Quadruplex ligands

Mergny JL et al. (2001) Telomerase inhibitors based on quadruplex ligands selected by a fluorescence assay. Proc Natl Acad Sci USA 98(6) 3062-3067... 

Haider SM, Parkinson GN, Neidle S (2003) Structime of a G-quadruplex-ligand complex. J Mol Biol 326(1) 117-125... 

This relatively small DCL yielded the first peptidic ligand for G-quadruplex DNA, providing a lead compound for the further development of novel G-quadruplex ligands with possible therapeutic utility. 

Nielsen, M. C. Ulven, T. Selective extraction of G-quadruplex ligands from a rationally designed scaffold-based dynamic combinatorial library. Chem. Ear. J. 2008,14, 9487-9490. 

This duplex-quadruplex competition is of special interest in the case of gene promoters. As discussed in the chapter by L. Hurley, guanine rich sequences are present in the promoter region of key oncogenes, and one can observe an induction of duplex to G-quadruplex transition in the presence of a quadruplex ligand and/or KCl. ... 

Data on the kinetics of bimolecular quadruplexes are scarce. Attachment of a long single-stranded tail to an oligonucleotide bearing two guanine blocks destabilizes and delays bimolecular quadruplex formation. This property has been used to determine the effect of quadruplex ligands on quadruplex forma-tion " " (see below). The second order rate constant for the formation of the bimolecular structure was estimated to be 3 x 10 M s in the absence of ligand. 

An important point concerning small quadruplex ligands would be to demonstrate that these molecules may not only bind to preformed quadruplexes, but also induce the formation of the multistranded structures starting from single strands. Kinetic analyses are well suited to answer to this question, as these modes of interactions should be reflected by differences in dissociation and association rates of the quadruplex, respectively. 

Figure 6 A small G-quadruplex ligand accelerates quadruplex formation, (a) Gel electrophoresis experiment. 100 [iM of TG3T was incubated at 4°C for 4.5 h in a pH 7.2 10 mM lithium cacodylate buffer supplemented with 110 mM KCl, in the presence of various concentrations (0-100 tM) of a specific quadruplex ligand, 360A. Samples were then loaded on a 20% acrylamide TBE IX gel supplemented with 20 mM KCl. Nucleic acids were revealed by UV-shadow using a 254 nm UV lamp. A retarded band reveals the formation of a quadruplex the presence of the quadruplex ligand induces a further retardation. (b) Isothermal (4°C) absorbance measurements at 295 nm of 31 vM TG4T in the absence (circle, left Y-scale) or presence (squares, right Y-square) of 10 iM 360A in a pH 7.2 10 mM lithium cacodylate buffer supplemented with 110 mM NaCl...

Several studies have shown that structural information derived from the modelling of quadruplex-ligand interactions can lead to new and more effective... 

The objective of this chapter is to present the latest view on the cellular mechanism(s) of action of G-quadruplex ligands and to discuss their potential use in therapy. 

Figure 2 Chemical formula of some G-quadruplex ligands...

G-quadruplex ligands behave as telomerase inhibitors. (A) Long-term treatment of A549 human cell line with a sub-toxic concentration of 12459 (0.04 i.M) leads to the cell-culture growth arrest after several weeks. (B) TRF decrease induced by 12459 in A549 cells. (C) Senescence induced by 12459, revealed by SA-b galactosidase activity at PD 50... 

Eor BRACO-19, an important decrease in the nuclear hTERT, together with the formation of cytoplasmic hTERT bound to ubiquitin may explain the telomerase activity down-regulation. Other antitumor agents have been also found to down-regulate telomerase activity and hTERT expression has been reported as a marker of cell proliferation. Thus, the simplest explanation for the effect of the G-quadruplex ligands to down-regulate telomerase activity is related to their antiproliferative activity. 

It was also observed earlier that G-quadruplex ligands indueed more rapid effects on cell growth than that initially expected for telomerase inhibition alone. Apoptosis and short-term response were observed with triazine derivatives (12459, 115405), telomestatin, and more recently with the pyridine dicarboxamide derivatives (307A, 360A). " Telomestatin induced the activation of ATM and Chk2 that corresponded to an activation of the DNA damage response. 12459 induced apoptosis through the mitochondrial pathway and also provoked the early activation of P53. ... 

Short-term and massive apoptosis were also observed from the interference of the telomere-capping function of telomerase when hTERT or hTR are modified by mutations.The observation that BRACO-19 causes chromosome end-to-end fusions associated with the appearance of pl6-associated senescence led to the proposal that G-quadruplex ligands mostly act to disrupt the telomere structure.Such telomeric dysfunction was also observed in cell lines treated with RHSP4 or with 307A and in cell lines resistant to 12459 with typical images of telophase bridges" (Figure 4). 

The evidence that the antiproliferative effect of G-quadruplex ligands is independent of the presence of telomerase activity also comes from two series of experiments. 

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