Development of Copper Complexes with Diimines and Dipicolinate as Anticancer Cytotoxic Agents.

Coordination complexes may act as anticancer agents. Among others, the formation of the complex may facilitate the ligand uptake by the cell. Searching for new copper compounds with cytotoxic activity, the complex Cu-dipicolinate was studied as a neutral scaffold to form ternary complexes with diimines. A series of [Cu(dipicolinate)(diimine)] complexes (where diimine: Phenanthroline, phen, 5-NO2-phenanthroline, 4-methyl-phenanthroline, neocuproine, 3,4,7,8-tetramethyl-phenanthroline, tmp, bathophenanthroline, bipyridine, dimethyl-bipyridine, as well as the ligand 2,2-dipyridil-amine, bam) were synthesized and characterized both in the solid state, including a new crystal structure of [Cu2(dipicolinate)2(tmp)2]·7H2O. Their chemistry in aqueous solution was explored by UV/vis spectroscopy, conductivity, cyclic voltammetry, and electron paramagnetic resonance studies. Their DNA binding was analyzed by electronic spectroscopy (determining Kb values), circular dichroism, and viscosity methods. The cytotoxicity of the complexes was assessed on human cancer cell lines MDA-MB-231, MCF-7 (breast, the first triple negative), A549 (lung epithelial) and A2780cis (ovarian, Cisplatin-resistant), and non-tumor cell lines MRC-5 (lung) and MCF-10A (breast). The major species are ternary, in solution and solid state. Complexes are highly cytotoxic as compared to Cisplatin. Complexes containing bam and phen are interesting candidates to study their in vivo activity in triple-negative breast cancer treatment.

New Copper(II)-L-Dipeptide-Bathophenanthroline Complexes as Potential Anticancer Agents-Synthesis, Characterization and Cytotoxicity Studies-And Comparative DNA-Binding Study of Related Phen Complexes.

Searching for new copper compounds which may be useful as antitumor drugs, a series of new [Cu(L-dipeptide)(batho)] (batho:4,7-diphenyl-1,10-phenanthroline, L-dipeptide: Gly-Val, Gly-Phe, Ala-Gly, Ala-Ala, Ala-Phe, Phe-Ala, Phe-Val and Phe-Phe) complexes were synthesized and characterized. To interpret the experimental IR spectra, [Cu(ala-gly)(batho)] was modelled in the gas phase using DFT at the B3LYP/LANL2DZ level of theory and the calculated vibrational frequencies were analyzed. Solid-state characterization is in agreement with pentacoordinate complexes of the general formula [Cu(L-dipeptide)(batho)]·x solvent, similar to other [Cu(L-dipeptide)(diimine)] complexes. In solution, the major species are heteroleptic, as in the solid state. The mode of binding to the DNA was evaluated by different techniques, to understand the role of the diimine and the dipeptide. To this end, studies were also performed with complexes [CuCl2(diimine)], [Cu(L-dipeptide)(diimine)] and free diimines, with phenanthroline, neocuproine and 3,4,7,8-tetramethyl-phenanthroline. The cytotoxicity of the complexes was determined on human cancer cell lines MDA-MB-231, MCF-7 (breast, the first triple negative), and A549 (lung epithelial) and non-tumor cell lines MRC-5 (lung) and MCF-10A (breast). [Cu(L-dipeptide)(batho)] complexes are highly cytotoxic as compared to cisplatin and [Cu(L-dipeptide)(phenanthroline)] complexes, being potential candidates to study their in vivo activity in the treatments of aggressive tumors for which there is no curative pharmacological treatment.

Tetramethyl-phenanthroline copper complexes in the development of drugs to treat cancer: synthesis, characterization and cytotoxicity studies of a series of copper(II)-L-dipeptide-3,4,7,8-tetramethyl-phenanthroline complexes.

