients (n=100) and healthy controls (n=100). Total RNA was extracted from saliva samples; cDNA was synthesized and real time quantitative PCR was used to measure the expression. Preliminary results showed that miRNAs were down regulated in saliva samples of patients as compared to healthy controls. Consequently, target genes of each miRNA were dysregulated in patients. miR-300 targets CDH-1 and ALB and there was a direct relationship of miR-300 with ALB. Both of them were down regulated in OSCC patients. We found a down regulation in the expression of miRNA-27a-3p and its target genes CACNA1I and ARID5B in the saliva samples of patients when compared with the healthy controls. This suggests that there is a direct relationship between the miRNA-27a-3p and its target genes CACNA1I and ARID5B. We identified that miRNA-340-5p targets BCL2 and CDKN1A genes; miR-340-5p was down regulated by targeting both genes and also down regulated their expression, which results in OSCC. miRNA-205-3p targets ABL1 gene and there is direct relationship between the two. Both of them were down regulated in OSCC patients and change in expression leads to OSCC. The change in expression pattern of miRNAs and their target genes suggests a potential diagnostic role of these miRNAs in early detection of OSCC by saliva samples at early stages. Citation Format: Muhammad Jawad Khan, Andleeb Zahra, Ghulam Rabia, Syeda Aba Ali, Shanza Sahar, Sobia Bibi, M. Qaiser Fatmi. Role of salivary miRNAs and their target genes in oral cancer patients as a potential diagnostic tool [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3700.

MicroRNA-155-5p Diminishes in Vitro Ovarian Cancer Cell Viability by Targeting HIF1α Expression.

Purpose: Ovarian cancer is the most lethal of gynecological malignancies. Recently, the development of microRNA (miRNA) -based therapeutics that could impact broad cellular programs, leading to inhibition of cancer cell viability, is gaining attention in the therapeutic landscape. The therapy is based on the presence of aberrant expressions of miRNA in cancer cells. Decreasing of tumor suppressor miRNA expression causes upregulation of oncoprotein, which worsens the prognosis of the ovarian cancer. Methods: miR-155-5p mimics were carried by chitosan nanoparticles using new nanotechnology methods. Cellular uptake of miRNA was assessed by fluorescence microscope while MTT and qPCR assay were used to determine miRNA profile and the effect of CS-NP/miRNA on SKOV3 cells. Results: Results of profiling validated using quantitative realtime-polymerase chain reaction (PCR) found one of the most altered tumor suppressor miRNAs, miR-155-5p was downregulated 892.15-fold. According to bioinformatic analysis we identified the miRNA could recognize and regulate HIF1α expression. Transfection of mimics for miR-155-5p showed significantly increased miR-155-5p endogen SKOV3 expression level compared to the control group. We found differences after transfection mimics for miR-155-5p 31.5 and 63 nanoMolar. Increasing of miR-155-5p endogen lead to diminished SKOV3 viability (by 30%; <0.05 at concentration 80 nanoMolar). These mimics may cause an increase in upregulated miR-155-5p endogen that can reduce HIF1α expression. Here we found 2-fold and 2.8-fold reduction of HIF1α expression level after transfection compared to the control group. Conclusion: According to these findings, the mimics miR-155-5p can inhibit ovarian cancer cell proliferation by regulating HIF1α expression.

