Symmetric and dissymmetric carbohydrate-phenyl ditriazole derivatives as DNA G-quadruplex ligands: Synthesis, biophysical studies and antiproliferative activity.

Here we present a novel G4-binding family of compounds based on a central core of phenyl ditriazole (PDTZ) modified with carbohydrates and phenyl pyrrolidinyl side-chains. Their synthesis was achieved using controlled click chemistry conditions to obtain both, symmetric and dissymmetric carb-PDTZ derivatives without any intermediate protecting steps through an optimized methodology. Binding of the new carb-PDTZ to a variety of G-quadruplex motifs was examined using different biophysical techniques. The symmetric carb-PDTZ derivatives were not able to stabilize G4, but the dissymmetric ones (containing one sugar and one phenyl pyrrolidinyl side-chain) did. Interestingly, the dissymmetric carb-PDTZ derivatives showed much higher G4 vs duplex DNA selectivity than the control compound PDTZ 1, which contains two phenyl pyrrodilinyl side-chains and no carbohydrates. Their potential antitumoral activity was also investigated by in vitro cytotoxicity measurements on different cancerous cell lines. All carb-PDTZ derivatives showed higher IC50 values than the control PDTZ 1, probably due to the lack of compound stability of some derivatives and to lower cellular uptake.

G-Quadruplex Identification in the Genome of Protozoan Parasites Points to Naphthalene Diimide Ligands as New Antiparasitic Agents.

G-quadruplexes (G4) are DNA secondary structures that take part in the regulation of gene expression. Putative G4 forming sequences (PQS) have been reported in mammals, yeast, bacteria, and viruses. Here, we present PQS searches on the genomes of T. brucei, L. major, and P. falciparum. We found telomeric sequences and new PQS motifs. Biophysical experiments showed that EBR1, a 29 nucleotide long highly repeated PQS in T. brucei, forms a stable G4 structure. G4 ligands based on carbohydrate conjugated naphthalene diimides (carb-NDIs) that bind G4's including hTel could bind EBR1 with selectivity versus dsDNA. These ligands showed important antiparasitic activity. IC50 values were in the nanomolar range against T. brucei with high selectivity against MRC-5 human cells. Confocal microscopy confirmed these ligands localize in the nucleus and kinetoplast of T. brucei suggesting they can reach their potential G4 targets. Cytotoxicity and zebrafish toxicity studies revealed sugar conjugation reduces intrinsic toxicity of NDIs.

Synthesis, Binding Properties, and Differences in Cell Uptake of†G-Quadruplex Ligands Based on Carbohydrate Naphthalene Diimide Conjugates.

The G-quadruplexes (G4s) are currently being explored as therapeutic targets in cancer and other pathologies. Six carbohydrate naphthalene diimide conjugates (carb-NDIs) have been synthesized as G4 ligands to investigate their potential selectivity in G4 binding and cell penetration. Carb-NDIs have shown certain selectivity for G4 structures against DNA duplexes, but different sugar moieties do not induce a preference for a specific G4 topology. Interestingly, when monosaccharides were attached through a short ethylene linker to the NDI scaffold, their cellular uptake was two- to threefold more efficient than that when the sugar was directly attached through its anomeric position. Moreover, a correlation between more efficient cell uptake of these carb-NDIs and their higher toxicity in cancerous cell lines has been observed. Carb-NDIs seem to be mainly translocated into cancer cells through glucose transporters (GLUT), of which GLUT4 plays a major role.

Synthesis, RNAi activity and nuclease-resistant properties of apolar carbohydrates siRNA conjugates.

Oligoribonucleotide conjugates carrying apolar carbohydrates at the 5'-end and the corresponding siRNA duplexes have been prepared using phosphoramidite chemistry. All the carbohydrate-siRNA derivatives were compatible with RNA interference machinery if transfected with oligofectamine. In the absence of a transfection agent, some of them exerted certain reduction of gene expression. Double-tailed permethylated glucose conjugated to siRNA through a long spacer inhibited gene expression up to 26% compared to the scrambled duplex. Such modifications contribute positively to the stability of oligoribonucleotides against 5'-exonuclease degradation.