“Synthesis and biological evaluation of thymoquinone derivatives and ON01910 derivatives as anticancer agents”


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Osama Chahrour2, Liu Xiangrui1, Lam Franki1, Ashraf Abdallah1, Carol Midgley3, Wang Shudong1
1University of Nottingham, Nottingham, United Kingdom,2University of Damascus, Damascus, Syrian Arab Republic,3The Open University, Milton Keynes, United Kingdom

Background

Polo-like kinase 1 (Plk1) is overexpressed in many human cancers, and considered as poor prognosis marker for cancer patients. Plk1 Polo box domain (PBD) is well validated target for the development of selective non ATP competitive Plk1 inhibitors for cancer therapy. Thymoquinone, The bioactive constituent of Nigella sativa volatile oil, and ON01910, a small synthetic molecule, are reported to inhibit Plk1 activity by targeting PBD.

Method

Thymoquinone, ON01910, and nineteen derivatives were synthesised. In vitro anti-proliferative effect was assessed by MTT assays against six human cell lines; HCT116, MCF7, MDA468, MDA231, M74D, and MRC5. Mitotic index, cell cycle, P53 stabilization, and caspase activation assays of selected compounds were assessed in one or more of Hela, MCF7, and HCT116 cells. Nocodazole synchronised-MCF7 lysate was used for western blotting and unsynchronized-Hela for cell staining to determine subcellular structural defects.

Results

Ligand based drug design resulted in the identification of the molecule OC2-23 which is ten times more active than the lead compound ON01910 in MTT assays. The biological evaluation showed that ON01910 blocked cells at G2/M phase, increased mitotic index and cellular P53, enhanced caspase3 activity, and caused the formation of aberrant multipolar mitotic spindle which is consistent with Plk1 role in centrosome maturation and bipolar spindle stabilisation. In contrast, thymoquinone induced a slight increase in the mitotic index, and caused cells to round up, bleb, and dye. Western blotting detecting Cdc25c, Plk1 substrate at the mitotic entry, suggested that Cdc25c phosphorylation is reduced by thymoquinone and not by ON01910. 

Conclusion

Structure activity relationship of thymoquinone and ON01910 was analysed and used in synthesising more active anti cancer agents. ON01910 caused the formation of multipolar spindle which is consistent with the effects of Plk1-PBD inhibition after the mitotic entry. Thymoquinone reduced the phosphorylation of Cdc25c which indicates Plk1-PBD inhibition at the mitotic entry.