Exosome-mediated Let7c-5p Delivery for Breast Cancer Therapeutic Development

Breast cancer is the second leading cause of death among women worldwide. Although chemical or biological drugs have been developed, they have limited therapeutic effects. To overcome this issue, alternative therapeutic agents based on nucleic acid drugs have been developed. Of the several nucleic acid-based drugs, micro-RNAs (miRNAs) are known to be drastically downregulated in cancer cells, and this endows cancer cells with certain abilities such as hyper-proliferation. To regulate these cancer-related miRNAs, including let7c-5p in cancer cells, low levels of cancer-inhibiting miRNAs in cancer cells can be upregulated by supplementing miRNA mimics. However, because miRNAs are unstable, a specific delivery vehicle is required for their efficient and safe delivery. Exosomes are nano-sized extracellular vehicles derived from cells. Because exosomes contain various biomolecules and are easily uptaken by cells, they are considered to be less toxic and less suitable as miRNA delivery vehicles. Accordingly, HEK293T cell-derived exosomes have been developed as delivery vehicles of let7c-5p for breast cancer treatment. To achieve effective delivery, exosomes were loaded with let7c-5p and then successfully delivered to MDA-MB-231 breast cancer cells. It was found that cancer cell proliferation and migration were significantly suppressed when cells were treated with let7c-5p-loaded exosomes. Downstream targets of let7c-5p in breast cancer cells were analyzed to demonstrate that let7c-5p was delivered by exosomes, released, and hybridized with its target mRNAs to exert its anti-cancer activity. Overall, human exosome-based let7c-5p delivery is a potential anti-cancer drug and can contribute to the development of highly effective breast cancer therapy.

MicroRNAs targeting cellular cholesterol: implications for combating anticancer drug resistance.

Over sixty percent of all mammalian protein-coding genes are estimated to be regulated by microRNAs (miRNAs), and unsurprisingly miRNA dysregulation has been linked with cancer. Aberrant miRNA expression in cancer cells has been linked with tumourigenesis and drug resistance. In the past decade, increasing number of studies have demonstrated that cholesterol accumulation fuels tumour growth and contributes to drug resistance, therefore, miRNAs controlling cholesterol metabolism and homeostasis are obvious hypothetical targets for investigating their role in cholesterol-mediated drug resistance in cancer. In this review, we have collated published evidences to consolidate this hypothesis and have scrutinized it by utilizing computational tools to explore the role of miRNAs in cholesterol-mediated drug resistance in breast cancer cells. We found that hsa-miR-128 and hsa-miR-223 regulate genes mediating lipid signalling and cholesterol metabolism, cancer drug resistance and breast cancer genes. The analysis demonstrates that targeting these miRNAs in cancer cells presents an opportunity for developing new strategies to combat anticancer drug resistance.

A microfluidic paper-based laser-induced fluorescence sensor based on duplex-specific nuclease amplification for selective and sensitive detection of miRNAs in cancer cells

MicroRNAs (miRNAs) are considered as the potential biomarkers for many cancers. To determine miRNAs in cancer cells is significant for realizing these diseases. In this work, a microfluidic paper-based laser-induced fluorescence sensor based on duplex-specific nuclease (DSN) amplification was developed and applied to selectively and sensitively determine miRNAs in cancer cells. An interface for laser-induced fluorescence detection was firstly applied to perform the sample detection on the paper-based chip. Under the optimal conditions, DSN (3 μL 0.10 U) and Taqman probes (2 μL 2.5 × 10−7 M) were preserved on the circles (Diameter 4 mm) of the folded paper chip. When miRNA solution was added, the mixed solution could trigger fluorescence signal amplification by cyclically digesting hybrids of miRNAs and Taqman probes by DSN. The whole determination, including sample heating process, could be accomplished within 40 min. The detection limits for miRNA-21 and miRNA-31 were 0.20 and 0.50 fM respectively, corresponding to only 1.0 and 1.5 zmol consumption of miRNAs. The testing of mismatched miRNAs showed that the method had good specificity. Finally, the method was applied to determine miRNA-21 and miRNA-31 in lysates of cancer cells of A549 and HeLa, and hepatocyte LO2. MiRNA-21 and miRNA-31 could be successfully found from the two cancer cells. The concentrations for miRNA-21 and miRNA-31 were 1.74 × 10−13 M and 6.29 × 10−14 M in HeLa cell lysate (3.75 × 104 cells/mL), 3.07 × 10−15 M and 3.28 × 10−15 M in A549 cell lysate (8.33 × 106 cells/mL) respectively. The recoveries ranged from 87.30% to 111.83%, indicating the results were reliable. The developed method was effective, selective and sensitive in the determination of miRNAs in cancer cells.

Cancer Cell Membrane Vesicle for Multiplex MicroRNA Imaging in Living Cells.

