MicroRNAs (miRNAs) constitute a large class of short RNAs (e.g., 20-24 nucleotides in length), whose main function is to posttranscriptionally regulate the expression of protein-coding genes. Their importance in tumorigenesis has been demonstrated over the past decade, and correspondingly, they have emerged as potential therapeutic molecules and targets. Liver cancer is one of the most common neoplastic diseases worldwide, and it currently has a poor prognosis owing to largely ineffective therapeutic options. Liver cancer is also an excellent model for testing miRNA-based therapy approaches as it can be easily targeted with the systemic delivery of oligonucleotides. In recent years, the role of miRNAs in hepatocellular carcinoma (HCC) has been established with molecular studies and the development of animal models. These studies have also provided the basis for evaluating the therapeutic potential of miRNAs, or anti-miRNAs. In general, the safety of miRNAs has been proven and antitumor activity has been observed. Moreover, because of the absence or presence of mild side effects, the prophylactic use of miRNA-based approaches may be foreseen.

MicroRNA-221 (miR-221) is one of the most frequently and consistently up-regulated microRNAs (miRNAs) in human cancer. It has been hypothesized that miR-221 may act as a tumor promoter. To demonstrate this, we developed a transgenic (TG) mouse model that exhibits an inappropriate overexpression of miR-221 in the liver. Immunoblotting and immunostaining confirmed a concomitant down-regulation of miR-221 target proteins. This TG model is characterized by the emergence of spontaneous nodular liver lesions in approximately 50% of male mice and by a strong acceleration of tumor development in 100% of mice treated with diethylnitrosamine. Similarly to human hepatocellular carcinoma, tumors are characterized by a further increase in miR-221 expression and a concomitant inhibition of its target protein-coding genes (i.e., cyclin-dependent kinase inhibitor [Cdkn]1b/p27, Cdkn1c/p57, and B-cell lymphoma 2–modifying factor). To validate the tumor-promoting effect of miR-221, we showed that in vivo delivery of anti-miR-221 oligonucleotides leads to a significant reduction of the number and size of tumor nodules. Conclusions : This study not only establishes that miR-221 can promote liver tumorigenicity, but it also establishes a valuable animal model to perform preclinical investigations for the use of anti-miRNA approaches aimed at liver cancer therapy.

The microRNA 125b is a double-faced gene expression regulator described both as a tumor suppressor gene (in solid tumors) and an oncogene (in hematologic malignancies). In human breast cancer, it is one of the most down-regulated miRNAs and is able to modulate ERBB2/3 expression. Here, we investigated its targets in breast cancer cell lines after miRNA-mimic transfection. We examined the interactions of the validated targets with ERBB2 oncogene and the correlation of miR-125b expression with clinical variables.

The down-regulation of miR-199 occurs in nearly all primary hepatocellular carcinomas (HCCs) and HCC cell lines in comparison with normal liver. We exploited this miR-199 differential expression to develop a conditionally replication-competent oncolytic adenovirus, Ad-199T, and achieve tumor-specific viral expression and replication. To this aim, we introduced four copies of miR-199 target sites within the 3’ UTR of E1A gene, essential for viral replication. As consequence, E1A expression from Ad-199T virus was tightly regulated both at RNA and protein levels in HCC derived cell lines, and replication controlled by the level of miR-199 expression. Various approaches were used to asses in vivo properties of Ad-199T. Ad-199T replication was inhibited in normal, miR-199 positive, liver parenchyma, thus resulting in reduced hepatotoxicity. Conversely, the intrahepatic delivery of Ad-199T in newborn mice led to virus replication and fast removal of implanted HepG2 liver cancer cells. The ability of Ad-199T to control tumor growth was also shown in a subcutaneous xenograft model in nude mice and in HCCs arising in immune-competent mice. In summary, we developed a novel oncolytic adenovirus, Ad-199T, which could demonstrate a therapeutic potential against liver cancer without causing significant hepatotoxicity.

hsa-mir-483 is located within intron 2 of the IGF2 gene. We have previously shown oncogenic features of miR-483-3p through cooperation with IGF2 or by independently targeting the proapoptotic gene BBC3/PUMA. Here we demonstrate that expression of miR-483 can be induced independently of IGF2 by the oncoprotein β-catenin through an interaction with the basic helix–loop–helix protein upstream stimulatory transcription factor 1. We also show that β-catenin itself is a target of miR-483-3p, triggering a negative regulatory loop that becomes ineffective in cells harboring an activating mutation of β-catenin. These results provide insights into the complex regulation of the IGF2/miR-483 locus, revealing players in the β-catenin pathway.

Purpose: Despite its indolent nature, chronic lymphocytic leukemia (CLL) remains an incurable disease. To establish the potential pathogenic role of miRNAs, the identification of deregulated miRNAs in CLL is crucial. Experimental Design: We analyzed the expression of 723 mature miRNAs in 217 early-stage CLL cases and in various different normal B-cell subpopulations from tonsils and peripheral blood. Results: Our analyses indicated that CLL cells exhibited a miRNA expression pattern that was most similar to the subsets of antigen-experienced and marginal zone–like B cells. These normal subpopulations were used as reference to identify differentially expressed miRNAs in comparison with CLL. Differences related to the expression of 25 miRNAs were found to be independent from IGHV mutation status or cytogenetic aberrations. These differences, confirmed in an independent validation set, led to a novel comprehensive description of miRNAs potentially involved in CLL. We also identified miRNAs whose expression was distinctive of cases with mutated versus unmutated IGHV genes or cases with 13q, 11q, and 17p deletions and trisomy 12. Finally, analysis of clinical data in relation to miRNA expression revealed that miR26a, miR532-3p, miR146-5p, and miR29c* were strongly associated with progression-free survival. Conclusion: This study provides novel information on miRNAs expressed by CLL and normal B-cell subtypes, with implication on the cell of origin of CLL. In addition, our findings indicate a number of deregulated miRNAs in CLL, which may play a pathogenic role and promote disease progression. Collectively, this information can be used for developing miRNA-based therapeutic strategies in CLL. Clin Cancer Res; 20(15); 4141–53. ©2014 AACR.