New compounds to fight cancer are needed due to cancer high incidence and lack of curative treatments for several classes of this disease. Metal-based coordination compounds offer a variety of molecules that can turn into drugs. Among them, coordination copper complexes are emerging as an attractive class of compounds for cancer treatment. A series of [Cu(L-dipeptide)(tmp)] (tmp = 3,4,7,8-tetramethyl-1,10-phenanthroline) complexes were synthesized and characterized in the solid state, including the determination of the crystalline structure of [Cu(Gly-Gly)(tmp)]·3.5 H2O and [Cu2Cl4(tmp)2]. The complexes were studied in solution, where the major species are also ternary ones. The lipophilicity of the complexes was determined and the binding to the DNA was evaluated, suggesting that it occurs in the DNA's major groove. The cytotoxicity of the complexes was evaluated on different cancer cell lines: human metastatic breast adenocarcinoma MDA-MB-231 (triple negative, ATCC: HTB-26), MCF-7 (ATCC: HTB-22), SK-BR-3 (ATCC: HTB-30), human lung epithelial carcinoma A549 (ATCC: CCL-185), cisplatin resistant-human ovarian carcinoma A2780cis (SIGMA) and nontumoral cell lines: MRC-5 (lung; ATCC: CCL-171) and MCF-10A (breast, ATCC: CRL-10317). [Cu(L-dipeptide)(tmp)] complexes are highly cytotoxic as compared to [Cu(L-dipeptide)(phenanthroline)] and cisplatin. Therefore, [Cu(L-dipeptide)(tmp)] complexes are promising candidates to have their in vivo activity further studied toward new treatments for triple negative breast cancer and other aggressive tumors for which there is no curative pharmacological treatment to the date.

Comparative Study of Antioxidant and Pro-Oxidant Properties of Homoleptic and Heteroleptic Copper Complexes with Amino Acids, Dipeptides and 1,10-Phenanthroline: The Quest for Antitumor Compounds.

In a search for new antitumoral agents, a series of homoleptic copper(II) complexes with amino acids and dipeptides, as well as heteroleptic complexes containing both dipeptides and 1,10-phenanthroline, were studied. Furthermore, a single-crystal structure containing alanyl-leucinato ([Cu3(AlaLeu)3(H2O)3(CO3)]·PF6·H2O), which is the first homotrinuclear carbonato-bridged copper(II) complex with a dipeptide moiety, is presented. To assess possible antitumor action mechanisms, we focused on the comparative analysis of pro- and antioxidant behaviors. Pro-oxidant activity, in which the reactive oxygen species (ROS) formed by the reaction of the complexes with H2O2 produce oxidative damage to 2-deoxy-d-ribose, was evaluated using the TBARS method. Additionally, the antioxidant action was quantified through the superoxide dismutase (SOD)-like activity, using a protocol based on the inhibitory effect of SOD on the reduction of nitrobluetetrazolium (NBT) by the superoxide anion generated by the xanthine/xanthine oxidase system. Our findings show that Cu-amino acid complexes are strong ROS producers and moderate SOD mimics. Conversely, Cu-dipeptide-phen complexes are good SOD mimics but poor ROS producers. The activity of Cu-dipeptide complexes was strongly dependent on the dipeptide. A DFT computational analysis revealed that complexes with high SOD-like activity tend to display a large dipole moment and condensed-to-copper charge, softness and LUMO contribution. Moreover, good ROS producers have higher global hardness and copper electrophilicity, lower copper softness and flexible and freely accessible coordination polyhedra.

Electrochemical, mechanistic, and DFT studies of amine derived diphosphines containing Ru(II)-cymene complexes with potent in vitro cytotoxic activity against HeLa and triple-negative breast cancer cells MDA-MB-231.

