MicroRNA profiling and prediction of recurrence/relapse-free survival in stage I lung cancer

About 30% stage I non-small cell lung cancer (NSCLC) patients undergoing resection will recur. Robust prognostic markers are required to better manage therapy options. MicroRNAs (miRNAs) are a class of small non-coding RNAs of 19-25 nt and play important roles in gene regulation in human cancers. The purpose of this study is to identify miRNA expression profiles that would better predict prognosis of stage I NSCLC. MiRNAs extracted from 527 stage I NSCLC patients were profiled on the human miRNA expression profiling v2 panel (Illumina). The expression profiles were analyzed for their association with cancer subtypes, lung cancer brain metastasis and recurrence/relapse free survival (RFS). MiRNA expression patterns between lung adenocarcinoma and squamous cell carcinoma differed significantly with 171 miRNAs, including Let-7 family members and miR-205. Ten miRNAs associated with brain metastasis were identified including miR-145*, which inhibit cell invasion and metastasis. Two miRNA signatures that are highly predictive of RFS were identified. The first contained 34 miRNAs derived from 357 stage I NSCLC patients independent of cancer subtype, whereas the second containing 27 miRNAs was adenocarcinoma specific. Both signatures were validated using formalin-fixed paraffin embedded and/or fresh frozen tissues in independent data set with 170 stage I patients. Our findings have important prognostic or therapeutic implications for the management of stage I lung cancer patients. The identified miRNAs hold great potential as targets for histology-specific treatment or prevention and treatment of recurrent disease.     

MicroRNA expression profiling in human ovarian cancer: miR-214 induces cell survival and cisplatin resistance by targeting PTEN

MicroRNAs (miRNA) represent a novel class of genes that function as negative regulators of gene expression. Recently, miRNAs have been implicated in several cancers. However, aberrant miRNA expression and its clinicopathologic significance in human ovarian cancer have not been well documented. Here, we show that several miRNAs are altered in human ovarian cancer, with the most significantly deregulated miRNAs being miR-214, miR-199a*, miR-200a, miR-100, miR-125b, and let-7 cluster. Further, we show the frequent deregulation of miR-214, miR-199a*, miR-200a, and miR-100 in ovarian cancers. Significantly, miR-214 induces cell survival and cisplatin resistance through targeting the 3'-untranslated region (UTR) of the PTEN, which leads to down-regulation of PTEN protein and activation of Akt pathway. Inhibition of Akt using Akt inhibitor, API-2/triciribine, or introduction of PTEN cDNA lacking 3'-UTR largely abrogates miR-214-induced cell survival. These findings indicate that deregulation of miRNAs is a recurrent event in human ovarian cancer and that miR-214 induces cell survival and cisplatin resistance primarily through targeting the PTEN/Akt pathway.     

宫颈癌及癌前病变组织中microRNAs的差异表达

目的 寻找与宫颈癌及宫颈癌前病变相关的microRNA。 方法 利用miRNA芯片,筛查宫颈 癌组织、宫颈上皮内瘤变及正常宫颈组织中差异表达的miRNA,并用实时定量RT-PCR在60份宫颈组 织标本中对4个miRNA进行验证。利用生物信息学对部分差异表达的miRNA的靶基因进行功能分析。 结果 与正常宫颈组织比较,宫颈癌及高级别宫颈病变（HSIL）中存差异表达的miRNAs,其中在宫 颈癌中下调最明显的是miR-218（下调倍数为0.175）,上调最明显的是miR-21（上调倍数为5.68）。 实时定量RT-PCR验证结果与miRNA芯片结果基本一致。功能分析显示预测的miR-218及miR-21的靶基 因与肿瘤的生长、侵袭转移有关。结论 宫颈癌及癌前病变中存在异常表达的miRNA,它们在宫颈癌 发生过程中可能起癌基因或抑癌基因的作用。     

Down-regulation of the microRNAs miR-34a, miR-127, and miR-200b in rat liver during hepatocarcinogenesis induced by a methyl-deficient diet

