We analyzed microRNA profiles in predominantly HBV-related hepatocellular carcinomas in men and women and the association of microRNA expression levels with survival and response to therapy with interferon alfa in three independent cohorts. We found that the expression of miR-26a and miR-26b in nontumor hepatic tissues was higher in women than in men, but the expression was significantly down-regulated in tumor samples, as compared with paired samples of noncancerous tissues, regardless of sex. Tumors with reduced miR-26 expression had a distinct gene-expression profile, and patients whose tumors had low miR-26 expression had shorter survival but were more likely to have a response to interferon alfa, as compared with patients whose tumors had high miR-26 expression.
Our results suggest that miR-26 may be a tumor suppressor and that miR-26 silencing in hepatocytes may contribute to the development of a more aggressive form of hepatocellular carcinoma in men. These hypotheses are supported by the following findings: miR-26 was expressed at higher levels in women than in men in the liver, where presumably more anticarcinogenic activities exist in women; miR-26 expression is silenced in a subgroup of patients who have reduced survival; and genes that are activated in tumors with reduced miR-26 expression are selectively enriched in signaling pathways between NF-κ
B and interleukin-6. It was recently reported that MyD8-dependent interleukin-6 induction by NF-κ
B differed between male and female mice.11
Intriguingly, estrogens inhibit interleukin-6 promoter activity, which may contribute to a decreased susceptibility to hepatocellular carcinoma in women. Whether such a link is functionally relevant to liver cancer in humans remains to be determined.24
Our results are consistent with these findings, since interleukin-6 expression was inversely correlated with miR-26 expression.
Our analyses revealed that miR-26 expression was an independent predictor of survival. However, when outcomes of therapy with interferon alfa were stratified, only patients whose tumors had reduced miR-26 expression had a favorable response to interferon alfa in two independent, prospective, randomized, controlled trials. These results indicate that miR-26 status in tumors may be a useful tool in estimating prognosis in patients with hepatocellular carcinoma and in assisting in the selection of patients who are likely to benefit from adjuvant therapy with interferon alfa to prevent relapse. In an evaluation of the effects of adjuvant interferon alfa therapy after liver resection or tumor ablation in seven randomized, controlled trials, Clavien5
concluded that all the trials showed modest beneficial effects of the drug. In addition, among multiple experimental agents, only a modest survival benefit has been observed with sorafenib.6
The poor efficacy of current systemic agents may stem from an inability to identify a subpopulation of patients who may have a favorable response to a particular therapy. Prospective studies will be necessary to determine whether interferon alfa might be used as a first-line therapy for patients with hepatocellular carcinoma who have undergone resection and who have tumors with reduced miR-26 expression.
It should be noted that our results involved Chinese patients with a high rate of HBV positivity (approximately 90%). Our findings therefore need to be confirmed by studies of tumors from non-Asian patients and tumors arising from other underlying liver diseases, such as hepatitis C, or associated with alcohol abuse.
The mechanisms behind the sensitivity of tumors with reduced miR-26 expression to therapy with interferon alfa are unclear. One plausible model is that such tumors have specific activation of the signaling pathway that is responsive to interferon alfa. Consistently, tumors with low miR-26 expression were distinct from those with high miR-26 expression in transcriptomic activities and were associated with poor survival. Many of the genes that are overexpressed in tumors with reduced miR-26 expression are related to cell immunity, such as those encoding proinflammatory and antiinflammatory cytokines (i.e., inter-leukin-1, interleukin-2, interleukin-10, and inter-leukin-17). Moreover, many signaling networks that are activated in tumors with low miR-26 expression are immune-associated, such as NF-κ
B–interleukin-6, interleukin-10, STAT3, and inter-feron alfa inducible factor signaling networks (Table 4
in the Supplementary Appendix). Tumors with low miR-26 expression may be characterized by a unique activation of interferon alfa signaling, possibly through the NF-κ
B–inter leukin-6 signaling pathway, and thus may be sensitive to growth inhibition mediated by interferon alfa through interleukin-6–STAT3 signaling.25
These hypotheses require evaluation.
In conclusion, we identified systematic differences in patterns of microRNA expression between liver tissues obtained from men and women with hepatocellular carcinoma. Tumors with low miR-26 expression were biologically distinct from those with high expression and were associated with reduced survival but had a favorable response to adjuvant therapy with inter-feron alfa.