Phenethyl caffeate benzo[kl]xanthene lignan with DNA interacting properties induces DNA damage and apoptosis in colon cancer cells

摘要

Aims: Phenethyl caffeate benzoxanthene lignan (PCBL) is a synthetic compound with DNA interacting, antiangiogenic, antiproliferative and tumor cell death inducing abilities. Though PCBL exhibits the qualities of a prospective antitumor agent, the basic mechanism of PCBL induced cell death remains unknown. This study aims to analyze the molecular mechanisms of PCBL induced cell death in tumor cells to further substantiate its antitumor abilities. Main methods: MTT assay was used for finding cell proliferation inhibition, flow cytometric analysis for the detection of cell cycle arrest, comet assay for DNA break detection and immunofluorescence for analyzing H2AX phosphorylation. Western blot analysis was used to detect the activation of different proteins related to DNA damage response and apoptosis. Key findings: PCBL inhibited proliferation of WiDr cells more efficiently than its analog, MCBL. Comet analysis of PCBL treated WiDr cells and activity of various DNA damage response proteins such as γ-H2AX, BRCA1, ATR and Chk1 in PCBL treated cells demonstrated the DNA damaging property of PCBL. Effector molecules of apoptosis such as caspase-3, caspase-7 and caspase-9 were found activated along with PARP cleavage in PCBL treated cells, suggesting apoptosis as the main mode of cell death. PCBL induced cell death was found associated with the activation of MAPK signaling. Inhibition of ERK, one of the MAPKs, by U0126 improved the apoptosis inducing ability of PCBL. Significance: In vitro findings suggest that PCBL works by initiating DNA damage and inducing apoptosis in cancer cells and thus could be considered for further preclinical studies.