The carbazole ligand 3 was synthesized, characterized and its binding interactions with human telomeric (22HT) G-quadruplex DNA in Na⁺ and K⁺-containing buffer were investigated by ultraviolet-visible (UV-Vis) spectrophotometry, fluorescence, circular dichroism (CD) spectroscopy, and DNA melting. The results showed that the studied carbazole ligand interacted and stabilized the intramolecular G-quadruplexes formed by the telomeric sequence in the presence of sodium and potassium ions. In the UV-Vis titration experiments a two-step complex formation between ligand and G-quadruplex was observed. Very low fluorescence intensity of the carbazole derivative in Tris HCl buffer in the presence of the NaCl or KCl increased significantly after addition of the 22HT G4 DNA. Binding stoichiometry of the ligand/G-quadruplex was investigated with absorbance-based Job plots. Carbazole ligand binds 22HT with about 2:1 stoichiometry in the presence of sodium and potassium ions. The binding mode appeared to be end-stacking with comparable binding constants of ~10(5) M(-1) as determined from UV-Vis and fluorescence titrations data. The carbazole ligand is able to induce formation of G4 structure of 22HT in the absence of salt, which was proved by CD spectroscopy and melting studies. The derivative of carbazole 3 shows significantly higher cytotoxicity against breast cancer cells then for non-tumorigenic breast epithelial cells. The cytotoxic activity of ligand seems to be not associated with telomerase inhibition.