RESUMO
Both EZH2 and its homolog EZH1 function as histone H3 Lysine 27 (H3K27) methyltransferases and repress the transcription of target genes. Dysregulation of H3K27 trimethylation (H3K27me3) plays an important role in the development and progression of cancers such as hepatocellular carcinoma (HCC). This study investigated the relationship between the expression of EZH1/2 and the level of H3K27me3 in HCC. Additionally, the role of EZH1/2 in cell growth, tumorigenicity, and resistance to sorafenib were also analyzed. Both the lentiviral knockdown and the pharmacological inhibition of EZH1/2 (UNC1999) diminished the level of H3K27me3 and suppressed cell growth in liver cancer cells, compared with EZH1 or EZH2 single knockdown. Although a significant association was observed between EZH2 expression and H3K27me3 levels in HCC samples, overexpression of EZH1 appeared to contribute to enhanced H3K27me3 levels in some EZH2lowH3K27me3high cases. Akt suppression following sorafenib treatment resulted in an increase of the H3K27me3 levels through a decrease in EZH2 phosphorylation at serine 21. The combined use of sorafenib and UNC1999 exhibited synergistic antitumor effects in vitro and in vivo. Combination treatment canceled the sorafenib-induced enhancement in H3K27me3 levels, indicating that activation of EZH2 function is one of the mechanisms of sorafenib-resistance in HCC. In conclusion, sorafenib plus EZH1/2 inhibitors may comprise a novel therapeutic approach in HCC.
Assuntos
Antineoplásicos/uso terapêutico , Benzamidas/uso terapêutico , Carcinoma Hepatocelular/terapia , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidores , Indazóis/uso terapêutico , Neoplasias Hepáticas/terapia , Piperazinas/uso terapêutico , Complexo Repressor Polycomb 2/antagonistas & inibidores , Piridonas/uso terapêutico , Sorafenibe/uso terapêutico , Idoso , Animais , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Regulação para Baixo/efeitos dos fármacos , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Feminino , Terapia Genética , Humanos , Neoplasias Hepáticas/genética , Masculino , Camundongos SCID , Pessoa de Meia-Idade , Complexo Repressor Polycomb 2/genéticaRESUMO
Hochuekkito (HET), a Kampo herbal medicine composed of ten medicinal plants, is traditionally used to improve the general state of patients with malignant diseases such as cancer. Recent studies showed that HET had an anticancer effect against several cancer cell lines in vitro by inducing apoptosis. However, high doses of HET may have cytotoxic effects attributed to saponins or detergentlike compounds. Therefore, the present study used low doses of HET (50 µg/ml), which did not affect cell viability, to evaluate its synergistic anticancer effects with cisplatin. HeLa cells were cultured for 24 h with 50 µg/ml HET, followed by cisplatin treatment for 24 h at various concentrations. Subsequently, the sensitivity of the cells to cisplatin was assessed using a colony survival and a crystal violet cell viability assay. Furthermore, cisplatininduced apoptosis was analyzed by flow cytometry. Proteins associated with cell viability and apoptosis, including phosphorylated (p)Akt, p53, Bcell lymphoma 2 (Bcl2), Bcl2associated X protein (Bax) and active caspase3 were analyzed by immunoblotting. The present study revealed that cell survival was decreased and apoptosis was increased in HeLa cells pretreated with HET prior to cisplatin treatment compared with HETuntreated cells. Furthermore, protein expression of p53 and active caspase3 was increased, while the expression of pAkt as well as the Bcl2/Bax ratio, an index of survival activity in cells, were decreased in the HETpretreated cells compared with those in HETuntreated cells following incubation with cisplatin. In conclusion, the present study indicated that HET enhanced cisplatininduced apoptosis of HeLa cells and that the administration of HET may therefore be clinically beneficial alongside apoptosisinducing chemotherapy.