Complexes with general formula [RuCl(η6-p-cymene)(P-NR-P)]X (R = CH2Py (Py = pyridine) - [1a]+, CH2Ph (Ph = phenyl) - [1b]+, Ph - [1c] and p-tol (p-tol = p-tolyl) - [1d]+; X = PF6- or BF4-) were evaluated as cytotoxic agents against two cancer cell lines (HeLa and MDA-MB-231). All metal complexes are active in the range of concentrations tested (up to 100 µmol L-1). The IC50 (µmol L-1) values for the metal complexes are lower than that found for cisplatin. The activities are up to 6- and 15-fold higher than cisplatin for HeLa and MDA-MB-231 cancer cell lines, respectively. Studies of DNA binding and DNA cleavage were performed. DNA binding studies revealed a modest hypochromic shift in the metal complexes electronic spectra, indicating a weak interaction with Kb values in the range of 1.7 × 103-1.6 × 104. Although the cleavage tests revealed that in the dark DNA is not a biological target for these metal complexes, upon blue light irradiation they are activated causing DNA cleavage. Electrochemical studies showed the presence of two independent redox processes, one attributed to the oxidation process of Ru2+ → Ru3+ (EC process) and the other one to the reduction of Ru2+ → Ru1+, which is further reduced to Ru0 (ECE mechanism). In both processes, coupled chemical reactions were observed. DFT calculations were performed to support the electrochemical/chemical behavior of the complexes. The reactivity of complex [1b]BF4 with CH3CN was evaluated and two complexes were isolated [2b]BF4 and [3b]BF4. The complex mer-[RuCl(CH3CN)3(P-NCH2Ph-P)]BF4 ([2b]BF4) was isolated after refluxing the precursor [1b]BF4 in CH3CN. Isomerization of [2b]BF4 in CH3CN resulted in the formation of fac-[RuCl(CH3CN)3(P-NCH2Ph-P)]BF4. An attempt to isolate the fac-isomer by adding diethyl ether was unsuccessful, and the complex [3b]BF4 was observed as the major component. The complex [Ru2(µ-Cl3)(CH3CN)2(P-NCH2Ph-P)2]BF4 ([3b]BF4) proved to be very stable and can be obtained from both the mer- and the fac-isomers. The molecular structures of [1b]BF4 and [3b]BF4 were solved by single-crystal X-ray diffraction.

Synthesis and structural characterization of a series of ternary copper(II)-L-dipeptide-neocuproine complexes. Study of their cytotoxicity against cancer cells including MDA-MB-231, triple negative breast cancer cells.

This work presents the synthesis and characterization of eight copper complexes [Cu(L-dipeptide)(neo)]·nH2O (neo = neocuproine) and their cytotoxic activities against tumor cell lines. The crystalline structure of [Cu(gly-val)(neo)]·3H2O, [Cu(gly-leu)(neo)]·H2O, [Cu(ala-gly)(neo)]·4H2O, [Cu(val-phe)(neo)]·4.5H2O and [Cu(phe-phe)(neo)]·3H2O were determined by single crystal X-ray diffraction. In all of them, the Cu(II) is pentacoordinated, in a square pyramidal environment. The coordination observed in solid state was retained in the major species in aqueous solution, as suggested by Electronic Paramagnet Resonance and UV-vis spectroscopies. The complexes were shown to have affinity for isolated DNA, as determined by Circular Dichroism experiments. Furthermore, biological experiments showed that all the complexes present high cytotoxic activity against the cell lines: MDA-MB-231, MCF-7 (human metastatic breast adenocarcinomas, the first triple negative), MCF-10A (human normal breast cells), A549 (human lung epithelial carcinoma) and MRC-5 (human lung epithelial cells). Together, these results suggest that these compounds are promising steps towards new effective drugs to treat cancer.

Experimental and theoretical studies of copper complexes with isomeric dipeptides as novel candidates against breast cancer.

