Synthesis and biological characterization of organoruthenium complexes with 8-hydroxyquinolines.

In this study we report the synthesis, characterization and a thorough biological evaluation of twelve organoruthenium-8-hydroxyquinolinato (Ru-hq) complexes. The chosen hqH ligands bear various halogen atoms in different positions which enables to study effect of the substituents on physico-chemical and biological properties. The determined crystal structures of novel complexes expectedly show the cymene ring, a bidentately coordinated deprotonated hq and a halide ligand (chlorido or iodido) coordinated to the ruthenium central ion. In previous studies the anticancer potential of organoruthenium complex with 8-hydroxyquinoline ligand clioquinol was well established and we have decided to perform an extended biological evaluation (antibacterial and antitumor activity) of the whole series of halo-substituted analogs. Beside the cytotoxic potential of studied compounds also the effect of two selected complexes (9 and 10) on apoptosis induction in MG-63 and A549 cells was also studied via externalization of phosphatidylserine at the outer plasma membrane leaflet. Both selected complexes that gave best preliminary cytotoxicity results contain bromo substituted hq ligands. Apoptosis induction results are in agreement with the cell viability assays suggesting the higher and more selective anticancer activity of complex 10 in comparison to complex 9 on MG-63 cells.

Decoding the anticancer activity of VO-clioquinol compound: the mechanism of action and cell death pathways in human osteosarcoma cells.

Vanadium compounds were studied in recent years by considering them as a representative of a new class of non-platinum metal anticancer drugs. However, a few challenges still remain in the discovery of new molecular targets of these new metallodrugs. Studies on cell signaling pathways related to vanadium compounds have scarcely been reported and so far this information is highly critical for identifying novel targets that play a key role in the antitumor actions of vanadium complexes. This research deals with the alterations in the intracellular signaling pathways promoted by an oxovanadium(iv) complex with the clioquinol (5-chloro-7-iodo-8-quinolinol), VO(CQ)2, on a human osteosarcoma cell line (MG-63). Herein are reported, for the first time, the antitumor properties of VO(CQ)2 and the relative abundance of 224 proteins (which are involved in most of the common intracellular pathways) to identify novel targets of the studied complex. Besides, full-length human recombinant AKT1 kinase was produced by using an IVTT system to evaluate the variation of relative tyrosin-phosphorylation levels caused by this compound. The results of the differential protein expression levels reveal several up-regulated proteins such as CASP3, CASP6, CASP7, CASP10, CASP11, Bcl-x, DAPK and down-regulated ones, such as PKB/AKT, DIABLO, among others. Moreover, cell signaling pathways involved in several altered pathways related to the PKC and AP2 family have been identified in both treatments (2.5 and 10 µM) suggesting the crucial antitumoral role of VO(CQ)2. Finally, it has been demonstrated that this compound (10 µM, 6 h) triggers a decrease of 2-fold in in situ AKT1 expression.

Copper(II) Complexes with Saccharinate and Glutamine as Antitumor Agents: Cytoand Genotoxicity in Human Osteosarcoma Cells.

