G-quadruplexes (G4) are non-canonical DNA secondary structures that form in guanine-rich nucleic acid sequences. They are composed of stacked guanine tetrads arising from Hoogsteen hydrogen bonding between four guanine bases and stabilised by a central monovalent cation. G4s are found throughout the genomes of many organisms, where they regulate key biological processes, including gene regulation and genome integrity. Small molecule ligands that bind G4s have been shown to modulate cellular functions such as gene expression, highlighting their emerging potential in therapeutic strategies. To date, the selective binding of small molecules to G4s remains a significant challenge, particularly due to competing interactions with double-helical DNA.
This work aims to develop a library of novel Hoechst-derived compounds, exploiting their strong nucleic acid affinity and tunable scaffold to achieve selective G4 binding. Circular dichroism spectroscopy, UV–Vis titrations, and Förster resonance energy transfer (FRET) assays were used to investigate the binding interactions of a series of Hoechst-derived compounds with double-stranded DNA and G4-forming sequences. These data provided insights into binding affinity, selectivity, and conformational effects, elucidating the molecular basis of G4 recognition. Further structure-activity relationship studies are expected to advance the rational design of small molecules capable of selective G4 targeting and modulation of their biological activity.