In the search for new cytotoxic drugs, two copper complexes with isomeric dipeptides (Ala-Phe and Phe-Ala) were developed in order to determine the influence of their different structures in the modulation of the chemical, biochemical and biological properties. Spectroscopic, voltammetric and equilibrium studies were performed providing information about the chemical properties. The superoxide dismutase (SOD) activity was studied and showed differences of IC50 for both Cu-Ala-Phe (IC50=4.5) and Cu-Phe-Ala (IC50=45). The computational results permitted to explain this behavior proposing that it is feasible that the O2- anion is attracted straight to the positive zone in Cu-Ala-Phe whereas for Cu-Phe-Ala this phenomenon would happen to a smaller extent. Confirming our previous studies, both complexes interacted with DNA. Molecular docking studies showed that the position of the phenyl ring modulates the complex-DNA affinity and in Cu-Ala-Phe the docked conformation allows the copper ion to face the DNA basis, giving rise to a more stable complex-DNA adduct than for Cu-Phe-Ala. In spite of the fact that Atomic Force Microscopy showed plasmid compactation and aggregation for both complexes, the image showed softer changes in the case of Cu-Ala-Phe in comparison with those produced by Cu-Phe-Ala. In order to evaluate the effect of Cu-Ala-Phe and Cu-Phe-Ala complexes against tumor cells, we have employed three aggressive metastatic breast adenocarcinoma cellular models, derived from human (MDA-MB-231 and MCF-7) and mouse (4T1) spontaneous tumors. These experiments showed that both Cu-dipeptide complexes have a similar cytotoxic effect against breast cancer cells, and lower toxicity against BJ non-tumor cells in comparison to Cisplatin.

Synthesis, structural characterization and cytotoxic activity of ternary copper(II)-dipeptide-phenanthroline complexes. A step towards the development of new copper compounds for the treatment of cancer.

In the search for new compounds with antitumor activity, coordination complexes with different metals are being studied by our group. This work presents the synthesis and characterization of six copper complexes with general stoichiometry [Cu(L-dipeptide)(phen)]·nH2O (were phen=1,10-phenanthroline) and their cytotoxic activities against tumor cell lines. To characterize these systems, analytical and spectroscopic studies were performed in solid state (by UV-visible, IR, X-ray diffraction) including the crystal structure of four new complexes (of the six complexes studied): [Cu(Ala-Phe)(phen)]·4H2O, [Cu(Phe-Ala)(phen)]·4H2O, [Cu(Phe-Val)(phen)]·4.5H2O and [Cu(Phe-Phe)(phen)]·3H2O. In all of them, the copper ion is situated in a distorted squared pyramidal environment. The phen ligand is perpendicular to the dipeptide, therefore exposed and potentially available for interaction with biological molecules. In addition, for all the studied complexes, structural information in solution using EPR and UV-visible spectroscopies were obtained, showing that the coordination observed in solid state is maintained. The lipophilicity, DNA binding and albumin interaction were also studied. Biological experiments showed that all the complexes induce cell death in the cell lines: HeLa (human cervical adenocarcinoma), MCF-7 (human metastatic breast adenocarcinoma) and A549 (human lung epithelial carcinoma). Among the six complexes, [Cu(Ala-Phe)(phen)] presents the lowest IC50 values. Taken together all these data we hypothesize that [Cu(Ala-Phe)(phen)] may be a good candidate for further studies in vivo.

High-dose selenium reduces ventilator-associated pneumonia and illness severity in critically ill patients with systemic inflammation.