BACKGROUND: Copper has shown to be useful in disorders with an inflammation origin such as cancer [1-3]. It has previously shown that Casiopeínas® interact with DNA and promote the disruption by a mechanism related to the increase in the level of free radicals [4-6] which confers antineoplastic potential. Objetive: The aim of the present work was to study the antitumor effects of a series of Cu(II) complexes with saccharinate (sac) and glutamate (gln): [Cu(sac)2(H2O)4].2H2O (Cu-sac), [Cu(gln)2] (Cu-gln) and Na2[Cu(sac)2 (gln)2].H2O (Cu-sac-gln). METHODS: We have investigated the action of these compounds on cell viability on human osteosarcoma cells MG-63. In particular, we pay special attention to the cyto and genotoxicity actions of these complexes and to the association to oxidative stress. RESULTS: The three complexes: Cu-sac, Cu-gln and Cu-sac-gln caused a decline in cell viability. The half-maximal inhibitory concentration in MG-63 cells for Cu-sac-gln is 170 µM, showing the strongest antiproliferative effect. Moreover, only Cu-sac-gln caused a decrease of the mitochondrial activity from 100 µM. Our results indicate that the copper(II) complexes studied here produce DNA damage and suggest that the rise of reactive oxygen species (ROS) is the central mechanism action. Genotoxicity studied by the Cytokinesis-block micronucleus (MN) assay and the Single cell gel electrophoresis (comet assay) could be observed in MG-63 cells treated with Cu-sac-gln from 100 and 50 µM, respectively. Cu-sac and Cu-gln also induced DNA damage; however their effect was definitively weaker. The generation of reactive oxygen species increased from 50 µM of Cu-sac-gln and Cu-sac and only from 250 µM of Cugln, as well as a reduction of the GSH/GSSG ratio from 50 µM. When cells were treated with several concentrations of the complexes in addition to a combination of 50 µM of vitamin C plus 50 µM of vitamin E, a total recovery in cell survival was obtained for Cu-gln in the whole range of tested concentrations while only a partial viability recovery was obtained from 250 µM of Cu-sac and Cu-sac-gln. CONCLUSION: Overall, our results point to a differential cyto- and genotoxicity of the three copper(II) complexes and demonstrate that the complexation with both ligands confers the most potent antitumor action in human osteosarcoma cells.

In vitro and in vivo antitumor effects of the VO-chrysin complex on a new three-dimensional osteosarcoma spheroids model and a xenograft tumor in mice.

Osteosarcoma (OS) is the most common primary tumor of bone, occurring predominantly in the second decade of life. High-dose cytotoxic chemotherapy and surgical resection have improved prognosis, with long-term survival for patients with localized disease. Vanadium is an ultra-trace element that after being absorbed accumulates in bone. Besides, vanadium compounds have been studied during recent years to be considered as representative of a new class of non-platinum antitumor agents. Moreover, flavonoids are a wide family of polyphenolic compounds that display many interesting biological effects. Since coordination of ligands to metals can improve the pharmacological properties, we report herein, for the first time, the in vitro and in vivo effects of an oxidovanadium(IV) complex with the flavonoid chrysin on the new 3D human osteosarcoma and xenograft osteosarcoma mice models. The pharmacological results show that VOchrys inhibited the cell viability affecting the shape and volume of the spheroids and VOchrys suppressed MG-63 tumor growth in the nude mice without inducing toxicity and side effects. As a whole, the results presented herein demonstrate that the antitumor action of the complex was very promissory on human osteosarcoma models, whereby suggesting that VOchrys is a potentially good candidate for future use in alternative antitumor treatments.

Deciphering the effect of an oxovanadium(iv) complex with the flavonoid chrysin (VOChrys) on intracellular cell signalling pathways in an osteosarcoma cell line.

Vanadium complexes were studied during recent years and considered as a representative of a new class of non-platinum metal antitumor agents in combination with their low toxicity. However, a few challenges still remain in the discovery of new molecular targets for these novel metal-based drugs. The study of cell signaling pathways related to vanadium drugs, which is highly critical for identifying specific targets that play an important role in the antitumor activity of vanadium compounds, is scarce. This research deals with the alterations in intracellular signaling pathways promoted by an oxovanadium(iv) complex with the flavonoid chrysin [VO(chrysin)2EtOH]2 (VOChrys) in a human osteosarcoma cell line (MG-63). Herein we report for the first time the effect of [VO(chrysin)2EtOH]2 on the relative abundance of 224 proteins, which are involved in the most common intracellular pathways. Besides, full-length human recombinant (FAK and AKT1) kinases are produced using an in situ IVTT system and then we have evaluated the variation of relative tyrosine-phosphorylation levels caused by the [VO(chrysin)2EtOH]2 compound. The results of the differential protein expression levels reveal that several proteins such as PKB/AKT, PAK, DAPK, Cdk 4, 6 and 7, FADD, AP2, NAK, and JNK, among others, were altered. Moreover, cell signaling pathways related to the PTK2B, FAK, PKC families suggests an important role associated with the antitumor activity of [VO(chrysin)2EtOH]2 was demonstrated. Finally, the effect of this compound on in situ expressed FAK and AKT1 is validated by determining the phosphorylation level, which decreased in the former and increased in the latter.

