Histone deacetylase 6 functions as a tumor suppressor by activating c-Jun NH2-terminal kinase-mediated beclin 1-dependent autophagic cell death in liver cancer

Abstract

Ubiquitin-binding histone deacetylase 6 (HDAC6) is uniquely endowed with tubulin deacetylase activity and plays an important role in the clearance of misfolded protein by autophagy. In cancer, HDAC6 has become a target for drug development due to its major contribution to oncogenic cell transformation. In the present study we show that HDAC6 expression was down-regulated in a large cohort of human hepatocellular carcinoma (HCC) patients, and that low expression of HDAC6 was significantly associated with poor prognosis of HCC patients in 5-year overall, disease-free, and recurrence-free survival. Notably, we observed that ectopic overexpression of HDAC6 suppressed tumor cell growth and proliferation in various liver cancer cells, and elicited increased LC3B-II conversion and autophagic vacuole formation without causing apoptotic cell death or cell cycle inhibition. In addition, the sustained overexpression of HDAC6 reduced the in vivo tumor growth rate in a mouse xenograft model. It was also found that HDAC6 mediated autophagic cell death by way of Beclin 1 and activation of the LC3-II pathway in liver cancer cells, and that HDAC6 overexpression activated c-Jun NH2-terminal kinase (JNK) and increased the phosphorylation of c-Jun. In contrast, the induction of Beclin 1 expression was blocked by SP600125 (a specific inhibitor of JNK) or by small interfering RNA directed against HDAC6. Conclusion: Our findings suggest that loss of HDAC6 expression in human HCCs and tumor suppression by HDAC6 occur by way of activation of caspase-independent autophagic cell death through the JNK/Beclin 1 pathway in liver cancer and, thus, that a novel tumor suppressor function mechanism involving HDAC6 may be amenable to nonepigenetic regulation. (HEPATOLOGY 2012;56:644-657) H epatocellular carcinoma (HCC) is an aggressive form of cancer, the fifth most common cancer, and the third leading cause of cancer death worldwide. 1 Surgery with curative intent is feasible for only 15% to 25% of patients and most HCC patients die from locally advanced or metastatic disease in a relatively short period of time. 2 Hepatitis B virus, hepatitis C virus, and aflatoxin B1 are well-known major causes of HCC. However, the overall survival of patients with HCC has not improved significantly over the past two decades, and the mechanisms responsible for the development and progression of HCC remain poorly understood. 3 To date, molecular targeted therapy has shown promise for the treatment of advanced Abbreviations: 3-MA, 3-methyladenine; FACS, fluorescence-activated cell sorting; FBS, fetal bovine serum; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HAT, histone acetyltransferase; HCC, hepatocellular carcinoma; HDAC, histone deacetylases; JNK, c-Jun NH2-terminal kinase; LC3, microtubule-associated protein 1 light chain 3; mRNA, messenger RNA; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; PARP, poly (ADP-ribose) polymerase; siRNA, small interfering RNA; TEM, transmission electron microscopy; TMA, tissue microarray.From the

HDAC2 Provides a Critical Support to Malignant Progression of Hepatocellular Carcinoma through Feedback Control of mTORC1 and AKT

“…S1C). In addition, we performed GSEA using three different cancer cell lines (liver, lung, and gastric cancer) to confirm HDAC2 signatures in these cell lines (7,9,10). Indeed, GSEA showed that EGFR system-related gene elements were commonly enriched in HDAC2 signatures observed in these cell lines (Fig.…”

“…S1A). Because the mRNA and protein levels of HDAC2 were upregulated in overt hepatocellular carcinoma, we next assessed the prognostic implications of HDAC2 expression in a large cohort of patients with hepatocellular carcinoma described in our previous study (10). First, 3,039 genes with expression patterns highly correlated with HDAC2 expression were selected for cluster analysis (P < 0.05, r > 0.4 or r < À0.4; Fig.…”

“…6D). We then investigated the mRNA expression patterns of several genes encoding a specific kinase or phosphatase in the phosphoinositide signaling pathway (these genes were identified during our previous microarray analysis of human hepatocellular carcinoma tissues (10). We found that the expressions of kinasecoding genes (PIK3CB, PIK3R2, PIP5K2C, and PIP5K2C) were upregulated, whereas the expressions of phosphatase-encoding genes (PTEN, INPP5B, and INPP5F) were downregulated in overt hepatocellular carcinoma (Supplementary Fig.…”

Limited efficacy of specific HDAC6 inhibition in urothelial cancer cells

“…Similarly variable results on HDAC6 expression levels have been published for hepatocellular cancers. 27,37,38 Furthermore, HDAC6 has been reported to stimulate proliferation in the latter tumor type. In breast cancer, HDAC6 expression was likewise variable 29 and particularly strong in small tumors with low histological grade.…”

“…52,53 In liver cancer, HDAC6 did not affect proliferation, too, but regulated autophagy. 38 Similarly, the newly developed HDAC6-specific inhibitor C1A generated a significant growth inhibitory effect across a number of different cancer cell lines by induction of apoptosis rather than cell cycle arrest, in contrast to the effect of the pan-HDAC inhibitor SAHA. 51 Our findings showing only a moderate induction of apoptosis after inhibition of enzyme activity with even higher concentrations of Tubacin, but no induction of apoptosis after diminution of protein expression by siRNA knockdown, indicate that neither HDAC6 protein expression nor enzymatic activity are critical for urothelial carcinoma cells in general.…”