PURPOSE: To confirm the pharmacodynamics and evaluate the efficacy of high-dose selenium (Se) administered by continuous infusion, following an initial loading bolus of selenite, on clinical outcome in critically ill patients with systemic inflammatory response syndrome (SIRS). METHODS: Prospective, placebo-controlled, randomized, single-blinded phase II study in a multidisciplinary university hospital intensive care unit (ICU). Two groups of patients with SIRS, age >18 years, and Acute Physiology and Chronic Health Evaluation (APACHE) II ≥15 (n = 35) were randomized to receive either placebo or intravenous selenite as a bolus-loading dose of 2,000 µg Se followed by continuous infusion of 1,600 µg Se per day for 10 days. Blood samples were analyzed before randomization (day 0) then at days 3, 7, and 10. Clinical outcome was assessed by Sequential Organ Failure Assessment (SOFA) score. Hospital-acquired pneumonia including ventilator-associated pneumonia (VAP), adverse events, and other safety parameters were monitored as secondary endpoints. RESULTS: SOFA score decreased significantly in the selenite group at day 10 (1.3 ± 1.2 versus 4.6 ± 2.0, p = 0.0001). Early VAP rate was lower in the selenite group (6.7% versus 37.5%, p = 0.04), and hospital-acquired pneumonia was lower after ICU discharge (p = 0.03). Glutathione peroxidase-3 (GPx-3) activity increased in both groups, reaching a maximum at day 7 (0.62 ± 0.24 versus 0.28 ± 0.14 U/mL, p = 0.001) in the selenite group. No adverse events attributable to selenite were observed. CONCLUSIONS: Daily infusion of 1,600 µg Se (as selenite), following an initial bolus of 2,000 µg, is novel and without short-term adverse events. High-dose parenteral selenite significantly increases Se status, improves illness severity, and lowers incidence of hospital-acquired pneumonia including early VAP for SIRS patients in ICU.

High-dose selenium for critically ill patients with systemic inflammation: pharmacokinetics and pharmacodynamics of selenious acid: a pilot study.

OBJECTIVE: Systemic inflammatory response syndrome is characterized by increased urinary excretion of selenium and low serum concentration. Repletion by parenteral selenite is the most efficacious form of supplementation. However, the optimum safe dose and mode of administration remain controversial. We aimed to determine pharmacokinetic and pharmacodynamic profiles of selenite and estimate a safe dose to optimize selenium status. METHODS: A prospective, randomized, pilot study in 20 patients with systemic inflammatory response syndrome compared a high-dose (HD) group that received a loading dose of selenium as selenite 15.18 micromol over 2 h and thereafter 10.12 micromol/d as a continuous intravenous infusion (CIV) for 10 d with a very-high-dose (VHD) group that received a loading dose of 25.30 micromol over 2 h and thereafter 20.24 micromol as a CIV for 10 d. Clinical outcome was evaluated by length of stay in the intensive care unit, incidence of ventilator-associated pneumonia, and Sequential Organ Failure Assessment score. RESULTS: Patients in group HD (n = 10, age 54 +/- 23 y) had an Acute Physiology and Chronic Health Evaluation II score of 23 +/- 5 and a Sequential Organ Function Assessment score of 10 +/- 2. Those in group VHD (n = 10, age 41 +/- 19 y) had scores of 21 +/- 7 and 8 +/- 3, respectively. Pharmacokinetic concentration/time curves for serum selenium overlapped but were independent of dose, whereas the pharmacodynamics were different, showing maximum glutathione peroxidase activity only with VHD. Glutathione peroxidase decreased after day 7 independently of the selenium dose. Clinical outcomes were similar in both groups. CONCLUSION: A bolus loading dose of selenite providing 2000 microg of selenium (25.30 micromol) followed by a CIV of 1600 microg/d (20.24 micromol/d) for 10 d is most effective at returning serum selenium to physiologic levels and safely maximizing glutathione peroxidase activity.

Serum selenium and glutathione peroxidase-3 activity: biomarkers of systemic inflammation in the critically ill?

