Suppression of STAT3 phosphorylation enhances the cytotoxicity of cucurbitacin B in B16F10 melanoma cells

Abstract

Cucurbitacin B (CuB) is a triterpenoid compound extracted from Cucurbitaceae plants, which has been shown to induce the phosphorylation (activation) of signal transducer and activator of transcription 3 (STAT3) in melanoma cells. In this study, we aimed to investigate whether inhibition of STAT3 phosphorylation could influence the cytotoxicity of CuB in melanoma cells. The results showed that pretreatment of B16F10 cells with AG490, an inhibitor of Janus kinase 2 (Jak2), enhanced the inhibitory effects of CuB on cell growth and G2/M phase arrest. DNA content analysis demonstrated that combined AG490 and CuB treatment increased the proportion of apoptotic cells, but decreased the ratio of tetraploid cells. Western blot analysis revealed that AG490 partially suppressed the levels of phosphorylated STAT3 (p-STAT3) and p38 (p-p38) in CuB-treated cells. Moreover, CuB-induced STAT3 phosphorylation could be completely blocked by SP600125, a specific inhibitor of c-Jun N-terminal kinase (JNK) signaling. In addition, SP600125 could also significantly augment CuB-induced apoptotic cell death. These results demonstrated that CuB-induced STAT3 activation was mediated by both JNK and Jak2 signaling and blocking STAT3 activation enhanced the cytotoxicity of CuB in B16F10 melanoma cells. Key words: AG490, cucurbitacin B, B16F10 melanoma cells, signal transducer and activator of transcription 3 (STAT3), c-Jun N-terminal kinase (JNK).