Proteomics analysis of bt-474 and skov-3 cancer cell lines treated with purified compounds from bee pollen and cerumen

Abstract

Breast and gynaecological cancers are the main causes of global cancer death in females. So far, several compounds have been identified from traditional medicine components and natural products that exhibited anticancer activity. Previously, α-mangostin (α-MG) and apigenin (APG) had been extracted and purified from cerumen of Tetragonula laeviceps and bee pollen of Apis mellifera respectively. Preliminary studies reported that these compounds express antiproliferative activity in several cancer cell lines. In this study, α-MG and APG were investigated for antiproliferation effect in breast cancer cell line BT-474 and ovarian adenocarcinoma cell line SKOV-3 using MTT assay, fluorescent staining coupled with flow cytometry analysis, caspase activity assay, and quantitative real-time PCR of interest genes. From BT-474 cell line result, both compounds caused necrotic death by induction of inflammation, as observation of COX2 gene upregulation. In SKOV-3 cell line, APG induced apoptosis via an intrinsic pathway while α-MG led to necrosis that was associated with upregulation of COX2 gene expression. Both compounds arrested cell progression at G2/M phase. In addition, the half proximal inhibition concentration (IC50) of both compounds in normal cell lines was higher than cancer cell lines representing less cytotoxicity on normal cells. Regarding the antiproliferation effect of APG in SKOV-3, it is interesting how APG regulated cellular mechanisms to suppress cell proliferation. To clarify this question, 11-plex TMT labelling phosphoproteomics was performed at an early response time to observe the global changes of phosphoproteins after APG exposure. Gene set enrichment analysis appeared that APG treatment altered regulation of transcription via epigenetics, histone modification, and organisation associated with demethylation, and activated various signalling pathways especially signalling through mitogen-activated protein kinase (MAPK). APG inhibited cyclin-dependent kinase 1, 2, and 4 (CDK1, CDK2, and CDK4) activities, and activated stress response and cell survival signalling from NetworKIN and Kinase set enrichment analysis (KSEA). Notably, inhibition of pyruvate dehydrogenase complex activity by enhancing activities of pyruvate dehydrogenase kinase (PDK1 and PDK2) presumed that conversion of energy metabolism and aerobic glycolysis occurred in SKOV-3 cells after exposure with APG. Taken together, α-MG and APG exhibit antiproliferative activity on a different mechanism and knowledge from this study provide a better understanding of cellular response to the compounds that could become useful as a therapeutic or co-treatment agent for cancer medication.