A new oxidovanadium(IV) complex of oxodiacetic acid and dppz: spectroscopic and DFT study. Antitumor action on MG-63 human osteosarcoma cell line.

The oxidovanadium(IV) complex of oxodiacetic acid (H2ODA) and dppz (dipyrido[3,2-a:2',3'-c] phenazine) of stoichiometry [VO(ODA)(dppz)]·3H2O could be synthesized for the first time by reaction between [VO(ODA)(H2O)2] and dppz. It was characterized by infrared and electronic spectroscopies. Its optimized molecular structure was obtained by DFT calculations, as it was impossible to grow single crystals adequate for crystallographic studies. The antitumor action of the complex on MG-63 human osteosarcoma cell line was also investigated. It was found that it caused a concentration-related inhibitory effect in the concentration range between 5 and 25 µM and diminished the cell viability ca. 45% in the range from 25 to 100 µM, without dose/response effects in this range. These biological effects are, in general, similar to those previously reported for the related [VO(ODA)(ophen)]·1.5H2O complex.

Biological evaluation of morin and its new oxovanadium(IV) complex as antioxidant and specific anti-cancer agents.

It is known that flavonoids possess, among others, antioxidant and antitumoral properties that depend on their molecular structure. The central objective if this study was to investigate the potential antioxidant and antiproliferative properties of the flavonol morin and its new oxovanadium(IV) complex (VOmor) that was synthesized in order to modify the morin chemical structure. Two osteoblast (UMR106 and MC3T3E1), two breast tumor (T47D and SKBR3) and breast epithelial cell lines in culture were used for the antitumoral determinations. Additionally, a comparative study of their antioxidant capacities using different radicals (DPPH, ABTS(+), OH, O2(-), ROO) was performed. Selected mechanisms of action were studied using the breast cancer cell lines. Results obtained show that morin and its complex behaved as good antioxidant agents for some of the radicals and that the complexation improved the behavior with respect to OH and O2(-) radicals being morin more effective as ROO scavenger. A considerable variation in sensitivity was observed in the breast cancer cells but non-specificity was found for the treatment of osteosarcoma. Moreover, the compounds did not affect the normal proliferation of the breast epithelial mammal cells. The mechanistic studies demonstrated that the complex did not generate reactive oxygen species in the cells (confirming the in vitro studies) and did not produce any damage of DNA. The plasmatic membrane was observed to be damaged only in the SKBR3 cell line. In contrast, the perturbation of the mitochondrial membrane potential and the activation of caspase 3/7 for the breast tumor cells revealed an apoptotic cell death process. All these results collectively suggested that VOmor complex could serve as promising pharmacologically active substance against breast cancer treatment.

Bis(oxalato)dioxovanadate(V) and bis(oxalato)oxoperoxo-vanadate(V) complexes: spectroscopic characterization and biological activity.

Two structurally related vanadium(V) complexes, K3[VO2(C2O4)2] · 3H2O and K3[VO(O2)(C2O4)2] · 1/2H2O, were thoroughly characterized by infrared, Raman, and electronic spectroscopies. The effect of both complexes on the viability of the human MG-63 osteosarcoma cells was tested using the MTT assay. The monoperoxo complex shows a very strong antiproliferative activity (at 100-µM concentration, this complex diminished the cell viability ca. 80 %), whereas the dioxo complex was inactive.

Hydroxylamido-amino acid complexes of oxovanadium(V). Toxicological study in cell culture and in a zebrafish model.

