Platinum(II) complexes bearing acetylide ligands containing nucleobase motifs are prepared and their impact on human topoisomerase II (TopoII) is evaluated. Both platinum( II) complexes [Pt(C∧N∧N)- (C≡CCHR)] (1a-c) and [Pt(tButerpy) (C≡CCHR)]+ (2a-c) (C∧N∧N=6-phenyl-2,2′- bipyridyl, tButerpy=4,4′,4″-tri-tert-butyl-2,2′: 6′,2″-terpyridyl, and R=(a) adenine, (b) thymine, and (c) 2-amino-6-chloropurine) are stable in aqueous solutions for 48 hours at room temperature. The binding constants (K) for the platinum(II) complexes towards calf thymus DNA are in the order of 10 dmmol as estimated by using UV/Vis absorption spectroscopy. Of the complexes examined, only complexes 1a-c are found to behave as intercalators. Both complexes 1a-c and 2a-c inhibit TopoII-induced relaxation of supercoiled DNA, while 2c is the most potent TopoII inhibitors among the tested compounds. Inhibition of DNA relaxation is detected at nanomolar concentrations of 2c. All of the platinum(II) complexes are cytotoxic to human cancer cells with IC values of 0.5-13.7 μm, while they are less toxic against normal cells CCD-19 Lu. © 2010 Wiley-VCH Verlag GmbH&Co. KGaA, Weinheim.