Altered expression of microRNAs (miRNAs) has been reported in diverse human cancers; however, the down-regulation or up-regulation of any particular miRNAs in cancer is not sufficient to address the role of these changes in carcinogenesis. In this study, using the rat model of liver carcinogenesis induced by a methyl-deficient diet, which is relevant to the hepatocarcinogenesis in humans associated with viral hepatitis C and B infections, alcohol exposure and metabolic liver diseases, we showed that the development of hepatocellular carcinoma (HCC) is characterized by prominent early changes in expression of miRNA genes, specifically by inhibition of expression of microRNAs miR-34a, miR-127, miR-200b, and miR-16a involved in the regulation of apoptosis, cell proliferation, cell-to-cell connection, and epithelial-mesenchymal transition. The mechanistic link between these alterations in miRNAs expression and the development of HCC was confirmed by the corresponding changes in the levels of E2F3, NOTCH1, BCL6, ZFHX1B, and BCL2 proteins targeted by these miRNAs. The significance of miRNAs expression dysregulation in respect to hepatocarcinogenesis was confirmed by the persistence of these miRNAs alterations in the livers of methyl-deficient rats re-fed a methyl-adequate diet. Altogether, the early occurrence of alterations in miRNAs expression and their persistence during the entire process of hepatocarcinogenesis indicate that the dysregulation of microRNAs expression may be an important contributing factor in the development of HCC.     

The miRNA-200 family and miRNA-9 exhibit differential expression in primary versus corresponding metastatic tissue in breast cancer

Metastases are the major cause of cancer-related deaths, but the mechanisms of the metastatic process remain poorly understood. In recent years, the involvement of microRNAs (miRNAs) in cancer has become apparent, and the objective of this study was to identify miRNAs associated with breast cancer progression. Global miRNA expression profiling was performed on 47 tumor samples from 14 patients with paired samples from primary breast tumors and corresponding lymph node and distant metastases using LNA-enhanced miRNA microarrays. The identified miRNA expression alterations were validated by real-time PCR, and tissue distribution of the miRNAs was visualized by in situ hybridization. The patients, in which the miRNA profile of the primary tumor and corresponding distant metastasis clustered in the unsupervised cluster analysis, showed significantly shorter intervals between the diagnosis of the primary tumor and distant metastasis (median 1.6 years) compared to those that did not cluster (median 11.3 years) (p<0.003). Fifteen miRNAs were identified that were significantly differentially expressed between primary tumors and corresponding distant metastases, including miR-9, miR-219-5p and four of the five members of the miR-200 family involved in epithelial-mesenchymal transition. Tumor expression of miR-9 and miR-200b were confirmed using in situ hybridization, which also verified higher expression of these miRNAs in the distant metastases versus corresponding primary tumors. Our results demonstrate alterations in miRNA expression at different stages of disease progression in breast cancer, and suggest a direct involvement of the miR-200 family and miR-9 in the metastatic process.     

MTA1 gene silencing inhibits invasion and alters the microRNA expression profile of human lung cancer cells

Metastasis-associated gene 1 (MTA1) is involved in the carcinogenesis and metastasis of many human carcinomas. However, its exact role in non-small cell lung cancer (NSCLC) is still unclear. Using immunohistochemistry analysis, we recently identified MTA1 to be associated with the progression of NSCLC. Here, we carried out further analysis on the effect of MTA1 knockdown in an NSCLC cell line on cell functions and the global microRNA (miRNA) expression profile. We succeeded in establishing the MTA1 knockdown NSCLC cell line using RNA interference (RNAi), and found that the silencing of MTA1 resulted in the effective inhibition of the invasive ability of NSCLC cells, but not of the cell growth in vitro. We performed an miRNA microarray analysis and demonstrated for the first time that MTA1 knockdown significantly changed the expression of some miRNAs in NSCLC cells. Among them, some have a well-characterized association with cancer progression, e.g. miR-125b, miR-210, miR-103, miR-194 and miR-500. In summary, it is evident from our results that MTA1 functions in regulating the invasive phenotype of lung cancer cells and this regulation may be through altered miRNA expression. The interaction between MTA1 and the miRNAs which contributes to lung cancer is worthy of further investigation.     

microRNA-320a inhibits tumor invasion by targeting neuropilin 1 and is associated with liver metastasis in colorectal cancer

