Hansjuerg Alder

MicroRNA-29 family reverts aberrant methylation in lung cancer by targeting DNA methyltransferases 3A and 3B

MicroRNAs (miRNAs) are small, noncoding RNAs that regulate expression of many genes. Recent studies suggest roles of miRNAs in carcinogenesis. We and others have shown that expression profiles of miRNAs are different in lung cancer vs. normal lung, although the significance of this aberrant expression is poorly understood. Among the reported down-regulated miRNAs in lung cancer, the miRNA (miR)-29 family (29a, 29b, and 29c) has intriguing complementarities to the 3′-UTRs of DNA methyltransferase (DNMT)3A and -3B (de novo methyltransferases), two key enzymes involved in DNA methylation, that are frequently up-regulated in lung cancer and associated with poor prognosis. We investigated whether miR-29s could target DNMT3A and -B and whether restoration of miR-29s could normalize aberrant patterns of methylation in non-small-cell lung cancer. Here we show that expression of miR-29s is inversely correlated to DNMT3A and -3B in lung cancer tissues, and that miR-29s directly target both DNMT3A and -3B. The enforced expression of miR-29s in lung cancer cell lines restores normal patterns of DNA methylation, induces reexpression of methylation-silenced tumor suppressor genes, such as FHIT and WWOX, and inhibits tumorigenicity in vitro and in vivo. These findings support a role of miR-29s in epigenetic normalization of NSCLC, providing a rationale for the development of miRNA-based strategies for the treatment of lung cancer.

A MicroRNA Signature of Hypoxia

ABSTRACT Recent research has identified critical roles for microRNAs in a large number of cellular processes, including tumorigenic transformation. While significant progress has been made towards understanding the mechanisms of gene regulation by microRNAs, much less is known about factors affecting the expression of these noncoding transcripts. Here, we demonstrate for the first time a functional link between hypoxia, a well-documented tumor microenvironment factor, and microRNA expression. Microarray-based expression profiles revealed that a specific spectrum of microRNAs (including miR-23, -24, -26, -27, -103, -107, -181, -210, and -213) is induced in response to low oxygen, at least some via a hypoxia-inducible-factor-dependent mechanism. Select members of this group (miR-26, -107, and -210) decrease proapoptotic signaling in a hypoxic environment, suggesting an impact of these transcripts on tumor formation. Interestingly, the vast majority of hypoxia-induced microRNAs are also overexpressed in a variety of human tumors.

The t(4;11) chromosome translocation of human acute leukemias fuses the ALL-1 gene, related to Drosophila trithorax, to the AF-4 gene

The ALL-1 gene located at human chromosome 11 band q23 is rearranged in acute leukemias with interstitial deletions or reciprocal translocations between this region and chromosomes 1, 4, 6, 9, 10, or 19. The gene spans approximately 100 kb of DNA and contains at least 21 exons. It encodes a protein of more than 3910 amino acids containing three regions with homology to sequences within the Drosophila trithorax gene, including cysteine-rich regions that can be folded into six zinc finger-like domains. The breakpoint cluster region within ALL-1 spans 8 kb and encompasses several small exons, most of which begin in the same phase of the open reading frame. The t(4;11) chromosome translocation results in two reciprocal fusion products coding for chimeric proteins derived from ALL-1 and from a gene on chromosome 4. This suggests that each 11q23 abnormality gives rise to a specific oncogenic fusion protein.

E2F1-Regulated MicroRNAs Impair TGFβ-Dependent Cell-Cycle Arrest and Apoptosis in Gastric Cancer

Deregulation of E2F1 activity and resistance to TGFbeta are hallmarks of gastric cancer. MicroRNAs (miRNAs) are small noncoding RNAs frequently misregulated in human malignancies. Here we provide evidence that the miR-106b-25 cluster, upregulated in a subset of human gastric tumors, is activated by E2F1 in parallel with its host gene, Mcm7. In turn, miR-106b and miR-93 regulate E2F1 expression, establishing a miRNA-directed negative feedback loop. Furthermore, upregulation of these miRNAs impairs the TGFbeta tumor suppressor pathway, interfering with the expression of CDKN1A (p21(Waf1/Cip1)) and BCL2L11 (Bim). Together, these results suggest that the miR-106b-25 cluster is involved in E2F1 posttranscriptional regulation and may play a key role in the development of TGFbeta resistance in gastric cancer.

