A Minimalistic Coumarin Turn-On Probe for Selective Recognition of Parallel G-Quadruplex DNA Structures

Abstract

G-quadruplex (G4) DNA structures are widespread in the human genome and are implicated in biologically important processes such as telomere maintenance, gene regulation, and DNA replication. Guanine-rich sequences with potential to form G4 structures are prevalent in the promoter regions of oncogenes, and G4 sites are now considered as attractive targets for anticancer therapies. However, there are very few reports of small “druglike” optical G4 reporters that are easily accessible through one-step synthesis and that are capable of discriminating between different G4 topologies. Here, we present a small water-soluble light-up fluorescent probe that features a minimalistic amidinocoumarin-based molecular scaffold that selectively targets parallel G4 structures over antiparallel and non-G4 structures. We showed that this biocompatible ligand is able to selectively stabilize the G4 template resulting in slower DNA synthesis. By tracking individual DNA molecules, we demonstrated that the G4-stabilizing ligand perturbs DNA replication in cancer cells, resulting in decreased cell viability. Moreover, the fast-cellular entry of the probe enabled detection of nucleolar G4 structures in living cells. Finally, insights gained from the structure–activity relationships of the probe suggest the basis for the recognition of parallel G4s, opening up new avenues for the design of new biocompatible G4-specific small molecules for G4-driven theranostic applications.

Self‐Assembled Peptidyl Aggregates for the Fluorogenic Recognition of Mitochondrial DNA G‐Quadruplexes

“…Despite significant advances in this field, the design of fluorescent "light-up" probes capable of selectively recognizing certain types of G4 DNAs over either single-stranded or duplex DNAs is highly challenging. [44][45][46][47][48] Moreover, most reported G4 fluorescent probes target nuclear G4s, whereas selective and sensitive probes for G4s in mitochondria are very rare (Table S7). [41,[49][50][51][52][53][54] Herein, we report a self-assembled peptidyl fluorescent probe (CV2, Figure 1b) that shows an excellent discrimination for G4 DNAs over single-stranded or duplex DNA.…”

“…Despite significant advances in this field, the design of fluorescent “light‐up” probes capable of selectively recognizing certain types of G4 DNAs over either single‐stranded or duplex DNAs is highly challenging [44–48] . Moreover, most reported G4 fluorescent probes target nuclear G4s, whereas selective and sensitive probes for G4s in mitochondria are very rare (Table S7) [41, 49–54] …”

Parallel G-Quadruplex DNA Structures from Nuclear and Mitochondrial Genomes Trigger Emission Enhancement in a Nonfluorescent Nano-aggregated Fluorine–Boron-Based Dye

“…In particular, noncanonical four-stranded DNA structures such as intercalated motifs (i-DNAs) and G4s have received increasing scientific attention, not only for being implicated in key biological processes, − but also for their predictable and controllable properties. Indeed, such structures can afford complex nanometer scale systems suited for molecular computing, , transport, , amplification of chirality, and biosensing − applications. While i-DNAs are predominantly formed under acidic conditions (pH < 7), − G4s are thermodynamically stable at physiological pH. , G4s are composed of stacked G-tetrads octa-coordinated by a central metal ion (usually K + ) and further stabilized by intraquartet hydrogen bonds. , Owing to their polynucleotide constitution and dynamic conformation, G4s show high polymorphism in solution.…”

“…The coumarin core can be combined with, for example, quinazolines, benzothiazoles, benzoimidazoles, and amidines to give compounds that combine fluorescent properties with G4 binding and stabilizing properties. , Along the same line, dipyrro-boradiazaindacenes (BODIPY) dyes have great potential in the field of fluorescent sensing and bioimaging because of their excellent photophysical properties, relatively high photostability, and neutral total charge. , Unfortunately, with some exceptions, − the strong intermolecular interactions exerted by the planar π-conjugated BODIPY-units, in aqueous solution, strongly quench the fluorescence intensity through activation of fast nonradiative decay pathways, narrowing their practical applications . Here, by building on this knowledge, we designed a benzimidazole–iminocoumarin fused with a fluorine–boron unit that resembles the BODIPY core to generate a self-assembled dye, hereafter 4b , capable of recovering its intense fluorescence in response to binding to parallel G4s through the recognition-induced disassembly of the nonemissive aggregates (Figure ).…”

Light-Induced Modulation of Chiral Functions in G-Quadruplex–Photochrome Systems

“…A set of different experiments by using a G4 specific antibody (BG4), fluorescent reporters, and G4 ChIP-seq showed an increased level of G4 structures in cancer cells compared to normal cells. ,− Furthermore, these globular non-B DNA structures have been also shown to play pivotal roles in regulating biological processes such as replication, transcription, translation, and splicing. , Therefore, they are recognized as potential therapeutic targets for small-molecule drugs in a range of disease classes including cancer . Although a plethora of molecules based on different recognition processes − have been reported to target G4 motifs, the development of light-responsive G4 ligands has received scant attention to date. , Molecular photoswitches based on an azobenzene unit, , stilbene-based and dithienylethene-based ligands, a (photo)­caged G4-binder, and a photochemically generated π-extended dicationic compound constitute rare examples of such molecules. Moreover, the design and implementation of molecules with tunable structural chirality to control G4 binding/function is still in its infancy. ,− …”