A new steroid derivative stabilizes g-quadruplexes and induces telomere uncapping in human tumor cells

Human telomeric DNA consists of tandem repeats of the sequence d( TTAGGG) with a 3 ' single- stranded extension ( the G- overhang). The stabilization of G- quadruplexes in the human telomeric sequence by small- molecule ligands inhibits the activity of telomerase and results in telomere uncapping, leading to senescence or apoptosis of tumor cells. Therefore, the search for new and selective G- quadruplex ligands is of considerable interest because a selective ligand might provide a telomere- targeted therapeutic approach to treatment of cancer. We have screened a bank of derivatives from natural and synthetic origin using a temperature fluorescence assay and have identified two related compounds that induce G- quadruplex stabilization: malouetine and steroid FG. These steroid derivatives have nonplanar and nonaromatic structures, different from currently known G- quadruplex ligands. Malouetine is a natural product isolated from the leaves of Malouetia bequaaertiana E. Woodson and is known for its curarizing and DNAbinding properties. Steroid FG, a funtumine derivative substituted with a guanylhydrazone moiety, interacted selectively with the telomeric G- quadruplex in vitro. This derivative induced senescence and telomere shortening of HT1080 tumor cells at submicromolar concentrations, corresponding to the phenotypic inactivation of telomerase activity. In addition, steroid FG induced a rapid degradation of the telomeric G- overhang and the formation of anaphase bridges, characteristics of telomere uncapping. Finally, the expression of protection of telomere 1 ( POT1) induced resistance to the growth effect of steroid FG. These results indicate that these steroid ligands represent a new class of telomere- targeted agents with potential as antitumor drugs.