MicroRNAs bind to Toll-like receptors to induce prometastatic inflammatory response

MicroRNAs (miRNAs) are small noncoding RNAs, 19–24 nucleotides in length, that regulate gene expression and are expressed aberrantly in most types of cancer. MiRNAs also have been detected in the blood of cancer patients and can serve as circulating biomarkers. It has been shown that secreted miRNAs within exosomes can be transferred from cell to cell and can regulate gene expression in the receiving cells by canonical binding to their target messenger RNAs. Here we show that tumor-secreted miR-21 and miR-29a also can function by another mechanism, by binding as ligands to receptors of the Toll-like receptor (TLR) family, murine TLR7 and human TLR8, in immune cells, triggering a TLR-mediated prometastatic inflammatory response that ultimately may lead to tumor growth and metastasis. Thus, by acting as paracrine agonists of TLRs, secreted miRNAs are key regulators of the tumor microenvironment. This mechanism of action of miRNAs is implicated in tumor–immune system communication and is important in tumor growth and spread, thus representing a possible target for cancer treatment.

MiR-15a and miR-16-1 cluster functions in human leukemia

MicroRNAs (miRNAs) are short noncoding RNAs regulating gene expression that play roles in human diseases, including cancer. Each miRNA is predicted to regulate hundreds of transcripts, but only few have experimental validation. In chronic lymphocytic leukemia (CLL), the most common adult human leukemia, miR-15a and miR-16-1 are lost or down-regulated in the majority of cases. After our previous work indicating a tumor suppressor function of miR-15a/16-1 by targeting the BCL2 oncogene, here, we produced a high-throughput profiling of genes modulated by miR-15a/16-1 in a leukemic cell line model (MEG-01) and in primary CLL samples. By combining experimental and bioinformatics data, we identified a miR-15a/16-1-gene signature in leukemic cells. Among the components of the miR-15a/16-1 signature, we observed a statistically significant enrichment in AU-rich elements (AREs). By examining the Gene Ontology (GO) database, a significant enrichment in cancer genes (such as MCL1, BCL2, ETS1, or JUN) that directly or indirectly affect apoptosis and cell cycle was found.

miR-221&222 regulate TRAIL resistance and enhance tumorigenicity through PTEN and TIMP3 downregulation.

Lung and liver cancers are among the most deadly types of cancer. Despite improvements in treatment over the past few decades, patient survival remains poor, underlining the need for development of targeted therapies. MicroRNAs represent a class of small RNAs frequently deregulated in human malignancies. We now report that miR-221&222 are overexpressed in aggressive non-small cell lung cancer and hepatocarcinoma cells, as compared with less invasive and/or normal lung and liver cells. We show that miR-221&222, by targeting PTEN and TIMP3 tumor suppressors, induce TRAIL resistance and enhance cellular migration through the activation of the AKT pathway and metallopeptidases. Finally, we demonstrate that the MET oncogene is involved in miR-221&222 activation through the c-Jun transcription factor.

MicroRNA Signatures Associated with Cytogenetics and Prognosis in Acute Myeloid Leukemia

