Ana Isabel Tomaz

DNA cleavage activity of VIVO(acac)2 and derivatives

The DNA cleavage activity of several beta-diketonate vanadyl complexes is examined. Vanadyl acetylacetonate, V(IV)O(acac)(2), 1, shows a remarkable activity in degrading plasmid DNA in the absence of any activating agents, air and photoirradiation. The cleaving activity of several related complexes V(IV)O(hd)(2) (2, Hhd=3,5-heptanedione), V(IV)O(acac-NH(2))(2) (3, Hacac-NH(2)=acetoacetamide) and V(IV)O(acac-NMe(2))(2) (4, Hacac-NMe(2)=N,N-dimethylacetoacetamide) is also evaluated. It is shown that 2 exhibits an activity similar to 1, while 3 and 4 are much less efficient cleaving agents. The different activity of the complexes is related to their stability towards hydrolysis in aqueous solution, which follows the order 1 approximately 2>>3 approximately 4. The nature of the pH buffer was also found to be determinant in the nuclease activity of 1 and 2. In a phosphate buffered medium DNA cleavage by these agents is much more efficient than in tris, hepes, mes or mops buffers. The reaction seems to take place through a mixed mechanism, involving the formation of reactive oxygen species (ROS), namely OH radicals, and possibly also direct cleavage at phosphodiester linkages induced by the vanadium complexes.

Interaction of vanadium(IV) with human serum apo-transferrin.

The interaction of V(IV)O-salts as well as of a few V(IV)O(carrier)n complexes with human serum transferrin (hTF) is studied focusing on the determination of the nature and stoichiometry of the binding of V(IV)O(2+) to hTF, as well as whether the conformation of hTF upon binding to V(IV)O(2+) or to its complexes is changed. Circular dichroism (CD) spectra measured for solutions containing V(IV)O(2+) and apo-hTF, and V(IV)O-maltol and apo-hTF, clearly indicate that hTF-V(IV)O-maltol ternary species form with a V(IV)O:maltol stoichiometry of 1:1. For V(IV)O salts and several V(IV)O(carrier)n complexes (carrier ligand=maltolato, dhp, picolinato and dipicolinato) (Hdhp=1,2-dimethyl-3-hydroxy-4-pyridinone) the maximum number of V(IV)O(2+) bound per mole of hTF is determined to be ~2 or lower in all cases. The binding of V(IV)O to apo-hTF most certainly involves several amino acid residues of the Fe-binding site, and as concluded by urea gel electrophoresis experiments, the formation of (V(IV)O)2hTF species may occur with the closing of the hTF conformation as is the case in (Fe(III))2hTF, which is an essential feature for the transferrin receptor recognition.

Novel ruthenium(II) cyclopentadienyl thiosemicarbazone compounds with antiproliferative activity on pathogenic trypanosomatid parasites.

Searching for new prospective antitrypanosomal agents, three novel Ru(II)-cyclopentadienyl compounds, [Ru(η(5)-C5H5)(PPh3)L], with HL=bioactive 5-nitrofuryl containing thiosemicarbazones were synthesized and characterized in the solid state and in solution. The compounds were evaluated in vitro on the blood circulating trypomastigote form of Trypanosoma cruzi (Dm28c strain), the infective form of Trypanosoma brucei brucei (strain 427) and on J774 murine macrophages and human-derived EA.hy926 endothelial cells. The compounds were active against both parasites with IC50 values in the micromolar or submicromolar range. Interestingly, they are much more active on T. cruzi than previously developed Ru(II) classical and organometallic compounds with the same bioactive ligands. The new compounds showed moderate to very good selectivity towards the parasites in respect to mammalian cells. The global results point at [RuCp(PPh3)L2] (L2=N-methyl derivative of 5-nitrofuryl containing thiosemicarbazone and Cp=cyclopentadienyl) as the most promising compound for further developments (IC50T. cruzi=0.41μM; IC50T. brucei brucei=3.5μM). Moreover, this compound shows excellent selectivity towards T. cruzi (SI>49) and good selectivity towards T. brucei brucei (SI>6). In order to get insight into the mechanism of antiparasitic action, the intracellular free radical production capacity of the new compounds was assessed by ESR. DMPO (5,5-dimethyl-1-pirroline-N-oxide) spin adducts related to the bioreduction of the complexes and to redox cycling processes were characterized. In addition, DNA competitive binding studies with ethidium bromide by fluorescence measurements showed that the compounds interact with this biomolecule.

Tracking antitumor metallodrugs: promising agents with the Ru(II)- and Fe(II)-cyclopentadienyl scaffolds

Research on the field of metal complexes for the treatment of cancer diseases has attracted increasing interest due to the urgency in finding more efficient and selective treatments. Owing to their wide structural diversity, organometallic complexes appear as potential alternatives to the design of new anticancer candidates. Herein, we review recent progress in our work toward the development of new drugs based on Ru(II)- and Fe(II)-cyclopentadienyl scaffolds. Their design and chemical properties are reviewed and correlated with their biological effects, in particular the key role that coligands play in the overall behavior of the complex.

