Titanium(IV) targets phosphoesters on nucleotides: implications for the mechanism of action of the anticancer drug titanocene dichloride

Abstract.

Reactions between the anticancer drug titanocene dichloride (Cp₂TiCl₂) and various nucleotides and their constituents in aqueous solution or N,N-dimethylformamide (DMF) have been investigated by ¹H and ³¹P NMR spectroscopy and in the solid state by IR spectroscopy. In aqueous solution over the pH* (pH meter reading in D₂O) range 2.3–6.5, CMP forms one new species with Ti(IV) bound only to the phosphate group. In acidic media at pH*<4.6, three species containing titanocene bound to the phosphate group of dGMP, AMP, dTMP and UMP are formed rapidly. The bases also appear to influence titanocene binding. Only one of these Ti(IV)-bound species can be detected in the pH* range of 4.6–6.5 in each case. The order of reactivity towards Cp₂TiCl₂(aq) at pH* ca. 3 is GMP>TMP≈AMP>CMP. At pH*>7.0, hydrolysis of Cp₂TiCl₂ predominated and little reaction with the nucleotides was observed. Binding of deoxyribose 5′-phosphate and 4-nitrophenyl phosphate to Cp₂TiCl₂(aq) via their phosphate groups was detected by ³¹P NMR spectroscopy, but no reaction between Cp₂TiCl₂(aq) and deoxyguanosine, 9-ethylguanine or deoxy-D-ribose was observed in aqueous solution. The nucleoside phosphodiesters 3′,5′-cyclic GMP and 2′,3′-cyclic CMP did not react with Cp₂TiCl₂(aq) in aqueous solution; however, in the less polar solvent DMF, 3′,5′-cyclic GMP coordination to {Cp₂Ti}²⁺ via its phosphodiester group was readily observed. Binding of titanocene to the phosphodiester group of the dinucleotide GpC was also observed in DMF by ³¹P NMR. The nucleoside triphosphates ATP and GTP reacted more extensively with Cp₂TiCl₂(aq) than their monophosphates; complexes with bound phosphate groups were formed in acidic media and to a lesser extent at neutral pH. Cleavage of phosphate bonds in ATP (and GTP) by Cp₂TiCl₂(aq) to form inorganic phosphate, AMP (or GMP) and ADP (or GDP) was observed in aqueous solutions. In addition, titanocene binding to ATP was not inhibited by Mg(II), but the ternary complex titanocene-ATP-Mg appeared to form. These reactions contrast markedly with those of the drug cisplatin, which binds predominantly to the base nitrogen atoms of nucleotides and only weakly to the phosphate groups. The high affinity of Ti(IV) for phosphate groups may be important for its biological activity.