More is not always better: finding the right trade-off between affinity and selectivity of a G-quadruplex ligand

Abstract

Guanine-rich nucleic acid sequences can fold into four-stranded G-quadruplex (G4) structures. Despite growing evidence for their biological significance, considerable work still needs to be done to detail their cellular occurrence and functions. Herein, we describe an optimized core-extended naphthalene diimide (cex-NDI) to be exploited as a G4 light-up sensor. The sensing mechanism relies on the shift of the aggregate-monomer equilibrium towards the bright monomeric state upon G4 binding. In contrast with the majority of other ligands, this novel cex-NDI is able to discriminate among G4s with different topologies, with a remarkable fluorescent response for the parallel ones. We investigate this sensing by means of biophysical methods, comparing the lead compound to a non-selective analogue. We demonstrate that mitigating the affinity of the binding core for G4s results in an increased selectivity and sensitivity of the fluorescent response. This is achieved by replacing positively charged substituents with diethylene glycol (DEG) side chains. Remarkably, the limit of detection values obtained for parallel G4s are more than one order of magnitude lower than those of the parallel-selective ligand N-methyl mesoporphyrin IX (NMM). Interestingly, the classical fluorescent intercalator displacement (FID) assay failed to reveal binding of cex-NDI to G4 because of the presence a ternary complex (G4-TO-cex-NDI) revealed by electrospray-MS. Our study thus provides a rational basis to design or modify existent scaffolds to redirect the binding preference of G4 ligands.

A Light‐up Logic Platform for Selective Recognition of Parallel G‐Quadruplex Structures via Disaggregation‐Induced Emission

“…This result clearly supports the formation of a c‐ MYC sG4– CQ4 complex via the direct interaction of the CQ π‐surfaces with the hydrophobic G‐tetrad core of the c‐ MYC sG4 DNA structure. It is important to note, that the linear response of the fluorescence intensity of CQ4 as a function of c‐MYC sG4 concentration provided a limit of detection (LOD) of 83.5±4.2 n m awarding this probe as one of the most sensitive parallel G4‐binders reported so far (Figure , inset) . Moreover, the optical changes arising in the absorption and emission profiles resulted in excellent one‐photon brightness (defined as the product of ϵ max × Φ F ) of 3.9×10 4 m −1 cm −1 .…”

Selective parallel G-quadruplex recognition by a NIR-to-NIR two-photon squaraine

“…This approach is reported in the literature and has been exploited in the design of selective G4-ligands. 21 – 23 The molecular design of dicyanovinyl squaraine revealed a suitable choice in terms of affinity and selectivity with G4s. 18 Additionally, the donor–acceptor–donor (D–A–D) nature of the chromophores 24 appeared very promising for multi-photon absorption processes at long-infrared wavelengths.…”

Trifunctionalized Naphthalene Diimides and Dimeric Analogues as G-Quadruplex-Targeting Anticancer Agents Selected by Affinity Chromatography

“…This design, with respect to the previously synthesized dimeric NDIs containing only alkylamines in their side chains [21], allows for the reduction of the unspecific interactions toward duplex DNA, typically resulting from the electrostatic attractions between the backbone phosphates and the protonated ligands. In this frame, the new NDIs should lose some affinity degrees toward the G4 targets but, in turn, gain enhanced G4 vs. duplex selectivity [23]. The second structural modification here introduced is related to the side chain (for the monomeric NDIs) or the linker connecting the two NDI units (for the dimeric NDIs).…”