The effect of phosphate on the nuclease activity of vanadium compounds

The nuclease activity of VO(acac)2 (1, acac = acetylacetone) and its derivatives VO(hd)2 (2, hd = 3,5-heptanedione), VO(Cl-acac)2 (3, Cl-acac = 3-chloro-2,4-pentanedione), VO(Et-acac)2 (4, Et-acac = 3-ethyl-2,4-pentanedione) and VO(Me-acac)2 (5, Me-acac = 3-methyl-2,4-pentanedione), is studied by agarose gel electrophoresis, UV-visible spectroscopy, cyclic and square wave voltammetry and (51)V NMR. The mechanism is shown to be oxidative and associated with the formation of reactive oxygen species (ROS). Hydrolytic cleavage of the phosphodiester bond is also promoted by 1, but at much slower rate which cannot compete with the oxidative mechanism. The generation of ROS is much higher in the presence of phosphate buffer when compared with organic buffers and this was attributed to the formation of a mixed-ligand complex containing phosphate, (V(IV)O)(V(V)O)(acac)2(HnPO4(n-3)), presenting a quasi-reversible voltammetric behavior. The formation of this species was further observed by Electrospray Ionization Mass Spectrometry (ESI-MS). Phosphate being an essential species in most biological media, the importance of the formation of mixed-ligand species in other vanadium systems is emphasized.

In Vivo Performance of a Ruthenium-cyclopentadienyl Compound in an Orthotopic Triple Negative Breast Cancer Model

BACKGROUND Ruthenium-based anti-cancer compounds are proposed as viable alternatives that might circumvent the disadvantages of platinum-based drugs, the only metallodrugs in clinical use for chemotherapy. Organometallic complexes in particular hold great potential as alternative therapeutic agents since their cytotoxicity involves different modes of action and present reduced toxicity profiles. OBJECTIVE During the last few years our research group has been reporting on a series of organometallic ruthenium(II)- cyclopentadienyl complexes with important cytotoxicity against several cancer cell lines, surpassing cisplatin in activity. We report herein preliminary in vivo studies with one representative compound of this family, with exceptional activity against several human cancer cell lines, including the glycolytic and highly metastatic MDAMB231 cell line used in this study. METHOD The anti-tumor activity of our compound was studied in vivo on N:NIH(S)II-nu/nu nude female mice bearing triple negative breast cancer (TNBC) orthotopic tumors. Administration of 2.5 mg/kg/day during ten days caused cell death mostly by necrosis (in vitro and in vivo), inducing tumor growth suppression of about 50% in treated animals when compared to controls. RESULTS The most remarkable result supporting the effectiveness and potential of this drug was the absence of metastases in the main organs of treated animals, while metastases were present in the lungs of all control mice, as revealed by histopathological and immunohistochemical analysis. CONCLUSION These in vivo studies suggest a dual effect for our drug not only by suppressing growth at the primary tumor tissue but also by inhibiting its metastatic behavior. Altogether, these results represent a benchmark and a solid starting point for future studies.

RuII(p-cymene) Compounds as Effective and Selective Anticancer Candidates with No Toxicity in Vivo

Ruthenium(II) complexes are currently considered a viable alternative to the widely used platinum complexes as efficient anticancer agents. We herein present the synthesis and characterization of half-sandwich ruthenium compounds with the general formula [Ru( p-cymene)(L-N,N)Cl][CF3SO3] (L = 3,6-di-2-pyridyl-1,2,4,5-tetrazine (1) 6,7-dimethyl-2,3-bis(pyridin-2-yl)quinoxaline (2)), which have been synthesized by substitution reactions from the precursor dimer [Ru( p-cymene)(Cl)(μ-Cl)]2 and were characterized by elemental analysis, mass spectrometry, 1H NMR, UV-vis, and IR spectroscopy, conductivity measurements, and cyclic voltammetry. The molecular structure for complex 2 was determined by single-crystal X-ray diffraction. The cytotoxic activity of these compounds was evaluated against human tumor cells, namely ovarian carcinoma A2780 and breast MCF7 and MDAMB231 adenocarcinoma cells, and against normal primary fibroblasts. Whereas the cytotoxic activity of 1 is moderate, IC50 values found for 2 are among the lowest previously reported for Ru( p-cymene) complexes. Both compounds present no cytotoxic effect in normal human primary fibroblasts when they are used at the IC50 concentration in A2780 and MCF7 cancer cells. Their antiproliferative capacity is associated with a combined mechanism of apoptosis and autophagy. A strong interaction with DNA was observed for both with a binding constant value of the same magnitude as that of the classical intercalator [Ru(phen)2(dppz)]2+. Both complexes bind to human serum albumin with moderate to strong affinity, with conditional binding constants (log Kb) of 4.88 for complex 2 and 5.18 for complex 1 in 2% DMSO/10 mM Hepes pH7.0 medium. The acute toxicity was evaluated in zebrafish embryo model using the fish embryo acute toxicity test (FET). Remarkably, our results show that compounds 1 and 2 are not toxic/lethal even at extremely high concentrations. The novel compounds reported herein are highly relevant antitumor metallodrug candidates, given their in vitro cytotoxicity toward cancer cells and the lack of in vivo toxicity.