Highly efficient cellular transfection and intracellular signal amplification is a prerequisite for low-abundant microRNA (miRNA) imaging and biomedical application. Herein, we report a functional cancer cell membrane (CM) vesicle, Au-P/DSN@CM (DSN, double-specific nucleases), which consists of Au nanoparticles modified with three types of fluorescent miRNA detection probes (Au-P) and DSN that simultaneously encapsulate in cancer CM. We find that the Au-P/DSN@CM could specifically target the cancer cell and transfect the cell with higher efficiency than Au nanoparticles. The internalized Au-P/DSN@CM could further specifically recognize the target miRNA and induce DSN-assisted target recycle signal amplification, leading to multiple miRNA simultaneous detection with high sensitivity. It successfully detects oncogenic miRNAs in MCF-7 cells with high sensitivity and is amenable to monitor the dynamic expression change of oncogenic miRNAs in cancer cells. Our study represents a promising gene delivery vector for cancer diagnosis and potential therapy.

MicroRNA-Based Combinatorial Cancer Therapy: Effects of MicroRNAs on the Efficacy of Anti-Cancer Therapies.

The susceptibility of cancer cells to different types of treatments can be restricted by intrinsic and acquired therapeutic resistance, leading to the failure of cancer regression and remission. To overcome this problem, a combination therapy has been proposed as a fundamental strategy to improve therapeutic responses; however, resistance is still unavoidable. MicroRNA (miRNAs) are associated with cancer therapeutic resistance. The modulation of dysregulated miRNA levels through miRNA-based therapy comprising a replacement or inhibition approach has been proposed to sensitize cancer cells to other anti-cancer therapies. The combination of miRNA-based therapy with other anti-cancer therapies (miRNA-based combinatorial cancer therapy) is attractive, due to the ability of miRNAs to target multiple genes associated with the signaling pathways controlling therapeutic resistance. In this article, we present an overview of recent findings on the role of therapeutic resistance-related miRNAs in different types of cancer. We review the feasibility of utilizing dysregulated miRNAs in cancer cells and extracellular vesicles as potential candidates for miRNA-based combinatorial cancer therapy. We also discuss innate properties of miRNAs that need to be considered for more effective combinatorial cancer therapy.

Abstract 5251: STAT1 and miR155 expression in human prostate cancer tissue

Abstract Background: Transcription factors (TFs) and microRNAs (miRNAs) form a gene regulatory network at transcriptional and posttranscriptional level. Increasing evidence shows that miRNAs regulates molecular pathways in cancer by targeting various oncogenes and tumor suppressor genes. However, the mutual regulations between TFs and miRNAs in cancer cells have not been well characterized. Signal transducer and activator of transcription 1 (STAT1) has been reported as a tumor suppressor, but recent evidence suggests that may also play a tumor-promoting role. An STAT1-miR155 feedback loop has been identified in normal tissues. We aimed to explore the STAT1 and miR155 expression in prostate cancer by a bioinformatics analysis combined with immunohistochemical evaluation of STAT1 in normal and prostate cancer tissues. Design: The expression of miR155 was explored in the dbDEMC2 database of Differentially Expressed miRNAs in Human Cancers. Potential targets of this miRNA were searched in the microRNA-target interactions database (miRTarBase), and STAT1 was selected. The protein expression level of STAT1 was analyzed in normal and tumor tissues, from the TCGA databases, by GEPIA bioinformatics tools; and by immunohistochemistry in a prostate tissue microarray (TMA), constructed from prostatectomy samples of patients with localized prostate cancer (PCa), including representative areas from PCa and benign prostate tissue (BPT). Immunohistochemical expression of STAT1 was evaluated by two pathologists, an H-score was obtained for each spot by microscopic assessment of the percentage of epithelial cells with positive staining. Differences in STAT1 expression were assessed by comparing H-Scores between BPT and PCa tissues using Mann-Whitney test, a p value less than 0.05 was considered statistically significant. The Ethical Review Board of the academic institution approved this study. Results: The analysis of miR155 and STAT1 expression in the Genome Cancer Atlas Database showed that both are overexpressed in a variety of tumors including prostate cancer. STAT1 expression at protein level was evaluated in 143 tissue cores representing PCa, and 78 tissue cores representing BPT. Mean STAT1 expression in cores with PCa, was significantly higher than mean of STAT1 expression in cores with BPT (p less than 0.0001). There was a higher mean staining score in PCa (210, 95% CI: 196,1-227,8), compared to BPT (105,5, 95% CI: 87,1-123,9). STAT1 staining was present predominantly in the cytoplasm, also in the membrane and nucleus, of prostate cancer tumor cells. Conclusions: Although miR155 and STAT1 are supposed to be involved in a negative auto-regulatory feedback mechanism, they were both found overexpressed in most of the prostate cancer tissues, which could be explained by the loss of the regulatory feedback loop equilibrium in the tumor environment. Additional studies to detect the in situ expression of miR155 are in progress to validate these results. Citation Format: Ines Benedetti, Lia Barrios Garcia, Juan Rebollo. STAT1 and miR155 expression in human prostate cancer tissue [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5251.

