Antiproliferative, proapoptotic and DNA methylation activity of sesqui- and diterpenoids in ovarian cancer cells

Ovarian cancer is a gynaecological malignancy associated with the highest level of death. Its resistance against anti-cancer drugs in recent time has exacerbated the problem. Hence, the search for novel drugs from plants used in traditional medicine is imperative. This study investigated the in vitro anti-ovarian cancer activities of Justicia insularis T. Anderson and mechanisms of action of its bioactive constituents, two plant-derived diterpenoids (andrographolide and triptolide) and four sesquiterpene lactones (dehydroleucodine, alantolactone, parthenolide and costunolide). The bioactive compounds from J. insularis were isolated and characterised using bioassay-guided fractionation, mass spectrometry and nuclear magnetic resonance spectroscopy. The antiproliferative activity of the sesqui- and diterpenoids was evaluated against human ovarian cancer cell lines (CIS-A2780, OVCAR- 8 and OVCAR-4) using Sulforhodamine B and trypan blue exclusion assays. Pro-apoptotic activity was evaluated using enzymatic assays and flow cytometry. Global DNA methylation and DNA methyltransferases (DNMTs) activity were measured using ELISA assay. Gene-specific CpG methylation and mRNA expression of panel of tumour suppressor genes were quantified using pyrosequencing and RT-qPCR respectively. The two major bioactive compounds in J. insularis were identified as diterpenoids, 16α/β-hydroxy-cleroda- 3,13(14) dien-15,16-olide and 16-oxocleroda-3,13(14) dien-15-oic acid. Sesqui- and diterpenoids showed strong growth inhibitory activity, decreased cell viability and induced apoptosis via intrinsic and extrinsic pathways. They possessed DNMT inhibitory activity, upregulated the expression of notable tumour suppressor genes. Furthermore, sesquiterpene lactones showed significant global and gene specific CpG methylation activity. In conclusion, sesqui- and diterpenoids possess strong anti-ovarian cancer activity and bear the potential to be developed as drug candidates for cancer treatment. Therefore, in vivo studies of their anti-ovarian cancer activity would be essential in the future