MicroRNAs (miRNAs) have been implicated in regulating diverse cellular pathways. Although there is emerging evidence that various miRNAs function as oncogenes or tumor suppressors in colorectal cancer (CRC), the role of miRNAs in mediating liver metastasis remains unexplored. The expression profile of miRNAs in liver metastasis and primary CRC tissues was analyzed by miRNA microarrays and verified by real-time polymerase chain reaction (PCR). In 62 CRC patients, the expression levels of miR-320a were determined by real-time PCR, and the effects on migration and invasion of miR-320a were determined using a transwell assay. miR-320a target genes were confirmed by luciferase assay, real-time PCR and western blot analysis. A set of miRNAs was found to be dysregulated in the liver metastasis tissues compared to matched primary CRC tissues, and the expression levels of miR-320a were significantly decreased in the liver metastasis tissues examined. miR-320a was correlated with tumor progression in CRC. miR-320a was downregulated in liver metastatic colon cancer cells and inhibited liver metastatic colon cancer cell migration and invasion. miR-320a directly binds to the 3'UTR of neuropilin 1 (NRP-1), a protein that functions as a co-receptor of vascular epithelial growth factor. miR-320a downregulated the expression of NRP-1 at both the mRNA and protein levels. These data demonstrated that miR-320a may be useful for identifying CRC patients that are at an elevated risk for developing liver metastasis. Our findings suggest that miR-320a may be a novel therapeutic candidate for the treatment of colorectal cancer.     

MicroRNA signatures in human ovarian cancer

Epithelial ovarian cancer (EOC) is the sixth most common cancer in women worldwide and, despite advances in detection and therapies, it still represents the most lethal gynecologic malignancy in the industrialized countries. Unfortunately, still relatively little is known about the molecular events that lead to the development of this highly aggressive disease. The relatively recent discovery of microRNAs (miRNA), a class of small noncoding RNAs targeting multiple mRNAs and triggering translation repression and/or RNA degradation, has revealed the existence of a new level of gene expression regulation. Multiple studies involving various types of human cancers proved that miRNAs have a causal role in tumorigenesis. Here we show that, in comparison to normal ovary, miRNAs are aberrantly expressed in human ovarian cancer. The overall miRNA expression could clearly separate normal versus cancer tissues. The most significantly overexpressed miRNAs were miR-200a, miR-141, miR-200c, and miR-200b, whereas miR-199a, miR-140, miR-145, and miR-125b1 were among the most down-modulated miRNAs. We could also identify miRNAs whose expression was correlated with specific ovarian cancer biopathologic features, such as histotype, lymphovascular and organ invasion, and involvement of ovarian surface. Moreover, the levels of miR-21, miR-203, and miR-205, up-modulated in ovarian carcinomas compared with normal tissues, were significantly increased after 5-aza-2'-deoxycytidine demethylating treatment of OVCAR3 cells, suggesting that the DNA hypomethylation could be the mechanism responsible for their overexpression. Our results indicate that miRNAs might play a role in the pathogenesis of human EOC and identify altered miRNA gene methylation as a possible epigenetic mechanism involved in their aberrant expression.     

Genes associated with liver metastasis of colon cancer, identified by genome-wide cDNA microarray

To uncover mechanisms underlying progression of colorectal carcinogenesis and to identify genes associated with liver metastasis, we analyzed expression profiles of 14 primary colorectal cancers (CRCs) with liver metastases, and compared them with profiles of 11 non-metastatic carcinomas and those of 9 adenomas of the colon. A hierarchical cluster analysis using data from a cDNA microarray containing 23,040 genes indicated that the cancers with metastasis had different expression profiles from those without metastasis, although a number of genes were commonly up-regulated in primary cancers of both categories. We documented 54 genes that were frequently up-regulated and 375 that were frequently down-regulated in primary tumors with metastases to liver, but not in tumors without metastasis. Subsequent quantitative PCR experiments confirmed that PRDX4, CKS2, MAGED2, and an EST (GenBank accession number BF696304) were expressed at significantly higher levels in tumors with metastasis. These data should contribute to a better understanding of the progression of colorectal tumors, and facilitate prediction of their metastatic potential.     

MicroRNA expression profile of MCF-7 human breast cancer cells and the effect of green tea polyphenon-60

This study reports for the first time the microRNA expression profile of human breast cancer MCF-7 cells and the effect of green tea. Although hundreds of miRNAs have been identified in humans, only a small proportion (25.6%) of miRNAs are expressed in MCF-7 cells. Low concentration treatment with Polyphenon-60 significantly alters the miRNA expression profile in MCF-7 cells. Twenty three miRNAs have been identified with differential expression after a 48 h treatment with 10 μg/ml Polyphenon-60 (green tea extract). These miRNAs include miR-21 and miR-27 that were found to be down-regulated following treatment with green tea. These two miRNAs have previously been identified as being overexpressed in MCF-7 breast cancer cells, with miR-21 specifically implicated in down-regulating the tumor suppressor gene, tropomyosin-1. This data supports the hypothesis that Polyphenon-60-induced modification of the breast cancer miRNA expression profile contributes to the efficacy of green tea treatment. The resulting decrease in carcinogenesis is further supported by the altered miRNA regulation of potential oncogenes and tumor-suppressor genes.     