MicroRNA-Based Diagnosis and Treatment of Metastatic Human Osteosarcoma.

Osteosarcoma is a malignant tumor of the bones that commonly occurs in young individuals. The 5-year survival rate of osteosarcoma patients is 60–70%. Metastasis to the lungs leads to death in 30–40% of osteosarcoma patients. Therefore, the development of effective strategies for early detection and treatment of this disease are important to improve the survival of osteosarcoma patients. However, metastatic markers for osteosarcoma and molecules that might be targeted for the treatment of metastatic osteosarcoma have not been identified yet. Therefore, the mechanism of metastasis to the lungs needs to be explored from a novel viewpoint. Recently, the aberrant expression of microRNAs (miRNAs) has been reported to be involved in the carcinogenesis and cancer progression of many cancers. Furthermore, miRNAs in the blood have been reported to show an aberrant expression unique to several cancers. Therefore, miRNAs are gaining attention as potential diagnostic markers for cancers. On the other hand, normalizing the dysregulated expression of miRNAs in cancer cells has been shown to alter the phenotype of cancer cells, and thus treatment strategies targeting miRNAs are also being considered. This review summarizes the abnormality of miRNA expression associated with the metastasis of osteosarcoma and describes the present situation and issues regarding the early diagnosis and development of treatment strategies for metastatic osteosarcoma based on the current understanding of this disease.

Involvement of the Dysregulation of miR-23b-3p, miR-195-5p, miR-656-5p, and miR-340-5p in Trastuzumab Resistance of HER2-Positive Breast Cancer Cells and System Biology Approach to Predict Their Targets Involved in Resistance.

Resistance to trastuzumab has become a limiting factor for therapeutic efficacy of human epidermal growth factor 2 (HER2)-positive breast cancer. Different expression levels of miRNAs in cancer cells have been associated with poor prognosis and response to chemotherapy. The aim of this study was to evaluate miRNAs that were thought to be associated with HER2-positive breast cancer chemoresistance. In this study, the relative expression of candidate miRNAs to U6 RNA was evaluated in trastuzumab-resistant and trastuzumab-sensitive cells using relative real-time PCR. Our results demonstrated that miR-23b-3p, miR-195-5p, miR-656-5p, and miR-340-5p were significantly dysregulated. For the first time in this study, these miRNAs were identified to be involved in trastuzumab resistance. TargetScan and miRDB were then used for predicting the potential targets of the candidate miRNAs. Our results also revealed that the predicted potential targets of these miRNAs were strongly associated with drug resistance pathways. As a relative expression of candidate miRNAs was statistically different in trastuzumab-resistant and trastuzumab-sensitive cells, their potential targets were involved in drug resistance pathways. We strongly hypothesized the dysregulation of miRNAs as a possible mechanism of trastuzumab resistance. We also assumed that the strategic manipulation of these regulatory networks might be a possible therapeutic strategy to improve the results of chemotherapy for this resistance. However, more research is needed to evaluate the role of these miRNAs in the acquisition of trastuzumab resistance.

