Browsing by Author "Pessoa, Joao Costa"
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- Copper Complexes with 1,10-Phenanthroline Derivatives: Underlying Factors Affecting Their CytotoxicityPublication . Nunes, Patrique; Correia, Isabel; Marques, Fernanda; Matos, Antonio Pedro; dos Santos, Margarida M. C.; Azevedo, Cristina G.; Capelo, Jose-Luis; Santos, Hugo M.; Gama, Sofia; Pinheiro, Teresa; Cavaco, Isabel; Pessoa, Joao CostaThe interpretation of in vitro cytotoxicity data of Cu(II)-1,10-phenanthroline (phen) complexes normally does not take into account the speciation that complexes undergo in cell incubation media and its implications in cellular uptake and mechanisms of action. We synthesize and test the activity of several distinct Cu(II)-phen compounds; up to 24 h of incubation, the cytotoxic activity differs for the Cu complexes and the corresponding free ligands, but for longer incubation times (e.g., 72 h), all compounds display similar activity. Combining the use of several spectroscopic, spectrometric, and electrochemical techniques, the speciation of Cu-phen compounds in cell incubation media is evaluated, indicating that the originally added complex almost totally decomposed and that Cu(II) and phen are mainly bound to bovine serum albumin. Several methods are used to disclose relationships between structure, activity, speciation in incubation media, cellular uptake, distribution of Cu in cells, and cytotoxicity. Contrary to what is reported in most studies, we conclude that interaction with cell components and cell death involves the separate action of Cu ions and phen molecules, not [Cu(phen)(n)] species. This conclusion should similarly apply to many other Cu-ligand systems reported to date.
- Evaluation of the binding of four anti-tumor Casiopeinas to human serum albuminPublication . Correia, Isabel; Borovic, Sladjana; Cavaco, Isabel; Matos, Cristina P.; Roy, Somnath; Santos, Hugo M.; Fernandes, Luz; Capelo, Jose L.; Ruiz-Azuara, Lena; Pessoa, Joao CostaThe metal complexes designated by Casiopeinas (R) are mixed-ligand Cu-II-compounds some of them having promising antineoplastic properties. We report studies of binding of Cu(glycinato)(4,7-dimethyl-1,10-phenanthroline) (Cas-II-Gly (1)), Cu(acetylacetonato)(4,7-dimethy1-1,10-phenanthroline) (Cas-III-Ea (2)), Cu(glycinato) (4,4'-dimethyl-2,2'-bipyridine) (Cas-W-Gly (3)) and Cu(acetylacetonato)(4,4'-dimethyl-2,2'-bipyridine) (Cas-IIIia (4)) to human serum albumin (HSA) by circular dichroism (CD), Electron paramagnetic resonance (EPR) and fluorescence spectroscopy. The results indicate that HSA may bind up to three molecules of the tested Casiopeinas. This is confirmed by inductively coupled plasma atomic absorption spectroscopy measurements of samples of HSA-Casiopeinas after passing by adequate size-exclusion columns. The binding of Cas-II-Gly to HSA was also confirmed by MALDI-TOF mass spectrometric experiments. In the physiological range of concentrations the Casiopeinas form 1:1 adducts with HSA, with conditional binding constants of ca. 1 x 10(9) (1), 4 x 10(7) (2), 1 x 10(6) (3) and 2 x 10(5) (4), values determined from the CD spectra measured, and the fluorescence emission spectra indicates that the binding takes place close to the Trp214 residue. Overall, the data confirm that these Casiopeinas may bind to HSA and may be transported in blood serum by this protein; this might allow some selective tumor targeting, particularly in the case of Cas-Il-Gly. In this work we also discuss aspects associated to the reliability of the frequently used methodologies to determine binding constants based on the measurement of fluorescence emission spectra of solutions containing low concentrations of proteins such as HSA and BSA, by titration with solutions of metal complexes.