Oxovanadium(V) complexes [VO(NH(2)O)(2)(val)] and [VO(NH(2)O)(2)(met)] caused inhibition of cell proliferation in two osteoblast cell lines, MC3T3-E1 and UMR106, as well as the viability of zebrafish eggs. In MC3T3-E1, both compounds inhibited cell proliferation (up to ca. 40% at 25 µM [VO(NH(2)O)(2)(val)] and 25% at 25 µM [VO(NH(2)O)(2)(met)]). This effect occurs in a dose response manner from 2.5 µM (p < 0.01) with a more deleterious action of [VO(NH(2)O)(2)(met)]. In UMR106 tumoral cells, [VO(NH(2)O)(2)(val)] inhibited cell proliferation up to 75% from 25 µM while [VO(NH(2)O)(2)(met)] behaved as an inhibitory agent in the whole range of concentrations (p < 0.01). Similar toxic effects were obtained from morphological studies in cell cultures. Moreover, the IC(50) values for both complexes in culture studies correlated with the IC(50) values obtained with an in vivo model of toxicity (FET test). Besides, the cytotoxicity evaluation in cell culture showed a decrease in mitochondrial activity which was stronger for [VO(NH(2)O)(2)(met)] than for [VO(NH(2)O)(2)(val)] (44% vs. 58% at 25 µM) in both cell lines (p < 0.001). Genotoxicity assessed by micronuclei induction also showed a stronger effect of [VO(NH(2)O)(2)(met)] in both cell lines. Besides, [VO(NH(2)O)(2)(val)] caused DNA damage determined by comet formation in MC3T3-E1 cells in the range of 2.5-25 µM, while this effect could not be observed in the osteosarcoma cells. On the other hand, [VO(NH(2)O)(2)(val)] enhanced ROS levels over basal up to 225% and 170% at 100 µM in MC3T3-E1 and UMR106 cells, respectively (p < 0.01). For [VO(NH(2)O)(2)(met)] a similar situation was observed, suggesting an important role for oxidative stress in the toxicity mechanism of action. Although both complexes showed interesting results that would deserve further drug development [VO(NH(2)O)(2)(val)] was more stable than [VO(NH(2)O)(2)(met)] in the solid state. Therefore, we consider that [VO(NH(2)O)(2)(val)] is a good candidate to be tested in in vivo models as a potential antitumoral agent.

Spectroscopic characterization of an oxovanadium(IV) complex of oxodiacetic acid and o-phenanthroline. Bioactivity on osteoblast-like cells in culture.

The oxovanadium(IV) complex of oxodiacetic acid (H(2)ODA) and o-phenanthroline of stoichiometry [VO(ODA)(ophen)]·1.5H(2)O, which presents the interesting tridentate OOO coordination, was thoroughly characterized by infrared, Raman, and electronic spectroscopies. The biological activity of the complex on the cell proliferation was tested on osteoblast-like cells (MC3T3E1 osteoblastic mouse calvaria-derived cells and UMR106 rat osteosarcoma-derived cells) in culture. The complex caused inhibition of cellular proliferation in both osteoblast cell lines in culture, but the cytotoxicity was stronger in the normal (MC3T3E1) than in the tumoral (UMR106) osteoblasts.

Dinuclear copper(II) complexes with valsartan. Synthesis, characterization and cytotoxicity.

Two novel dinuclear complexes involving the antihypertensive drug valsartan and copper(II) ion have been prepared in water and DMSO. The complex compositions were determined as: [Cu(vals)(H(2)O)(3)](2).6H(2)O and [Cu(vals)(H(2)O)(2)DMSO](2).2H(2)O. They were thoroughly characterized by elemental and thermal analysis, spectrophotometric titrations and UV-visible, diffuse reflectance, FTIR, Raman and EPR spectroscopies. No effect of the ligand on two tested osteoblastic cell lines in culture (one normal MC3T3E1 and one tumoral UMR106) was observed in concentrations up to 100 µM. Higher concentrations of Valsartan are required to induce cytotoxicity in both cell lines. The antiproliferative effect of the tested complex ([Cu(vals)(H(2)O)(3)](2).6H(2)O) in a dose-response manner, was higher in the UMR106 osteoblastic cell line than that of the MC3T3E1 normal line at concentrations ≥100 µM. Morphological alterations are in accordance with proliferative observations.

Cyto- and genotoxicity of a vanadyl(IV) complex with oxodiacetate in human colon adenocarcinoma (Caco-2) cells: potential use in cancer therapy.