MiRNAs 484 and 210 regulate Pax-5 expression and function in breast cancer cells.

Recent studies have enabled the identification of important factors regulating cancer progression, such as paired box gene 5 (Pax-5). This transcription factor has consistently been associated to B-cell cancer lesions and more recently solid tumors including breast carcinoma. Although Pax-5 downstream activity is relatively well characterized, aberrant Pax-5 expression in a cancer-specific context is poorly understood. To investigate the regulation of Pax-5 expression, we turned to micro RNAs (miRNAs), small non-coding RNA molecules that regulate key biological processes. Extensive studies show that miRNA deregulation is prevalent in cancer lesions. In this study, we aim to elucidate a causal link between differentially expressed miRNAs in cancer cells and their putative targeting of Pax-5-dependent cancer processes. Bioinformatic prediction tools indicate that miRNAs 484 and 210 are aberrantly expressed in breast cancer and predicted to target Pax-5 messenger RNA (mRNA). Through conditional modulation of these miRNAs in breast cancer cells, we demonstrate that miRNAs 484 and 210 inhibit Pax-5 expression and regulate Pax-5-associated cancer processes. In validation, we show that these effects are probably caused by direct miRNA/mRNA interaction, which are reversible by Pax-5 recombinant expression. Interestingly, miRNAs 484 and 210, which are both overexpressed in clinical tumor samples, are also modulated during epithelial-mesenchymal transitioning and hypoxia that correlate inversely to Pax-5 expression. This is the first study demonstrating the regulation of Pax-5 expression and function by non-coding RNAs. These findings will help us better understand Pax-5 aberrant expression within cancer cells, creating the possibility for more efficient diagnosis and treatments for cancer patients.

Extracellular vesicles produced in B cells deliver tumor suppressor miR-335 to breast cancer cells disrupting oncogenic programming in vitro and in vivo

The successful implementation of miRNA (miR) therapies in humans will ultimately rely on the use of vehicles with improved cellular delivery capability. Here we tested a new system that leverages extracellular vesicles (EVs) laden with a tumor suppressor miRNA (miR-335) produced in B cells by plasmid DNA induction (iEVs). We demonstrate that iEVs-335 efficiently and durably restored the endogenous miR-335 pool in human triple negative breast cancer cells, downregulated the expression of the miR-335 target gene SOX4 transcription factor, and markedly inhibited tumor growth in vivo. Remarkably, iEVs-335 mediated transcriptional effects that persisted in tumors after 60 days post orthotopic implantation. Genome-wide RNASeq analysis of cancer cells treated in vitro with iEVs-335 showed the regulation of a discrete number of genes only, without broad transcriptome perturbations. This new technology may be ideally suited for therapies aimed to restore tumor suppressor miRNAs in cancer cells, disrupting the oncogenic program established after escape from miRNA control.

Abstract 1067: Tenascin C in colorectal cancer stroma is a predictive marker for liver metastasis and is a potent target of miR-198 as identified by microRNA analysis

