Highly water-soluble ruthenium(II) terpyridine coordination compounds form stable adducts with blood-borne metal transporting proteins

Abstract

Three coordination compounds of ruthenium(II), belonging to a recently synthesised series of water-soluble compounds of general formula mer-[Ru(L3)(N-N)Cl]Cl, where L3 = 4'-chloro-2,2':6',2″-terpyridine (Cl-tpy), N-N = ethylenediamine (en), 1,2-diaminocyclohexane (dach) or 2,2'-bipyridine (bpy), have shown strong binding to calf thymus DNA and moderate in vitro cytotoxicity towards cancer cell lines. Knowing that serum proteins play a crucial role in the transport and deactivation of ruthenium drugs, we have conducted a detailed study of their interactions with two major metal-transporting serum proteins, albumin and transferrin, and it is presented herein. Ruthenated protein adducts were formed with various concentrations of the three compounds and then separated from the unbound portions by ultrafiltration through 10 kDa cut-off centrifugal filter units. The stoichiometry of binding was determined using inductively coupled plasma optical emission spectrometry. One mol of albumin bound up to 7, 8.5 and 1.5 mol of compound 1 ([Ru(Cl-tpy)(en)Cl][Cl]), 2 ([Ru(Cl-tpy)(dach)Cl][Cl] and 3 ([Ru(Cl-tpy)(bpy)Cl][Cl]), respectively. One mol of transferrin bound up to 3, 3.5 and 0.4 mol of 1, 2 and 3, respectively. The affinity of albumin and transferrin for the three ruthenium compounds was evaluated using fluorescence quenching. The binding constants for 1 and 2 lay within the range 104–105 M−1, suggesting moderate-to-strong attachment to albumin. Both compounds showed much lower affinity for transferrin (102–103 M−1). Compound 3 bound weakly to each studied protein. High resolution ESI qTOF mass spectra of albumin before and after binding of 1 revealed the high stoichiometry of binding. Although the binding of the compounds 1–3 to albumin and transferrin did not affect proteins’ secondary structure much, their tertiary structures underwent some alterations, as deduced from the circular dichroism study. Changes in the stability of albumin, after binding to compounds 1–3 were examined by differential scanning calorimetry.

Keywords

• Anticancer drugs
• Ru(II) terpyridine coordination compounds
• Serum albumin binding
• Serum transferrin binding
• CD spectra
• ESI qTOF MS spectra