- Exploring the cytotoxic activity of new phenanthroline salicylaldimine Zn(II) complexesPublication . Matos, Cristina P.; Addis, Yemataw; Nunes, Patrique; Barroso, Sonia; Alho, Irina; Martins, Marta; Matos, Antonio P. A.; Marques, Fernanda; Cavaco, Isabel; Pessoa, Joao Costa; Correia, IsabelZinc(II) complexes bearing N-salicylideneglycinate (Sal-Gly) and 1,10-phenanthroline (phen) or phenanthroline derivatives [NN= 5-chloro-1,10-phenanthroline, 5-amine-1,10-phenanthroline (amphen), 4,7-diphenyl-1,10-phenanthroline (Bphen) and 5,6-epoxy-5,6-dihydro-1,10-phenanthroline] are synthesized. Complexes formulated as [Zn(NN)(2)(H2O)(2)](2+) (NN = phen and amphen), are also prepared. The cytotoxicity of the compounds is evaluated towards a panel of human cancer cells: ovarian (A2780), breast (MCF7) and cervical (HeLa), as well as non-tumoral V79 fibroblasts. All compounds display higher cytotoxicity than cisplatin (IC50 = 22.5 +/- 5.0 mu M) towards ovarian cells, showing IC50 values in the low micromolar range. Overall, all compounds show higher selectivity for the A2780 cells than for the non tumoral cells and higher selectivity indexes (IC50(V79)/IC50(A2780) than cisplatin. [Zn(Sal-Gly)(NNI(H2O)] complexes induce caspase-dependent apoptosis in A2780 cells, except [Zn(Sal-Gly)(Bphen)(H2O)], one of the most cytotoxic of the series. The cellular uptake in the ovarian cells analyzed by Inductively Coupled Plasma mass spectrometry indicates different Zn distribution profiles. Transmission electronic microscopy shows mitochondria alterations and apoptotic features consistent with caspase activationells incubated with EZn(Sal-Gly)(amphen)(H2O)] present additional nuclear membrane alterations in agreement with significant association with the nucleus. The increase of reactive oxygen species and lipid peroxidation forms could be related to apoptosis induction. [Zn(NN)(2)(H2O)(2)](2+) complexes have high ability to bind DNA through intercalation/groove binding, and circular dichroism data suggests that the main type of species that interact with DNA is [Zn(NN)](2+). Studies varying the % of fetal bovine serum (1-15%) in cell media show that albumin binding decreases the complex activity, indicating that distinct speciation of Zn- and phen-containing species in cell media may affect the cytotoxicity.
- Exploring the therapeutic potential of Cu(II)-complexes with ligands derived from pyridoxalPublication . Nunes, Patrique; Marques, Fernanda; Cavaco, Isabel; Pessoa, Joao Costa; Correia, IsabelThree new copper(II) complexes formulated as [Cu(L)(X)], where X = H2O or Cl and H2L is a Schiff base (H2L1,2) or its reduced version ((H3LCl)-Cl-3) derived from pyridoxal, are prepared, as well as two ternary complexes [Cu(L) (phen)] also containing 1,10-phenanthroline. All compounds are characterized by the usual techniques: elemental analyses, ESI mass spectrometry, UV-Vis absorption, FTIR and EPR spectroscopies. The ligands co-ordinate the Cu(II) center forming complexes with square-planar based geometries. Their antioxidant properties are evaluated with a radical scavenging activity assay, with one of the ligand precursors showing activity higher than the positive control, ascorbic acid. The antiproliferative activity of all compounds is evaluated against two cancer cell lines: ovarian (A2780) and breast (MCF7). All complexes show moderate to excellent activity with the ternary Cu-complexes showing IC50 values between 0.7 and 9.3 mu M after 24 h of incubation, values much lower than those reported for cisplatin, the reference drug. The hydrolytic stability of the complexes and their ability to bind albumin and DNA are evaluated by spectroscopic techniques, showing that the compounds bind bovine serum albumin. The [Cu(L)(phen)] complexes show ability to target DNA via intercalation.
