Targeting g4 dna structures with novel triphenylamine derivatives

Zusammenfassung

Apart from double-stranded DNA (dsDNA), a wide variety of alternative non-canonical DNA structures are well known, such as i-motifs, hairpins, triplexes or G-quadruplexes (G4). This latter sort of DNA is formed by stacks of two or more guanine tetrads, which are hold together through hydrogen bonds and electrostatic interactions. Within the human genome, G4 are mainly located in telomeres and promoter regions of several genes. Therefore, G4 may play an essential role in different biological processes, for instance transcription and expression of genetic information, as well as genomic stability. Consequently, G4 DNA has been identified as a potential target for antitumour therapy. In this context, several research groups have sought to design small molecules, known as G4 binders, with the aim to target G4 DNA in vivo. The typical molecular structure of potential G4 binders includes a π-delocalised core and positively charged side chains, enabling the establishment of π-π interactions with the G-tetrad, in addition to electrostatic interactions with the negatively charged phosphate backbone. The work presented deals with the design of novel polyamine-based compounds as G4 binders. The central core of the compounds and their side chains have been systematically modified, aiming to get insight into the influence of the molecular structure on the G4 affinity. Since the protonation state of the ligands modulates their interaction with DNA, their acid-base behaviour have been studied by potentiometric and spectroscopic techniques. In terms of G4 stabilisation, different G4 topologies have been assessed by fluorescence spectroscopy, Förster Resonance Energy Transfer (FRET) assays and circular dichroism. Some of the compounds have proved to be G4 selective fluorescent sensors, being able to distinguish between G4 DNA and dsDNA. Those compounds that have demonstrated to be potential G4 binders show a high positive charge at physiological pH, which may hamper their cell uptake and consequently their therapeutic effect. In order to overcome this limitation, they have been encapsulated inside liposome nanoparticles, which act as delivery vehicles. Furthermore, a specific liposome formulation including an aptamer has been designed in order to target the cell nucleus. The cytotoxicity of both compounds and liposomes nanoparticles has been studied in different tumour cell lines, proving the outstanding antitumour effect of the targeted nanoparticles. Finally, confocal fluorescence microscopy studies have confirmed the nuclear uptake of the targeted formulation.