Down-regulation of miR-126 expression in colorectal cancer and its clinical significance

MicroRNAs (miRNAs) are non-coding RNAs that regulate the expression of target mRNAs. Altered expression of specific miRNAs in several tumor types has been reported. However, the expression levels of miR-126 in colorectal cancers are unclear. In this study, we compared the expression of miR-126 between colorectal cancer tissues and non-tumor tissues. The expression levels of miR-126 in colorectal cancer tissues were significantly lower than those in non-tumor tissues, and miR-126 overexpression inhibited the growth of colorectal cancer cells. This new information may help to clarify the molecular mechanisms involved in colorectal carcinogenesis and to indicate that miR-126 may be a novel suppressive microRNA and help to differentiate between malignant and normal colorectal tissue.     

Identification of aberrantly expressed miRNAs in rectal cancer

Disturbance of miRNA expression may play a key role in the initiation and progression of colorectal cancer (CRC). CRC should be viewed as a heterogeneous disease, but previous studies have only screened dysregulated miRNAs in CRC from a panel of 96, 145, 287 and 455 miRNAs, respectively. It is necessary to identify new aberrantly expressed miRNAs in rectal cancer. In this study tissue samples were derived from patients undergoing a surgical procedure to remove a portion of cancers. The expression profile of 904 miRNAs was analyzed using a miRCURY? LNA Array from 6-paired rectal cancers and normal tissues. The expression levels of 4 miRNAs were compared by real-time PCR between colon and rectal cancer, and also the expression levels of metastatic miRNAs in different stages of rectal cancer were analyzed. We found that 67 miRNA precursors are upregulated in rectal cancer (p<0.05) and 21 of those have never been reported in colorectal cancer (CRC); 39 miRNA precursors are downregulated (p<0.05) and 24 novel dysregulated miRNAs were identified in rectal cancer. miR-31, miR-126 and miR-143 are differentially expressed between colon cancer and rectal cancer. Here, we report an miRNA profile of rectal cancer, and we identified differential expression patterns of miRNAs between rectal and colon cancers. This novel information may suggest the potential roles of these miRNAs in the diagnosis of rectal cancer.     

Identification of new aberrantly expressed miRNAs in intestinal-type gastric cancer and its clinical significance

miRNAs are small 19 to 22 nucleotide sequences of RNA that negatively regulate gene expression. miRNA expression profiles may become useful biomarkers for diagnostics, prognosis and prediction of response to treat, and it could be a powerful tool for cancer prevention and therapeutics. Several miRNA expression profiles of miRNAs in gastric cancer have been reported, but these studies screened only few miRNAs and samples used in experiments include several different subtypes of gastric cancers, which decrease the sensitivity to identify new aberrant miRNAs. In this study, a miRNA expression profile was identified by miRCURY LNA Array (v.14.0) between intestinal-type gastric cancers and normal tissues. Forty miRNA precursors were up-regulated and thirty-six miRNA were down-regulated in intestinal-type gastric cancers (p<0.01). Sixteen new miRNAs were found in intestinal-type gastric cancers. Seventeen new miRNAs were found in intestinal-type gastric cancers. miR-145, miR-27a, miR-494 are differently expressed between intestinal-type and diffuse-type gastric cancers. miR-32, miR-182 and miR-143 dysregulated expression levels are related with different pathological stages of intestinal-type gastric cancers (p<0.01). Taken together, aberrantly expressed miRNAs may offer new clues to tumorigenesis of gastric cancers. miR-32, miR-182 and miR-143 may be potential diagnostic biomarkers for intestinal-type gastric cancers.     

miR-199a在消化系统肿瘤中的相关研究进展

microRNA（miRNA）是一类内源性非编码短链RNA。研究发现miRNA具有癌基因或抑癌基因样作用,参与多种恶性肿瘤的演进,是肿瘤发生、发展过程中的重要分子,其中miR-199a最受关注。miR-199a在消化系统肿瘤中表达上调,发挥着类似于癌基因的作用。miR-199a通过对靶基因的调控,增强肿瘤细胞的增殖、浸润及转移能力,且与消化系统肿瘤患者的无复发生存时间与总生存时间密切相关。体外研究证实,抑制miR-199a表达或阻碍其与靶基因的相互作用在消化系统肿瘤的治疗中具有广阔的应用前景。本文就miR-199a在消化系统肿瘤的调控机制、增殖与凋亡、侵袭、耐药与转移中的 作用,及其临床价值作一综述。