- Interaction of [(VO)-O-IV(acac)(2)] with Human Serum Transferrin and AlbuminPublication . Correia, Isabel; Chorna, Ielyzaveta; Cavaco, Isabel; Roy, Somnath; Kuznetsov, Maxim L.; Ribeiro, Nadia; Justino, Goncalo; Marques, Fernanda; Santos-Silva, Teresa; Santos, Marino F. A.; Hugo, M. Santos E.; Capelo, Jose L.; Doutch, James; Pessoa, Joao CostaVO(acac)(2)] is a remarkable vanadium compound and has potential as a therapeutic drug. It is important to clarify how it is transported in blood, but the reports addressing its binding to serum proteins have been contradictory. We use several spectroscopic and mass spectrometric techniques (ESI and MALDI-TOF), small-angle X-ray scattering and size exclusion chromatography (SEC) to characterize solutions containing [VO(acac)(2)] and either human serum apotransferrin (apoHTF) or albumin (HSA). DFT and modeling protein calculations are carried out to disclose the type of binding to apoHTF. The measured circular dichroism spectra, SEC and MALDI-TOF data clearly prove that at least two VOacac moieties may bind to apoHTF, most probably forming [(VO)-O-IV(acac)(apoHTF)] complexes with residues of the HTF binding sites. No indication of binding of [VO(acac)(2)] to HSA is obtained. We conclude that (VO)-O-IV-acac species may be transported in blood by transferrin. At very low complex concentrations speciation calculations suggest that [(VO)(apoHTF)] species form.
- Naphthoylhydrazones: coordination to metal ions and biological screeningPublication . Ribeiro, Nadia; Galvao, Adelino M.; Gomes, Clara S. B.; Ramos, Helena; Pinheiro, Rute; Saraiva, Lucilia; Ntungwe, Epole; Isca, Vera; Rijo, Patricia; Cavaco, Isabel; Ramilo-Gomes, Filipa; Guedes, Rita C.; Pessoa, Joao Costa; Correia, IsabelWe report the synthesis of 3-hydroxyl-2-naphthoylhydrazones containing pyrrole (HL1), furane (HL2) and thiophene (HL3) moieties and their V(IV)O-, Cu(II)- and Zn(II)-complexes. All compounds are characterized by the usual analytical techniques and coordination of the ligands to the metal ions is discussed based on spectroscopic data (FTIR, UV-vis, EPR and NMR) as well as CAMB3LYP DFT/TDDFT calculations, indicating the formation of neutral ML2 type complexes. The photophysical properties of ligands and complexes are disclosed. The binding to Bovine Serum Albumin (BSA) is evaluated in detail using several spectroscopic tools. Circular dichroism shows that the compounds, and particularly the ligand precursors, stabilize BSA, increasing its a-helical content. Fluorescence studies indicate the formation of 1 : 1 protein-compound adducts, which is corroborated by molecular docking studies that show the interaction between Trp 213 of BSA and the naphthalene rings. The general toxicity is evaluated using the Artemia salina lethality assay, with all compounds showing general toxicity towards the brine shrimp model. The cytotoxicity on human cancer cells (H1299, MCF7, and HCT116) is assessed for all compounds and the half-maximal inhibitory concentration (IC50) values are in the range from 0.57 to 27.35 mu M. Compounds containing L-1 (pyrrole derivative) are the most cytotoxic, with the vanadium and zinc complexes performing better than the copper ones, and some of them depicting IC50 values lower than 1.1 mM. However, selectivity needs improvement as the compounds show toxicity towards Artemia salina and normal fibroblasts.