The complex of vanadyl(IV) cation with oxodiacetate, VO(oda) caused an inhibitory effect on the proliferation of the human colon adenocarcinoma cell line Caco-2 in the range of 25-100 µM (P < 0.001). This inhibition was partially reversed by scavengers of free radicals. The difference in cell proliferation in the presence and the absence of scavengers was statistically significant in the range of 50-100 µM (P < 0.05). VO(oda) altered lysosomal and mitochondria metabolisms (neutral red and MTT bioassays) in a dose-response manner from 10 µM (P < 0.001). Morphological studies showed important transformations that correlated with the disassembly of actin filaments and a decrease in the number of cells in a dose response manner. Moreover, VO(oda) caused statistically significant genotoxic effects on Caco-2 cells in the low range of concentration (5-25 µM) (Comet assay). Increment in the oxidative stress and a decrease in the GSH level are the main cytotoxic mechanisms of VO(oda). These effects were partially reversed by scavengers of free radicals in the range of 50-100 µM (P < 0.05). Besides, VO(oda) interacted with plasmidic DNA causing single and double strand cleavage, probably through the action of free radical species. Altogether, these results suggest that VO(oda) is a good candidate to be evaluated for alternative therapeutics in cancer treatment.

Antioxidant, DNA cleavage, and cellular effects of silibinin and a new oxovanadium(IV)/silibinin complex.

A new complex of the oxovanadium(IV) cation with the flavolignan silibinin has been synthesized and characterized. Vanadium compounds show interesting biological and pharmacological properties and some of them display antitumoral actions. Flavonoids are part of a larger group of antioxidant compounds called polyphenols which may inhibit the proliferation and growth of cancer cells. The antioxidant and antitumoral effects of silibinin and its oxovanadium(IV) complex were investigated. Silibinin acted as a very strong antioxidant and its complexation with oxovanadium(IV) improved this behavior. Besides, the generation of reactive oxygen species (ROS) by this compound was favored in tumoral (UMR106) cells and correlated with the deleterious behavior in the proliferation of this cell line. Conversely, silibinin did not exert any effect on the proliferation of normal osteoblasts (MC3T3E1). The cytotoxic action and ROS generation of the oxovanadium(IV) complex was more effective in tumoral cells. This behavior was not consistent with cleaving DNA of plasmid DNA pA1 because no significant cleaving activity was observed in both cases. These results suggest that the main deleterious mechanisms may take place through cytotoxic effects more than genotoxic actions. A comparison with our own findings on the behavior of other flavonoids and their vanadyl(IV) complex has also been performed.

Spectroscopic behavior and biological activity of K2[VO(O2)NTA].2H2O.

The dihydrated potassium salt of the complex anion [VO(O(2))NTA](2-) (NTA = nitrilotriacetate anion, [N(CH(2)-COO)(3)](3-)) was thoroughly characterized by electronic and vibrational (infrared and Raman) spectroscopies. The bioactivity of the complex on the cell proliferation was tested on three cell lines in culture (UMR106 rat osteosarcoma-derived cells, Caco-2 derived from a human colon adenocarcinoma, and RAW 264.7, a macrophage murine cell line).

Universal versus selective screening for the detection, control and prognosis of gestational diabetes mellitus in Argentina.

In all, 1,702 unselected pregnant women from the city of La Plata were tested for gestational diabetes mellitus (GDM) and evaluated to determine GDM prevalence and risk factors. In women with GDM, we evaluated compliance with guidelines for GDM management, and perinatal complications attributable to GDM. GDM prevalence was 5.8%, and its risk factors were pre-gestational obesity, previous hyperglycaemia, age > 30 years, previous GDM (and its surrogate markers). In primi-gravida (PG) subjects, GDM was equally prevalent in the presence (4.2%) or absence (4.0%) of risk factors. In multi-gravida (MG) women, although risk factors doubled the prevalence of GDM (8.6%), in the absence of risk factors GDM prevalence was similar to that of PG women (3.9%). Half of all women with GDM received inadequate post-diagnosis obstetric control, and this induced a fourfold increase in infant perinatal complications. In conclusion, all non-hyperglycaemic 24-28-week pregnant women should be tested for GDM, although particular attention must be paid to MG women with risk factors.

Biological effects of a complex of vanadium(V) with salicylaldehyde semicarbazone in osteoblasts in culture: mechanism of action.

