The performance of circulating biomarkers for the diagnosis of hepatocellular carcinoma (HCC) is sub-optimal. In this study we tested circulating microRNAs as biomarkers for HCC in cirrhotic patients by performing a two stage study: a discovery phase conducted by microarray and a validation phase performed by qRT-PCR in an independent series of 118 patients. Beside miRNAs emerged from the discovery phase, miR-21, miR-221, miR-519d were also tested in the validation setting on the basis of literary and tissue findings. Deregulated microRNAs were assayed in HCC-derived cells in the intracellular compartment, cell culture supernatant and exosomal fraction. Serum and tissue microRNA levels were compared in 14 patients surgically treated for HCC. From the discovery study, it emerged that seven circulating microRNAs were differentially expressed in cirrhotic patients with and without HCC. In the validation set, miR-939, miR-595 and miR-519d were shown to differentiate cirrhotic patients with and without HCC. MiR-939 and miR-595 are independent factors for HCC. ROC curves of miR-939, miR-595 and miR-519d displayed that AUC was higher than AFP. An exosomal secretion of miR-519d, miR-21, miR-221 and miR-1228 and a correlation between circulating and tissue levels of miR-519d, miR-494 and miR-21 were found in HCC patients. Therefore, we show that circulating microRNAs deserve attention as non-invasive biomarkers in the diagnostic setting of HCC and that exosomal secretion contributes to discharging a subset of microRNAs into the extracellular compartment.

The aberrant expression of miR-221 is a hallmark of human cancers, including hepatocellular carcinoma (HCC), and its involvement in drug resistance, together with a proved efficacy of anti-miR-221 molecules, strengthen its role as an attractive target candidate in the oncologic field. The discovery of biomarkers predicting the response to treatments represents a clinical challenge in the personalized treatment era. This study aimed to investigate the possible role of miR-221 as a circulating biomarker in HCC patients undergoing sorafenib treatment as well as to evaluate its contribution to sorafenib resistance in advanced HCC. A chemically induced HCC rat model and a xenograft mouse model, together with HCC-derived cell lines were employed to analyze miR-221 modulation by Sorafenib treatment. Data from the functional analysis were validated in tissue samples from surgically resected HCCs. The variation of circulating miR-221 levels in relation to Sorafenib treatment were assayed in the animal models and in two independent cohorts of patients with advanced HCC. MiR-221 over-expression was associated with Sorafenib resistance in two HCC animal models and caspase-3 was identified as its target gene, driving miR-221 anti-apoptotic activity following Sorafenib administration. Lower pre-treatment miR-221 serum levels were found in patients subsequently experiencing response to Sorafenib and an increase of circulating miR-221 at the two months assessment was observed in responder patients. MiR-221 might represent a candidate biomarker of likelihood of response to Sorafenib in HCC patients to be tested in future studies. Caspase-3 modulation by miR-221 participates to Sorafenib resistance. .

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CEUS misdiagnosed as HCC a significantly higher number of ICC lesions in cirrhotic patients than CT and MRI. However, some CEUS contrast features can help suspect ICC, especially in some cases with inconclusive CT or MRI.

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SUMMARY BackgroundGadoxetic acid (Gd-EOB-DTPA) is a 'hepatocyte-specific' contrast agent for magnetic resonance (MR) in both the vascular and the hepatobiliary phases.

These data confirm that the HYPER-HYPO pattern at CEUS is definitely diagnostic for HCC in cirrhosis and that the HYPER-ISO pattern has a similar PPV, indicating that this pattern is highly suspicious for HCC. The ISO-ISO pattern was found in > 50% of recurrent nodules and indicates a high risk of HCC.

The overexpression of microRNA-221 (miR-221) is reported in several human cancers including hepatocellular carcinoma, and its targeting by tailored treatments has been proposed. The evidence supporting the role of miR-221 in cancer is growing and has been mainly focused on the discovery of miR-221 targets as well as on its possible therapeutic exploitations. However, the mechanism sustaining miR-221 aberrant expression remains to be elucidated. In this study, MDM2 (E3 ubiquitin-protein ligase homolog), a known p53 (TP53) modulator, is identified as a direct target of miR-221, and a feed-forward loop is described that sustains miR-221 aberrant expression. Interestingly, miR-221 can activate the p53/mdm2 axis by inhibiting MDM2 and, in turn, p53 activation contributes to miR-221 enhanced expression. Moreover, by modulating the p53 axis, miR-221 impacts cell-cycle progression and apoptotic response to doxorubicin in hepatocellular carcinoma-derived cell lines. Finally, CpG island methylation status was assessed as a causative event associated with miR-221 upregulation in hepatocellular carcinoma cells and primary tumor specimens. In hepatocellular carcinoma-derived cell lines, pharmacologically induced DNA hypomethylation potentiated a significant increase in miR-221 expression. These data were confirmed in clinical specimens of hepatocellular carcinoma in which elevated miR-221 expression was associated with the simultaneous presence of wild-type p53 and DNA hypomethylation.Implications: These findings reveal a novel miR-221-sustained regulatory loop that determines a p53-contextspecific response to doxorubicin treatment in hepatocellular carcinoma. Mol Cancer Res; 12(2); 203-16. Ó2013 AACR.Introduction microRNAs (miRNA) play a pivotal role in cancer cell biology. Because of their ability to simultaneously control several cancer-related pathways, miRNAs represent attractive therapeutic targets. Each miRNA triggers different molecular events depending both on the cell type and on the molecular context. Indeed, the same miRNA can exert diverse functions in different settings (1). Thus, it is not