- New Cu(II) complexes with pyrazolyl derived schiff base ligands: synthesis and biological evaluationPublication . Ribeiro, Nadia; Roy, Somnath; Butenko, Nataliya; Cavaco, Isabel; Pinheiro, Teresa; Alho, Irina; Marques, Fernanda; Avecilla, Fernando; Pessoa, Joao Costa; Correia, IsabelSince the discovery of cisplatin there has been a continuous pursuit for new metallodrugs showing higher efficacies and lower side effects. In this work, new copper(II) complexes (C1-C6) of Schiff bases derived from pyrazolyl were developed. Through condensation of 5-methyl-1H-pyrazole-3-carbohydrazide with different aromatic aldehydes - pyridoxal, salicylaldehyde, 3-methoxy-2-hydroxybenzaldehyde, 3-ethoxy-2-hydroxybenzaldehyde and 2-hydroxynaphthene-l-carbaldehyde a set of new pyrazole based "ONO" tridentate Schiff bases were obtained in moderate to good yields - L1-L6, as well as their Cu(II)-complexes. All compounds were characterized by analytical techniques and their molecular formulae established. The antioxidant potential of all compounds was tested, yielding low activity in most cases, with the exception of L1 and C5. The Cu(II) complexes were tested for their aqueous stability, and for their interaction with biological molecules, namely DNA and HSA (human serum albumin), through fluorescence quenching experiments (and electrophoresis for DNA). With the exception of C3, all the synthesized complexes were able to interact with DNA and HSA. Their cytotoxic activity against two cancer cell lines (MCF7 - breast and PC3 - prostate) was also evaluated. Complexes C5 and C6, with larger aromatic systems, showed much higher cytotoxicity (in the low mu M range), than C1-C4, as well as IC50 values much lower than cisplatin. For C6 the results suggest that the mechanisms of cell death do not seem to be mediated by apoptosis, through caspases 3/7 activation, but by involving membrane potential and imbalance in physiological elements such as P, K and Ca.
- Therapeutic potential of vanadium complexes with 1,10-phenanthroline ligands, quo vadis? Fate of complexes in cell media and cancer cellsPublication . Nunes, Patrique; Correia, Isabel; Cavaco, Isabel; Marques, Fernanda; Pinheiro, Teresa; Avecilla, Fernando; Pessoa, Joao Costa(VO)-O-IV-complexes formulated as [(VO)-O-IV(OSO3)(phen)(2)] (1) (phen = 1,10-phenanthroline), [(VO)-O-IV(OSO3) (Me(2)phen)(2)] (2) (Me(2)phen = 4,7-dimethyl-1,10-phenanthroline) and [(VO)-O-IV(OSO3)(amphen)(2)] (3) (amphen = 5-amino-1,10-phenanthroline) were prepared and stability in cell incubation media evaluated. Their cytotoxicity was determined against the A2780 (ovarian), MCF7 (breast) and PC3 (prostate) human cancer cells at different incubation times. While at 3 and 24 h the cytotoxicity differs for complexes and corresponding free ligands, at 72 h incubation all compounds are equally active presenting low IC50 values. Upon incubation of A2780 cells with 1-3, cellular distribution of vanadium in cytosol, membranes, nucleus and cytoskeleton, indicate that the uptake of V is low, particularly for 1, and that the uptake pattern depends on the ligand. Nuclear microscopic techniques are used for imaging and elemental quantification in whole PC3 cells incubated with 1. Once complexes are added to cell culture media, they decompose, and with time most V-IV oxidizes to V-V-species. Modeling of speciation when [(VO)-O-IV(OSO3)(phen)(2)] (1) is added to cell media is presented. At lower concentrations of 1, (VO)-O-IV- and phen-containing species are mainly bound to bovine serum albumin, while at higher concentrations [(VO)-O-IV (phen)n](2+)-complexes become relevant, being predicted that the species taken up and mechanisms of action operating depend on the total concentration of complex. This study emphasizes that for these (VO)-O-IV-systems, and probably for many others involving oxidovanadium or other labile metal complexes, it is not possible to identify active species or propose mechanisms of cytotoxic action without evaluating speciation occurring in cell media.