Carcinogen-induced hepatic tumors in KLF6+/− mice recapitulate aggressive human hepatocellular carcinoma associated with p53 pathway deregulation

Authors

Mirko Tarocchi, Rebekka Hannivoort, Yujin Hoshida, Ursula E. Lee, Diana Vetter, Goutham Narla, Augusto Villanueva, Moshe Oren, Josep M. Llovet, Scott L. Friedman

Abstract

Inactivation of KLF6 is common in hepatocellular carcinoma (HCC) associated with hepatitis C virus (HCV) infection, thereby abrogating its normal antiproliferative activity in liver cells. The aim of the study was to evaluate the impact of KLF6 depletion on human HCC and experimental hepatocarcinogenesis in vivo. In patients with surgically resected HCC, reduced tumor expression of KLF6 was associated with decreased survival. Consistent with its role as a tumor suppressor, KLF6+/− mice developed significantly more tumors in response to the chemical carcinogen diethyl nitrosamine (DEN) than wild-type animals. Gene expression signatures in both surrounding tissue and tumors of KLF6+/− mice closely recapitulated those associated with aggressive human HCCs. Expression microarray profiling also revealed an increase in Mdm2 mRNA in tumors from KLF6+/− compared with KLF6+/+ mice, which was validated by way of quantitative real-time polymerase chain reaction and western blot analysis in both human HCC and DEN-induced murine tumors. Moreover, chromatin immunoprecipitation and cotransfection assays established the P2 intronic promoter of Mdm2 as a bona fide transcriptional target repressed by KLF6. Whereas KLF6 overexpression in HCC cell lines and primary hepatocytes led to reduced MDM2 levels and increased p53 protein and transcriptional activity, reduction in KLF6 by small interfering RNA led to increased MDM2 and reduced p53. Our findings indicate that KLF6 deficiency contributes significantly to the carcinogenic milieu in human and murine HCC and uncover a novel tumor suppressor activity of KLF6 in HCC by linking its transcriptional repression of Mdm2 to stabilizing p53.