Abstract
Nucleic acid sequences containing repeats of short G-tracts can fold into four-stranded structures termed G-quadruplexes. Tertiary DNA structures of this type may be formed in telomeric DNA repeats as well as in regulatory sequences of protooncogenes and genes involved with proliferation such as c-myc, c-kit, KRAS, PDGF-A and Bcl-2. Small molecules that can selectively bind and stabilize G-quadruplex structures are of increasing interest as potentially novel anticancer agents since they can, for example, promote selective telomere destabilization or down-regulate oncogene expression in cancer cells. The majority of quadruplex-binding molecules studied to date possess planar aromatic chromophores such as acridines, anthraquinones, indoloquinolines, macrocyclic oligoamides and porphyrins. We have discovered a novel class of biaryl polyamides which show high selectively towards G-quadruplexes compared to duplex DNA. These molecules also have significant biological activity in several cancer cell lines. Molecular modeling studies suggest that these biaryl polyamides form planar U-shaped structures that match the surface area dimensions of a terminal G-quartet in quadruplex structures but cannot readily be accommodated within the minor groove of duplex DNA. The molecules described are part of an 84-member solution-phase library, the design of which allowed positioning of the biaryl units at either end of the polyamide core in order to introduce ligand diversity. Assessment of G-quadruplex DNA stabilization was carried out with a FRET-based DNA melting assay using three different quadruplex sequences: human telomeric G-quadruplex (h-telo), two c-kit promoter G-quadruplexes (c-kit1 and c-kit2), and a hairpin DNA sequence with a duplex structure as control. Six biaryl polyamides were selected from the initial screen and these were evaluated further using different biophysical techniques. All six compounds showed a high level of selectivity towards G-quadruplex compared to duplex DNA structure. At 1 to 10 µM concentrations they had 20- to 50-fold selectivity for G-quadruplex compared to duplex DNA. The inability of these molecules to bind to duplex DNA was confirmed by DNAseI footprinting experiments against Hex A and Hex B sequences in which they failed to provide any footprints at concentrations >50 µM. A short-term growth inhibitory experiment against four different cancer cell lines, MCF-7, A549, HT29 and GIST882, gave low micromolar to sub-micromolar IC50 values. CD titrations of the six most promising ligands with h-telo, c-kit1 and c-kit2 quadruplex-forming DNA sequences showed concentration-dependent enhancement of major CD signals, and in some cases the ligands induced the folding of a particular quadruplex structure.
Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 5354.
100th AACR Annual Meeting-- Apr 18-22, 2009; Denver, CO