Abstract Background: Liver metastasis in colorectal cancer (CRC) is still one of the major determinants of survival. Tumor stroma has important roles in the development of colorectal cancer (CRC) metastasis. MicroRNAs (miRNAs) are known to regulate the expression of genes involved in many physiological conditions, including cancer. Previous studies reported important roles of miRNAs in cancer cells in CRC development and metastasis. However, only few studies have focused on miRNA expression in CRC stroma. In this study, we revealed the miRNA expression profiles in tumor stroma by miRNA array analysis using Laser captured microdissection (LCM) to identify miRNAs and their target genes as novel molecular markers involved in CRC liver metastasis. Methods: Tumor stroma was isolated from formalin-fixed, paraffin-embedded tissues of primary CRCs with or without liver metastasis by LCM, and miRNA expression was analyzed using TaqMan miRNA arrays. Results: Hierarchical clustering classified 16 CRCs into two groups according to the existence of synchronous liver metastasis. Combinatory target prediction identified tenascin C (TNC) as a predicted target of miR-198, one of the top 10 miRNAs downregulated in tumor stroma of CRCs with synchronous liver metastasis. Immunohistochemical analysis of TNC in 139 primary CRCs revealed that a high staining intensity was correlated with synchronous liver metastasis (P&amp;lt;0.001). TNC staining intensities in the tumor stroma of 83 CRC liver metastases revealed a higher staining in liver metastases than in primary CRCs independently of preoperative chemotherapy. Univariate and multivariate analyses revealed that the TNC staining intensity was an independent prognostic factor to predict postoperative overall survival (P=0.005; n=139) and liver metastasis-free survival (P=0.001; n=128). To evaluate the biological link between miR-198 and TNC, we first checked expression levels of miR-198 in the 16 samples in miRNA array data, and compared them with TNC staining intensities. The CRC samples expressing miR-198 tended to have lower levels of TNC expression in comparison with those without miR-198 expression, although the difference was not statistically significant (P=0.237). Next, we transfected a miR-198 mimic or a miR-198 inhibitor into SW620 human colon cancer cells and CCD-18Co human colon fibroblast cells and analyzed the expression change of TNC protein by western blotting. The relative protein expression of TNC was suppressed by treatment with the miR-198 mimic and increased by miR-198 inhibitor. Conclusions: Alterations of miRNAs in CRC stroma appear to form a metastasis-permissive environment that can elevate TNC to promote liver metastasis. TNC in primary CRC stroma has the potential to be a novel biomarker to predict postoperative prognosis. Citation Format: Tomohiro Murakami, Hirotoshi Kikuchi, Hisahito Ishimatsu, Sanshiro Kawata, Amane Hirotsu, Tomohiro Matsumoto, Yusuke Ozaki, Yoshihiro Hiramatsu, Kinji Kamiya, Megumi Baba, Satoshi Baba, Masatoshi Kitagawa, Takanori Sakaguchi, Hiroyuki Konno, Hiroya Takeuchi. Tenascin C in colorectal cancer stroma is a predictive marker for liver metastasis and is a potent target of miR-198 as identified by microRNA analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1067.

Anti-tumor roles of both strands of the miR-455 duplex: their targets SKA1 and SKA3 are involved in the pathogenesis of renal cell carcinoma.

Recent studies revealed that some passenger strands of miRNAs acted as anti-tumor or oncogenic miRNAs in cancer cells. In this study, we focused on miR-455-5p (the passenger strand) and miR-455-3p (the guide strand) based on microRNA (miRNA) expression signatures of cancer cells. Both miR-455-5p and miR-455-3p were downregulated in renal cell carcinoma (RCC) tissues and low expression of these miRNAs was significantly associated with poor prognosis. Cancer cell proliferation, migration and invasive abilities were significantly inhibited by ectopic expression of miR-455-5p and miR-455-3p. To identify their oncogenic targets, we applied a combination of genome-wide gene expression and in silico miRNA database analyses. We focused on spindle and kinetochore-associated proteins, SKA1 and SKA3 and demonstrated direct regulation of SKA1 by miR-455-5p and SKA3 by miR-455-3p in RCC cells. Our present data demonstrated overexpression of SKA3 in RCC clinical specimens. Moreover, the study showed that the miR-455-3p/SKA3 axis contributed to cancer cell aggressiveness. Analytic strategies based on anti-tumor miRNAs, including passenger strands of miRNAs, are effective approaches for the elucidation of the molecular pathogenesis of RCC.

MicroRNAs, Regulatory Messengers Inside and Outside Cancer Cells.

MicroRNAs (miRNAs) are a class of short non-coding RNAs (ncRNAs) with typical sequence lengths of 19-25 nucleotides and extraordinary abilities to regulate gene expression. Because miRNAs regulate multiple important biological functions of the cell (proliferation, migration, invasion, apoptosis, differentiation, and drug resistance), their expression is highly controlled. Genetic and epigenetic alterations frequently found in cancer cells can cause aberrant expression of miRNAs and, consequently, of their target genes. The tumor microenvironment can also affect miRNA expression through soluble factors (e.g., cytokines and growth factors) secreted by either tumor cells or non-tumor cells (such as immune and stromal cells). Furthermore, like hormones, miRNAs can be secreted and regulate gene expression in recipient cells. Altered expression levels of miRNAs in cancer cells determine the acquisition of fundamental biological capabilities (hallmarks of cancer) responsible for the development and progression of the disease.