Vanadium compounds display important pharmacological actions in vivo and in vitro systems. Semicarbazones are versatile ligands with therapeutic effects. Herein, we report the effects of V(V)O(2)(salicylaldehydesemicarbazone) (V(V)-Salsem) on two osteoblast cell lines in culture (MC3T3-E1 and UMR106). V(V)-Salsem inhibited cell proliferation in a dose response manner. At 100muM, the complex caused an inhibition of ca. 48% and 38% for the normal and the tumoral osteoblasts, respectively (p<0.001). This inhibition could be partially reversed to 35% and 28% by NAC (N-acetylcysteine) and a mixture of vitamins E and C. Changes in cell proliferation correlated with morphological alterations and the disruption of actin cytoskeleton fibers. The complex also enhanced the level of ROS (reactive oxygen species) up to ca. 100% over basal in both cell lines. Activation of ERK signalling cascade was also observed. These events led to apoptosis (up to 44% in MC3T3-E1 and 33% in UMR106 cells). Scavengers of ROS and inhibitors of ERK cascade allowed to elucidate the mechanisms involved in the cytotoxicity. In conclusion, V(V)-Salsem displayed cytotoxic effects on osteoblasts in culture through the production of free radicals and the activation of ERK cascade. These mechanisms triggered the apoptotic events that conveyed to cell death.

Potential antitumoral properties of a new copper complex with santonic acid.

A new copper(II) complex of santonic acid [Cu(2)(sant)(4)(H(2)O)(2)].2(1/2)H(2)O has been prepared and characterized by electronic, vibrational, EPR spectral studies, and stability determinations in solution. The presence of two antiferrromagnetically coupled copper centers in the solid state was detected by EPR. The dinuclear Cu(II) complex crystallizes in the tetragonal P4(3)2(1)2 space group, with a=b=14.498(3), c=64.07(1)A. Biological studies indicate that the complex displays interesting potential antitumoral actions.

Vanadium(IV) complexes inhibit adhesion, migration and colony formation of UMR106 osteosarcoma cells.

Vanadium is a trace element widely distributed in the environment. In vertebrates it is mainly stored in bone tissue. The unique cellular environment in the bone and the variety of interactions that mediate cancer metastasis determine that certain types of cancer, such as breast and prostate cancer, preferentially metastize in the skeleton. Since this effect usually signifies serious morbidity and grave prognosis there is an increasing interest in the development of new treatments for this pathology. The present work shows that vanadium complexes can inhibit some parameters related to cancer metastasis such as cell adhesion, migration and clonogenicity. We have also investigated the role of protein kinase A in these processes.

Losartan and its interaction with copper(II): biological effects.

Losartan, the potassium salt of 2-n-butyl-4-chloro-5-hydroxymethyl-1-[(2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]imidazol, is an efficient antihypertensive drug. The vibrational FTIR and Raman spectra of Losartan (its anionic and protonated forms) are discussed. In addition, the copper(II) complex of Losartan was obtained and characterized as a microcrystalline powder. The metal center is bound to the ligand through the nitrogen atoms of the tetrazolate moiety as determined by vibrational spectroscopy. The compound is a dimer with the metal centers in a tetragonal distorted environment but the presence of a monomeric impurity has been determined by EPR spectroscopy. The antioxidant properties of the complex (superoxide dismutase mimetic activity) and its effect on the proliferation and morphology of two osteoblast-like cells in culture are reported. The new compound exerted more toxic effects on tumoral cells than the copper(II) ion and Losartan.

Regulation of advanced glycation end product (AGE) receptors and apoptosis by AGEs in osteoblast-like cells.

Advanced glycation end products (AGEs) have been proposed as the pathological mechanisms underlying diabetic chronic complications. They may also play a role in the pathogenesis of diabetic osteopenia, although their mechanisms of action remain unclear. We investigated the protein (immunofluorescence) and gene expression (realtime RT-PCR) of two receptors for AGEs, RAGE and galectin-3, as well as their regulation by AGEs, and the apoptotic effect on osteoblast-like cells (UMR106 and MC3T3E1) in culture. AGEs up-regulated the expression of RAGE and galectin-3 in both cells lines. These effects were accompanied by an increase in the corresponding mRNA in the non-tumoral MC3T3E1 but not in the osteosarcoma UMR106 cells. Finally, we demonstrated that a 24 h exposure to AGEs induced apoptosis in both cell lines. Thus, AGEs-receptors may play important roles in the bone alterations described in aging and diabetic patients.