OBJECTIVES: To confirm the influence of systemic inflammatory response syndrome (SIRS) on selenium (Se) levels and prospectively evaluate the relationship between serum Se concentration [Se], glutathione peroxidase activity [GPx-3] and injury severity in patients at the time of intensive care unit (ICU) admission. DESIGN: Prospective, observational study. SETTING: Multidisciplinary University Hospital ICU. PATIENTS AND PARTICIPANTS: A total of 36 ICU patients and 23 healthy volunteer subjects (HVS). MEASUREMENTS AND RESULTS: Healthy volunteer subjects were designated as controls (Group 1). ICU patients were divided into three groups: without SIRS (Group 2); with SIRS (Group 3); with SIRS and multiple organ dysfunction syndrome (MODS) (Group 4). The latter groups had APACHE II scores >15. [GPx-3] and [Se] were determined by standard methods within the first 48 h of admission to ICU. Kruskal-Wallis and Mann-Whitney U test were used for analysis of non-parametric continuous variables. The predictive value of [Se] and [GPx-3] for SIRS was calculated using a receiver operating characteristics (ROC) analysis. In SIRS and MODS patients [GPx-3] and [Se] decreased significantly (P = 0.0001 and P = 0.002, respectively). After ICU admission [GPx-3] and [Se] had a predictive value for SIRS ([GPx-3] sensitivity: 90%, specificity: 86.2% (cut-off value: 0.5 U/mL); [Se]: sensitivity 90%, specificity 72.4% (cut-off value: 60 microg/L). [Se] had predictive value for ICU mortality (P = 0.034). CONCLUSIONS: Systemic inflammatory response syndrome and MODS were associated with early decreases in [Se] and [GPx-3]. Low [Se] and [GPx-3] after ICU admission had a predictive value for SIRS, which may aid future selection of patients who could benefit from Se supplementation.

Weak exchange interaction supported by a biologically relevant long chemical bridge in a Cu-peptide model compound.

The copper complex of the dipeptide L-alanyl-L-phenylalanine, catena-(L-alaninate-L-phenylalaninate-copper(II) monohydrate), identified as Cu(II)Ala-Phe, provides a convenient system to study a weak exchange interaction between unpaired spins transmitted through a biologically relevant long chemical bridge (18.34 A). In this complex, the copper ions are arranged in two symmetry-related anisotropic layers parallel to the ab plane at 13.17 A, separated by a double layer of water molecules. The equatorial-equatorial bridge considered as the most relevant path for exchange interactions between copper ions in neighbor layers contains 11 diamagnetic atoms (including three hydrogens), with two covalent amidate bridges plus three weak and moderate H bonds that go across the water layer. This interaction was studied using electron paramagnetic resonance in single-crystal samples, at 9.5 and 34.5 GHz. The measured magnitude of the interlayer interaction, |J3|/kB = 1.7(2) x 10(-3) K, is discussed in terms of values obtained for similar paths in other model compounds and in proteins. These results in model systems provide information that may be important in understanding biological functions at the molecular level.

Characterization of a Cu(II) complex of sulfadimethoxine.

The molecular structure of [Cu(sulfadimet)(2)].SO(CH(3))(2) (sulfadimet=sulfadimethoxine=4-p-aminobenzenesulfonamido-2,6-dimethoxypyrimidine) was determined by single crystal X-ray diffractometry. It crystallizes in the monoclinic space group P2(1)/c with Z=4. The Cu(II) cation is in a distorted CuN(5) square pyramidal coordination, involving four sulfadimethoxine molecules, one of them acting as a bidentate ligand. The infrared spectrum is briefly discussed on the basis of the structural peculiarities of the complex.

Synthesis and characterization of three new Cu(II)-dipeptide complexes.

Three new copper(II) complexes of stoichiometry [Cu(L-dipeptide)].nH(2)O, containing as ligands the dipeptides L-alanine-L-isoleucine, L-alanine-L-threonine and L-alanine-L-tyrosine were prepared. They were characterized by single crystal X-ray diffractometry, and electronic and infrared spectroscopy. In all cases, the Cu(II) cation has essentially the same elongated square pyramidal coordination, being equatorially cis coordinated by a N(2)O(2) arrangement of ligand atoms and axially by a carbonyl oxygen atom. The compounds show rather similar polymeric structures which resemble those recently reported for the [Cu(ala-val)] and [Cu(ala-phe)] complexes. The electronic and infrared spectra are briefly discussed on the basis of the structural peculiarities of the complexes. Superoxide dismutase (SOD)-like activity was also tested for the compounds.