Synthesis, characterization and cytotoxic studies of water soluble [(η5-C5H5)2Mo(thionucleobase/thionucleoside)]Cl complexes in breast and colon cancer cell lines
Graphical abstract
Four new water soluble molybdenocene complexes, [(η5-C5H5)2Mo(L)]Cl (L = 6-mercaptopurine, 2-amino-6-mercaptopurine, (−)-2-amino-6-mercaptopurine ribose and 6-mercaptopurine ribose) were synthesized in aqueous solution at pH 7.0. The thionucleobases as ancillary ligands enhance water solubility and cytotoxic activity of the molybdenocenes on MCF-7 cancer cell line must likely as a result of synergism between both moieties.
Research highlights
► Mo(IV) could be coordinated by the nearby amino acids in the estrogen receptor ligand binding domain behaving as an agonist. ► Synergism between Cp2Mo2+ and 2-amino-6-mercaptopurine leads to highly antitumor species in MCF-7. ► Incorporation of Cp2Mo2+ into the thionucleobase increases transport across the cell membrane.
Introduction
During the last century, metals have been used to treat fungal infections, arthritis, ulcers and other conditions. However, the interest of metals in medicine was awakened by the accidental discovery of cis-platin, Pt(NH3)2Cl2, as potential anticancer agent in 1970 [1]. In subsequent years, other metals were explored for medicinal applications ranging from cancers to diagnostic, immunoassay, biosensors, enzymes and proteins [2].
The first non-platinum complex tested in clinical trials as antitumor agent was cis-[(CH3CH2O)2(bzac)2Ti(IV)], bzac = 1-phenylbutane-1,3-dioanate [3]. This complex is active against a wide variety of ascites and solid tumors [3], [4]. Other cis-[X2(bzac)2Ti(IV)], X = halides, complexes have been investigated exhibiting similar biological activity as the ethoxide complex [4]. However, the interest for non-platinum complex proceeded slowly because of the pharmacological efficacy and potent antitumor properties of cis-platin.
The discovery of metallocene-based organometallic anticancer agent, titanocene dichloride (Cp2TiCl2), in 1979 by Köpf and Köpf-Maier [5] stimulated much interest to investigate other non-platinum complexes with a different mechanism of anticancer activity. Other diacido metallocene complexes, Cp2MX2 (Cp = cyclopentadienyl, M = Ti, V, Nb, Mo; X = halides and pseudo-halides), have been investigated for antitumor activity [6], [7], [8], [9], [10], [11]. Among these metallocene complexes, Cp2TiCl2 has been studied in details but it possesses a major drawback: hydrolyzes extensively at physiological pH [12]. In contrast, molybdenocene dichloride is stable at physiological pH, producing Cp2Mo(H2O)(OH)+ [13]. Thus, there is a great potential to synthesize water soluble and stable molybdenocene species and study their biological properties.
Several water-soluble thiols derivatives of molybdenocene have been prepared and cell uptake and cytotoxic studies were pursued by Harding and coworkers [14]. It was found that the molybdenocenes containing thiols as ancillary ligands are robust toward hydrolysis but yielded non-cytotoxic complexes [14]. Strong coordination of molybdenocene by the thiolates apparently hinders the formation of vacant coordination sites on the metal center and the net result is inactivation. In the past, we reported a series of water soluble molybdenocenes containing thionucleobases [15]. These species were synthesized under non-aqueous solution. These complexes are stable at physiological pH but under these conditions, three or more species are produced in solution with unknown chemical formula. However, it is unclear as to which species have anticancer properties. Therefore, we decided to revisit the synthesis of molybdenocene–thionucleobase/thionulceoside complexes in aqueous solution at pH of 7.0, leading to the formation of more water stable species with features different to the previously synthesized molybdenocene–thionucleobase/thionulceoside complexes under non-aqueous solution. Herein we report the synthesis, characterization and cytotoxic activity of these molybdenocenes in HT-29 colon cancer and MCF-7 breast cancer cell lines.
Section snippets
Synthesis and spectroscopic characterization of [Cp2Mo(L)]Cl
At neutral pH molybdenocene dichloride undergoes chloride hydrolysis forming a stable species, Cp2Mo(OH)(H2O)+ [13], [16]. Taking advantage of the stability of molybdenocene dichloride at neutral pH, we performed the reaction at pH 7.0. The reaction of Cp2MoCl2 with the corresponding thionulceobase or thionucleoside in degassed water at pH 7.0 leads to the formation of, after column chromatography using lipophilic Sephadex, orange solids of general formula [Cp2Mo(L)]Cl, L = 6-mercaptopurine,
Concluding remarks
We have synthesized a series of molybdenocene–thionucleobase and –thionucleoside complexes in aqueous solution at pH 7.0. This simple synthetic route provided highly soluble and stable complexes in water. Two coordination isomers (S(6), N(7) and S(6), N(1)) were observed by NMR spectroscospy, forming 5-member and 4-member chelate rings. We hypothesize that S(6), N(7) coordination is the major isomer in aqueous solution.
The cytotoxic properties of these new species were investigated on HT-29
Methods and materials
All starting materials were obtained from Aldrich and used without further purification. The purity of molybdenocene dichloride and ligands were checked by IR and/or by 1H NMR spectroscopy to determine possible decomposition. Water was doubly distilled, deionized and thoroughly saturated with dried nitrogen. All solvents for NMR measurements were 99.9% D purity grade. The 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (yellow tetrazolium MTT) was obtained from Sigma and used as
Acknowledgements
E.M. acknowledges the NIH-MBRS SCORE Programs at both the University of Puerto Rico Mayagüez and the Ponce School of Medicine (PSM) for financial support via NIH-MBRS-SCORE Program grant #S06 GM008103-37 and #S06 GM008239-23 and the PSM-Moffitt Cancer Center Partnership 1U56CA126379-01 grant. In addition, EM thanks NSF-MRI Program for providing funds for the purchase of the 500 MHz NMR instrument.
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