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.