G-quadruplexes (G4s) are secondary DNA structures which are held together by Hoogsten hydrogen bonding, with the further aid of cations (Na+ or K+). Recent publications have highlighted that G4s are over-represented in regulatory, nucleosome-depleted regions of the human genome which include promoter regions of genes involved in replication, oncogenes, introns, untranslated regions and at the 3’-end of telomeric DNA.1 The occurrence of G4-DNA in gene regulatory regions have made them an attractive target for transcriptional regulation in the oncology field, where a large number of G4-DNA stabilizing small molecules have been developed to trap quadruplexes in cellular DNA, down-regulate oncogene transcription, promote telomere disruption and induce growth arrest in cancer cells.2,3
In this work, a series of novel tri(carbazole)benzene derivatives were synthesized to target G4-DNA. These compounds possess C3-symmetry, an extended aromatic chromophore and positively charged side-chains, which are all important features for G4-targeting ligands. Our results indicate that for optimal G4 stabilization, a C3-alkyl side-chain length was necessary, where reducing the chain length by one CH2 unit, resulted in decreased G4-stabilization. Furthermore, all four of the highest stabilizing G4-ligands demonstrated high selectivity for G4-DNA over duplex DNA, which is a critical requirement in the design of G4-specific ligands given the ubiquitous nature of duplex DNA. Interestingly, among these four G4-ligands the highest G4 over duplex selectivity was observed for the ligand with the lower G4-stabilization highlighting that a high ligand-induced G4-stabilisation does not guarantee high G4 over duplex specificity. All four ligands were able to stabilize two or more quadruplexes, including the human telomeric quadruplex and the KRAS promoter quadruplex, demonstrating that these ligands able to have multiple targets which is an important feature given there are multiple genes that are dysregulated in cancer.