A topoisomerase II poison screen of ethnomedicinal Thai plants using a yeast cell-based assay Suphattra Sangmalee a , Areerat Laorpaksa b , Suchada Sukrong a,n a Department of Pharmacognosy and Pharmaceutical Botany, CU-Drug & Health Product Innovation Center, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok 10330, Thailand b Department of Biochemistry and Microbiology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok 10330, Thailand article info Available online 14 May 2012 Keywords: Anticancer DNA topoisomerase II Saccharomyces cerevisiae Thai traditional medicine Yeast cell-based assay abstract Ethnopharmacological relevance: The plants used in this study had previously been identified and used by Thai folk practitioners, who had knowledge of Thai traditional medicine, as alternative treatments for cancer. Investigation into the mechanism of the Topoisomerase II (Top2) poison of these plants may give rise to new drug leads for cancer treatment. Aim of the study: This study aimed to screen ethnomedicinal plants used in Thai traditional medicine for Top2 poison activity using a yeast cell-based assay and also to validate the traditional uses of these plants by examining the Top2 poison activity. Materials and methods: Thirty Thai medicinal plants were harvested and identified. Plant methanol extracts were prepared and screened in vitro using a yeast cell-based assay. Mutant yeast strains carrying the top2-1 allele, which encodes a temperature-sensitive topoisomerase, were used to establish the yeast spot test. Strains carrying this mutation grow normally at 25 1C and generally have a wild-type drug sensitivity. These yeast strains are able to grow at 30 1C, but the Top2 activity is greatly reduced at this temperature, which causes the strains to be highly drug resistant to anti-Top2 agents. Cell growth was measured by colony survival after exposure to various concentrations of extracts at different temperatures. Results: The extracts of six out of thirty ethnomedicinal plants, Curcuma longa, C. zedoaria, Derris scandens, Grangea maderaspatana, Stephania pierrei and S. suberosa, were found to have Top2 poison activity against the yeast cells. Conclusions: The yeast screening system confirmed the proposed anti-cancer mechanisms of plants used in Thai traditional medicine by traditional doctors. & 2012 Elsevier Ireland Ltd. All rights reserved. 1. Introduction The current trend in cancer treatment research is the devel- opment of drugs with defined molecular targets. The conceptual idea behind this approach is that knowledge of a drug’s mechan- ism of action provides a target for a specific site of action that is also specific for the type of cancer. These drugs aim to improve clinical results by targeting the cancer cells and minimally affecting healthy cells (Pezzuto, 1997). DNA topoisomerases have emerged as important targets for anticancer agents. These enzymes catalyse the relaxation of supercoiled DNA via a concerted mechanism for transient DNA cleavage and reannealing (Nitiss et al., 1992). DNA topoisome- rases can be divided into two classes: type I (Top1) enzymes, which transiently break and rejoin one DNA strand at a time, and type II (Top2) enzymes, which break and rejoin both strands of the double-stranded DNA (Reid et al., 1998). The activity of Top2 is different from that of Top1, as it is essential for the viability of growing cells. Top2 enzymes are also essential for DNA-mediated processes and for proliferation, which is characterised by a high level of Top2 expression (Robert and Larsen, 1998; McClendon and Osheroff, 2007). Top2 enzymes have been shown to be clinically important targets for cancer chemotherapy, and their inhibitors are central components of many therapeutic regimens (Reid et al., 1998). Many drugs target Top2 by trapping the covalent intermediate, termed the cleavage complex, converting the transient DNA cleavage into a stable DNA break and then covalently attaching the enzyme to the target DNA molecule (Liu, 1989; Nitiss et al., 1993; Nitiss, 2009). These drugs are often referred to as Top2 poisons because their anticancer activity is due to the trapping of Top2 rather than to the inhibition of Top2 activity. The agents that have been shown to inhibit Top2 by this mechanism include amsacrine, doxorubicin and etoposide (Meczes et al., 1997; Patel et al., 1997; Hammonds et al., 1998). Contents lists available at SciVerse ScienceDirect journal homepage: www.elsevier.com/locate/jep Journal of Ethnopharmacology 0378-8741/$ - see front matter & 2012 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jep.2012.05.013 n Corresponding author. Tel.: þ66 2 2188264; fax: þ66 2 2545195. E-mail address: suchada.su@chula.ac.th (S. Sukrong). Journal of Ethnopharmacology 142 (2012) 432–437