A quadruple therapy synergistically blocks proliferation and promotes apoptosis of hepatoma cells.

ABSTRACT

Effective therapy for advanced hepatocellular carcinoma (HCC) is lacking. Conventional chemotherapy was judged to be ineffective. We previously demonstrated that the histone deacetylase inhibitor Trichostatin A (TSA) blocks growth of HCC cells in vitro. The anti-tumoral effect of a combination of more than 2 classes of drugs remains unexplored. Four hepatoma cell lines were incubated with increasing concentrations of Tamoxifen (TAM), 9-cis retinoic acid (CRA), the methioninaminopeptidase inhibitor TNP-470 and TSA as single agents and in combination. Anti-proliferative and pro-apoptotic effects were assessed using BrdU-incorporation, FACS analysis and immunocytochemistry. Central pro- and anti-apoptotic proteins were measured by semi-quantitative Western blotting and substrate assays. All single substances inhibited proliferation and induced apoptosis in HCC cells only at high concentrations. The combination of TAM/CRA/TNP/TSA multiplied the anti-tumoral effects, reaching up to 93% inhibition of proliferation and 63% induction of apoptosis after 24 h in Hep1B cells. Pro-apoptotic factors bax and caspase 3 were highly increased with quadruple therapy, while anti-apoptotic bcl-2 decreased to undetectable levels. Fibroblasts remained largely unaffected. While the single substances were not effective on hepatoma cells in tolerable doses, their combination significantly increases anti-tumoral efficacy. Combination therapy with biomodulators is a promising treatment option for HCC.