Abstract
Evodiamine (EVO), an alkaloidal compound isolated from Evodia rutaecarpa (Juss.), has been reported to affect many physiological functions. Topoisomerase inhibitors have been developed in a variety of clinical applications. In the present study, we report the topoisomerase I (TopI) inhibitory activity of EVO, which may have properties that lead to improved therapeutic benefits. EVO is able to inhibit supercoiled plasmid DNA relaxation catalyzed by TopI. Upon treatment 0~10 μM EVO TopI was depleted in MCF-7 breast cancer cells in a concentration-dependent and time-dependent manner in 0~120 min. A K-SDS precipitation assay was performed to measure the extent of Top I-trapped chromosomal DNA. The ability of EVO to cause the formation of a TopI-DNA complex increased in a concentration-dependent manner, in that the DNA trapped increased by 24.2% in cells treated with 30 μM. The results suggest that EVO inhibits TopI by stabilizing the enzyme and DNA covalent complex.
Highlights
DNA topoisomerase enzymes regulate the topological state of DNA that is crucial for initiation and elongation during DNA synthesis
A phenolic oxygen attacks a DNA phosphodiester bond, forming an intermediate in which the 3′ end of the broken strand is covalently attached by an O4-phosphodiester bond to topoisomerase I (TopI) tyrosine
The cytotoxic abilities of EVO (Figure 1) and CPT were tested against human breast MCF-7 carcinoma cells, which express high levels of TopI
Summary
DNA topoisomerase enzymes regulate the topological state of DNA that is crucial for initiation and elongation during DNA synthesis. A phenolic oxygen attacks a DNA phosphodiester bond, forming an intermediate in which the 3′ end of the broken strand is covalently attached by an O4-phosphodiester bond to TopI tyrosine. The religation step consists of transesterification involving a nucleophilic attack by the hydroxyl oxygen at the 5′ end of the broken strand. Some TopI- and TopII-targeted drugs are reported to stabilize the covalent topoisomerase-DNA complex, thereby preventing religation [2]. TopI-DNA complexes can be trapped and purified because the enzymatic religation is no longer functional. The inhibitor-trapped TopI-cleavable complex triggers replication fork arrest and breakage to generate a DNA break that is responsible for the G2/M arrest and activation of DNA damage signals including nuclear factor κB activation, p53 upregulation, Chk phosphorylation, and ATM/ATR activation [3]. We report the TopI-DNA trapping activity by EVO, which suggests beneficial effects of EVO for the development of a variety of therapeutic applications
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