Abstract
β-2-himachalen-6-ol (HC) is a major sesquiterpene that was isolated from the Lebanese Daucus carota ssp. carota and was shown to possess potent anticancer activity against various cancer cell lines. The aim of the present study was to explore the in vitro and in vivo anticancer activities of HC and to investigate the possible mechanisms of action involved. Lebanese Daucus carota spp. carota plant oil was extracted from dried mature umbels with methanol/acetone (1/1) and HC was purified using silica gel column chromatography to 92% purity as confirmed by GC-MS and NMR. The in vitro effect of HC on cell viability was tested against a panel of colon cancer cell lines (Caco-2, HT-29, SK-Co, LoVo, SW-1116 and T-84), a normal colon cell line CCD-33Co, the HaCaT-ras II-4 epidermal squamous carcinoma and the 4T1 breast cancer cell lines. HC displayed potent cytotoxic activity as measured by the WST-1 cell proliferation assay against all cancer cell lines with an IC50 ranging between 8-14.5μg/ml and an IC90 between 19-36μg/ml. No significant cytotoxic effect was observed on the normal colon cell line at the concentrations used. Upon treatment with HC, flow cytometry and western blot analyses showed similar results for both the SW1116 and HaCaT-ras II-4 cell lines, with an observed accumulation of cells in the sub-G1 apoptotic phase and decreased cell population in S and G2/M phases indicating cell cycle arrest at sub-G1phase. A decrease in the p-Erk, p- Akt, anti-apoptotic protein (Bcl-2) and an increase in pro-apoptotic proteins (p53, Bax, p21, cleaved Caspase-3 and PARP) was also observed. The in vivo anticancer effect of HC was investigated using DMH-induced colon, DMBA/TPA-induced skin carcinogenesis and 4T1 breast metastatic murine models of cancer. HC proved to be effective in reducing cancer incidence and size in all the experiments with a relatively low hepato-toxicity. The present study established that HC is a potent and relatively safe anticancer compound that induces apoptosis through the inhibition of the MAPK/Erk and PI3K/ Akt pathways.