Moscatilin induces apoptosis in human colorectal cancer cells: a crucial role of c-Jun NH2-terminal protein kinase activation caused by tubulin depolymerization and DNA damage

Purpose: To study the effect of moscatilin (purified from the stem of orchid Dendrobrium loddigesii) on the proliferation of human colorectal Cancer HCT-116 cells in vitro and in vivo.

Experimental design: The growth inhibition of moscatilin was screened on several human Cancer cell lines. The effect of moscatilin on tubulin was detected in vitro. Following moscatilin treatment on HCT-116 cells, c-Jun NH(2)-terminal protein kinase (JNK) and Caspase activation was studied by Western blot analysis, and DNA damage was done by Comet assay. Specific JNK Inhibitor SP600125 was cotreated to reverse moscatilin-induced Apoptosis. Tumor growth inhibition of moscatilin was done on HCT-116 xenograft models.

Results: Moscatilin induced a time-dependent arrest of the cell cycle at G(2)-M, with an increase of cells at sub-G(1). Moscatilin inhibited tubulin polymerization, suggesting that it might bind to tubulins. Moscatilin also induced the phosphorylation of JNK1/2. SP600125 significantly inhibited the activation of caspase-9 and Caspase-3 and the subsequent moscatilin-induced Apoptosis. The data suggest that JNK activation may contribute to moscatilin-mediated Apoptosis signaling. A parallel experiment showed that SP600125 significantly inhibits Taxol- and vincristine-induced HCT-116 cell Apoptosis. This suggests that the JNK activation may be a common mechanism for tubulin-binding agents. Moreover, moscatilin induces DNA damage, phosphorylation of H2AX and p53, and up-regulation of p21. Our HCT-116 xenograft models show the in vivo efficacy of moscatilin.

Conclusions: In summary, our results suggest that moscatilin induces Apoptosis of colorectal HCT-116 cells via tubulin depolymerization and DNA damage stress and that this leads to the activation of JNK and mitochondria-involved intrinsic Apoptosis pathway.