Involvement of aberrantly expressed microRNAs in the pathogenesis of head and neck squamous cell carcinoma.

MicroRNAs (miRNAs) are small noncoding RNAs that act as fine-tuners of the post-transcriptional control of protein-coding or noncoding RNAs by repressing translation or cleaving RNA transcripts in a sequence-dependent manner in cells. Accumulating evidence have been indicated that aberrantly expressed miRNAs are deeply involved in human pathogenesis, including cancers. Surprisingly, these small, single-stranded RNAs (18-23 nucleotides) have been shown to function as antitumor or oncogenic RNAs in several types of cancer cells. A single miRNA has regulating hundreds or thousands of different mRNAs, and individual mRNA has been regulated by multiple different miRNAs in normal cells. Therefore, tightly controlled RNA networks can be disrupted by dysregulated of miRNAs in cancer cells. Investigation of novel miRNA-mediated RNA networks in cancer cells could provide new insights in the field of cancer research. In this review, we focus on head and neck squamous cell carcinoma (HNSCC) and discuss current findings of the involvement of aberrantly expressed miRNAs in the pathogenesis of HNSCC.

Simultaneous Surface-Enhanced Raman Spectroscopy Detection of Multiplexed MicroRNA Biomarkers.

Simultaneous detection of cancer biomarkers holds great promise for the early diagnosis of cancer. In the present work, an ultrasensitive and reliable surface-enhanced Raman scattering (SERS) sensor has been developed for simultaneous detection of multiple liver cancer related microRNA (miRNA) biomarkers. We first proposed a novel strategy for the synthesis of nanogap-based SERS nanotags by modifying gold nanoparticles (AuNPs) with thiolated DNA and nonfluorescent small encoding molecules. We also explored a simple approach to a green synthesis of hollow silver microspheres (Ag-HMSs) with bacteria as templates. On the basis of the sandwich hybridization assay, probe DNA-conjugated SERS nanotags used as SERS nanoprobes and capture DNA-conjugated Ag-HMSs used as capture substrates were developed for the detection of target miRNA with a detection limit of 10 fM. Multiplexing capability for simultaneous detection of the three liver cancer related miRNAs with the high sensitivity and specificity was demonstrated using the proposed SERS sensor. Furthermore, the practicability of the SERS sensor was supported by the successful determination of target miRNA in cancer cells. The experimental results indicated that the proposed strategy holds significant potential for multiplex detection of cancer biomarkers and offers the opportunity for future applications in clinical diagnosis.

MTDH promotes glioma invasion through regulating miR-130b-ceRNAs.

Cell invasion is crucial for high mortality and recurrence rate in glioma. Epithelial-mesenchymal transition (EMT) is an important step in cancer invasion. Metadherin (MTDH) contributes to EMT in several cancers, but the role and mechanism of MTDH in EMT-like process of glioma remain unknown. Here we demonstrate that MTDH was overexpressed in glioma tissues and cells and induced EMT-like change and invasion of glioma cells. Interestingly, MTDH could modulate the expression of a group of glioma-related miRNAs. In particular, MTDH upregulated miR-130b transcription via acting as a coactivator of NF-kB. MiR-130b promoted EMT-like change and invasion of glioma cells through targeting multiple EMT-related genes, including PTEN, PPP2CA and SMAD7. In addition, PTEN acted as the competing endogenous RNA (ceRNA) to affect PPP2CA and SMAD7 expression, and inhibited EMT-like change in glioma cells. Furthermore, miR-130b mediated EMT-like change induced by MTDH, and MTDH inhibited the expression levels of PTEN, PPP2CA and SMAD7. Taken together, we reveal a novel mechanism that MTDH induces EMT-like change and invasion of glioma via the regulation of miR-130b-ceRNAs, providing the first direct link between MTDH and miRNAs in cancer cells.