Differential sensitization of hepatocytes by inhibitors of histone deacetylases against death receptor-induced apoptosis

Aims: Preclinical and clinical studies suggest that inhibition of histone/protein deacetylation by targeting histone deacetylases (HDACs) represents a novel approach in cancer therapy [1]. However, only recently it became evident that HDAC inhibitors by themselves do not only induce apoptosis but also render cells sensitive to death receptor agonists, i.e. TNF, TRAIL, and CD95 ligand (CD95L), respectively [2, 3]. For a comprehensive understanding of the pharmacodynamic properties of HDAC inhibitors (HDIs) it is madatory to elucidate and profile these sensitizing effects. Results: HepG2 cells pre-incubated with subtoxic concentrations of protein biosynthesis inhibitors (i.e. actinomycin D, cycloheximide or alpha-amanitin) are known to be prone to apoptosis induced by death receptor agonists. In the additional presence of HDIs, these cells were much more susceptible to TRAIL or CD95 elicited apoptosis in a time- and concentration-dependent manner. In contrast, HDIs failed to sensitize against TNF-induced cell death. Upon direct activation of the intrinsic death signaling pathway via UV radiation or in a cell-free system, HDIs did not further increase cell death and caspase–3 activation. However, in the presence of HDIs, an increased rate of apoptosis and enhanced activation of downstream caspases was observed when the extrinsic death signaling pathway was activated by transient expression of caspase–8. Following incubation with HDIs, expression levels of various pro-apoptotic proteins such as caspases, Bax, Bid and FADD remained largely unchanged whereas proteins with anti-apoptotic properties, i.e. FLIP and XIAP were markedly down-regulated. These findings in HepG2 cells were largely confirmed in primary murine hepatocytes. Moreover, in mouse livers perfused in situ, HDIs strongly exacerbated CD95-mediated liver injury as determined by detection of alanine aminotransferase in the perfusate. Conclusions: This study reveals that HDIs can selectively sensitize hepatocytes against TRAIL and CD95L- but not against TNF-induced apoptosis. As a mechanistic rationale, we provide evidence for a preferential activation of proteins associated with the extrinsic signaling pathway of apoptosis. These results are discussed in the context of the pharmacodynamic properties of HDIs of e.g. hepatocellular carinoma, i.e. under conditions where hepatic death receptors are known to be upregulated.

Literatur: 1. McLaughlin F et al. Histone deacetylase inhibitors open new doors in cancer therapy. Biochem Pharmacol 2004, 68:1139-44 2. Chopin V et al. Synergistic induction of apoptosis in breast cancer cells by cotreatment with butyrate and TNF-alpha, TRAIL, or anti-Fas agonist antibody involves enhancement of death receptors' signaling and requires P21(waf1). Exp Cell Res. 2004, 298:560-73. 3. Nakata S et al. Histone deacetylase inhibitors upregulate death receptor 5/TRAIL-R2 and sensitize apoptosis induced by TRAIL/APO2-L in human malignant tumor cells. Oncogene. 2004; 23:6261-71.

Schlüsselwörter

HepG2 - TNF - apoptosis - death receptors