Article pubs.acs.org/IC Conjugation of Organoruthenium(II) 3-(1H-Benzimidazol-2- yl)pyrazolo[3,4-b]pyridines and Indolo[3,2-d]benzazepines to Recombinant Human Serum Albumin: a Strategy To Enhance Cytotoxicity in Cancer Cells Iryna N. Stepanenko,† Angela Casini,‡,⊥ Fabio Edafe,‡ Maria S. Novak,† Vladimir B. Arion,*,† Paul J. Dyson,*,‡ Michael A. Jakupec,† and Bernhard K. Keppler*,† †Institute of Inorganic Chemistry, University of Vienna, Waḧringer Strasse 42, A-1090 Vienna, Austria ‡Institut des Sciences et Ingeńierie Chimiques, Ecole Polytechnique Fed́eŕale de Lausanne (EPFL), CH-1015 Lausanne, Switzerland *S Supporting Information ABSTRACT: Following our strategy of coupling cyclin-dependent kinase (Cdk) inhibitors with organometallic moieties to improve their physicochemical properties and bioavailability, five organoruthenium complexes (1c−5c) of the general formula [RuCl(η6- arene)(L)]Cl have been synthesized in which the arene is 4-formylphenoxyacetyl-η6- benzylamide and L is a Cdk inhibitor [3-(1H-benzimidazol-2-yl)-1H-pyrazolo[3,4- b]pyridines (L1−L3) and indolo[3,2-d]benzazepines (L4 and L5)]. All of the compounds were characterized by spectroscopic and analytical methods. Upon prolonged standing (2−3 months) at room temperature, the dimethyl sulfoxide (DMSO) solutions of 1c and 2c−HCl afforded residues, which after recrystallization from EtOH and EtOH/H2O, respectively, were shown by X-ray diffraction to be cis,cis-[RuIICl2(DMSO)2(L1)]·H2O and mer-[RuIICl(DMSO)3(L2−H)]·H2O. Compound 5c, with a coordinated amidine unit, undergoes E/Z isomerization in solution. The antiproliferative activities and effects on the cell cycle of the new compounds were evaluated. Complexes 1c−5c are moderately cytotoxic to cancer cells (CH1, SW480, A549, A2780, and A2780cisR cell lines). Therefore, in order to improve their antiproliferative effects, as well as their drug targeting and delivery to cancer cells, 1c−5c were conjugated to recombinant human serum albumin, potentially exploiting the so-called “enhanced permeability and retention” effect that results in the accumulation of macromolecules in tumors. Notably, a marked increase in cytotoxicity of the albumin conjugates was observed in all cases. ■ INTRODUCTION Numerous strategies have been developed for the effective delivery of anticancer drugs to tumor tissue to improve their selectivity and, consequently, to reduce drug side effects.1−4 By using passive and active targeting strategies, cancer nano- therapeutics, based on polymers (polymeric nanoparticles, micelles, or dendrimers), lipids (liposomes), viruses (viral nanoparticles), and carbon nanotubes, leads to an enhancement of the intracellular concentration of drugs in cancer cells, usually without being blocked by P-glycoprotein, a protein responsible for multidrug resistance.5 These emerging approaches, mainly applied to organic anticancer drugs (e.g., doxorubicin, paclitaxel),6 have also been used successfully to deliver inorganic drugs, namely, platinum(II) and platinum(IV) complexes.7 Serum albumin has been observed to accumulate in solid tumors and, consequently, has been exploited as a drug-delivery system,8 involving both albumin conjugates for the delivery of anticancer agents and albumin nanoparticles for drug encapsulation. Interestingly, albumin conjugates with metho- trexate and a doxorubicin derivative and an albumin paclitaxel nanoparticle (nab-paclitaxel; Abraxane) have been evaluated in clinical trials.8,9 Albumin conjugates of the platinum(II) anticancer drug carboplatin were shown to be as, or more, effective than carboplatin in reducing the tumor size of nude mice bearing human breast tumors and, in some cases, were less toxic.10 Even if in a less advanced stage of development, an organometallic ruthenium compound has also been conjugated to recombinant human serum albumin (rHSA), with a considerable increase (ca. 20-fold) in cytotoxicity observed (see below).11,12 Organometallic ruthenium(II) arene complexes are currently under intensive investigations as anticancer agents,13−16 with several groups contributing to their design. Within this frame, and as part of our ongoing studies on targeted chemotherapy,17 involving the development of inhibitors of upregulated receptors and growth factors in cancer cells, we have studied the effect of metal coordination (Ga, Ru, Os, and Cu) of some cyclin-dependent kinase (Cdk) inhibitors (indolo[3,2-d]- benzazepines (paullones),18−23 indolo[3,2-c]quinolines,24 and 3-(1H-benzimidazol-2-yl)-1H-pyrazolo[3,4-b]pyridines25) on Received: August 18, 2011 Published: November 23, 2011 © 2011 American Chemical Society 12669 dx.doi.org/10.1021/ic201801e | Inorg. Chem. 2011, 50, 12669−12679