Chapter 18 - Anticancer effects of the active fraction from clove in vitro and in vivo

Abstract

Previous studies from our laboratory have shown that the extract of clove possesses a potent anticancer effect both in vitro and in vivo. In the present study, we further evaluated the effect of active fraction extracted and isolated from clove (AFC) on induction of cellular apoptosis in human colorectal cancer HCT-116 cells by morphological observation, flow cytometry, and Western blotting analysis. The results showed that AFC was able to induce typical apoptosis in HCT-116 cells. AFC also induces autophagy by increases of punctuating microtubule-associated protein 1A/1B-light chain 3 (LC3) dots, the levels of LC3-II, and Beclin-1 protein. Moreover, autophagy inhibitors 3-MA, and bafilomycin A1 (BA) potentiate the pro-apoptotic activity of AFC in HCT-116 cells. AFC also inhibits the phosphorylation of PI3K/Akt/mTOR signaling pathway. These studies provide evidence for the understanding of the anticancer mechanism of clove and the scientific rationale for AFC to be further developed as a promising novel anticancer agent for the treatment of colorectal cancer clinically. Cloves (Syzygium aromaticum), a traditional Chinese medicinal herb, display broad biological activity. In the present study, the aqueous extract of clove (AEC) was prepared and its anticancer effects were studied. MTS analysis revealed that AEC was able to inhibit cancer cell growth in vitro on several cancer cell lines; the IC50 is around 150μg/mL for human pancreatic ASPC-1 and human colon HT-29 cancer cells. Treatment of the cancer cells with AEC also diminished the colony formation significantly in both human pancreatic ASPC-1 cancer cells and human colon HT-29 cancer cells. In vivo study revealed that AEC inhibited the tumor growth significantly in the HT-29 xenograft mice model. Transmission electron microscope (TEM), flow cytometry assay and fluorescence microscope analysis confirmed that AEC is capable of inducing cell autophagy. Further study showed that the AMPK/ULK pathway plays an important role in AEC-induced autophagy in cancer cells. Analysis of AEC components was performed by liquid chromatograph-mass spectrometer (LC-MS) approach, and more than nine constitutes were identified in the AEC fraction. The study provides evidence that AEC has the potential to be developed as a novel anticancer agent or as an adjuvant in cancer chemotherapy.