MicroRNAs (miRNAs) are small RNAs of 19 to 25 nucleotides that are negative regulators of gene expression. To determine whether miRNAs are associated with cytogenetic abnormalities and clinical features in acute myeloid leukemia (AML), we evaluated the miRNA expression of CD34(+) cells and 122 untreated adult AML cases using a microarray platform. After background subtraction and normalization using a set of housekeeping genes, data were analyzed using Significance Analysis of Microarrays. An independent set of 60 untreated AML patients was used to validate the outcome signatures using real-time polymerase chain reaction. We identified several miRNAs differentially expressed between CD34(+) normal cells and the AML samples. miRNA expression was also closely associated with selected cytogenetic and molecular abnormalities, such as t(11q23), isolated trisomy 8, and FLT3-ITD mutations. Furthermore, patients with high expression of miR-191 and miR-199a had significantly worse overall and event-free survival than AML patients with low expression (overall survival: miR-191, P = .03; and miR-199a, P = .001, Cox regression). In conclusion, miRNA expression in AML is closely associated with cytogenetics and FLT3-ITD mutations. A small subset of miRNAs is correlated with survival.

Relation between microRNA expression and progression and prognosis of gastric cancer: a microRNA expression analysis

BACKGROUND Analyses of microRNA expression profiles have shown that many microRNAs are expressed aberrantly and correlate with tumorigenesis, progression, and prognosis of various haematological and solid tumours. We aimed to assess the relation between microRNA expression and progression and prognosis of gastric cancer. METHODS 353 gastric samples from two independent subsets of patients from Japan were analysed by microRNA microarray. MicroRNA expression patterns were compared between non-tumour mucosa and cancer samples, graded by diffuse and intestinal histological types and by progression-related factors (eg, depth of invasion, metastasis, and stage). Disease outcome was calculated by multivariable regression analysis to establish whether microRNAs are independent prognostic factors. FINDINGS In 160 paired samples of non-tumour mucosa and cancer, 22 microRNAs were upregulated and 13 were downregulated in gastric cancer; 292 (83%) samples were distinguished correctly by this signature. The two histological subtypes of gastric cancer showed different microRNA signatures: eight microRNAs were upregulated in diffuse-type and four in intestinal-type cancer. In the progression-related signature, miR-125b, miR-199a, and miR-100 were the most important microRNAs involved. Low expression of let-7g (hazard ratio 2.6 [95% CI 1.3-4.9]) and miR-433 (2.1 [1.1-3.9]) and high expression of miR-214 (2.4 [1.2-4.5]) were associated with unfavourable outcome in overall survival independent of clinical covariates, including depth of invasion, lymph-node metastasis, and stage. INTERPRETATION MicroRNAs are expressed differentially in gastric cancers, and histological subtypes are characterised by specific microRNA signatures. Unique microRNAs are associated with progression and prognosis of gastric cancer. FUNDING National Cancer Institute.

The detection of differentially expressed microRNAs from the serum of ovarian cancer patients using a novel real-time PCR platform

OBJECTIVE To determine the utility of serum miRNAs as biomarkers for epithelial ovarian cancer. METHODS Twenty-eight patients with histologically confirmed epithelial ovarian cancer were identified from a tissue and serum bank. Serum was collected prior to definitive therapy. Fifteen unmatched, healthy controls were used for comparison. Serum was obtained from all patients. RNA was extracted using a derivation of the single step Trizol method. The RNA from 9 cancer specimens was compared to 4 normal specimens with real-time PCR using the TaqMan Array Human MicroRNA panel. Twenty-one miRNAs were differentially expressed between normal and patient serum. Real-time PCR for the 21 individual miRNAs was performed on the remaining 19 cancer specimens and 11 normal specimens. RESULTS Eight miRNAs of the original twenty-one were identified that were significantly differentially expressed between cancer and normal specimens using the comparative C(t) method. MiRNAs-21, 92, 93, 126 and 29a were significantly over-expressed in the serum from cancer patients compared to controls (p<.01). MiRNAs-155, 127 and 99b were significantly under-expressed (p<.01). Additionally, miRs-21, 92 and 93 were over-expressed in 3 patients with normal pre-operative CA-125. CONCLUSION We demonstrate that the extraction of RNA and subsequent identification of miRNAs from the serum of individuals diagnosed with ovarian cancer is feasible. Real-time PCR-based microarray is a novel and practical means to performing high-throughput investigation of serum RNA samples. miRNAs-21, 92 and 93 are known oncogenes with therapeutic and biomarker potential.