Erika Ferrari

Curcuminoids as potential new iron-chelating agents: spectroscopic, polarographic and potentiometric study on their Fe(III) complexing ability

The pKa values of curcumin and diacetylcurcumin are, here doubtless, determined by means of spectroscopic and potentiometric measurements, and the enolic proton is the more acidic one. The interaction of Fe 3 + with curcumin and diacetylcurcumin, in water/methanol 1:1 solution, leads to the formation of the complex species [FeH2CU(OH)2] and [FeDCU(OH)2 ]( H 2CU and DCU=curcumin or diacetylcurcumin monoanion, respectively) which prevails near pH 7. At more basic condition the prevailing species are [FeH2CU(OH)3] − and [FeDCU(OH)3] − , which prevent metal hydroxide precipitation. 1 H NMR data state that the dissociated -diketo moiety of the ligands is involved in metal chelation. The pKa value of the deprotonation reaction is strongly anticipated by the metal ion, as shown by UV spectral data. The stability constants, evaluated from potentiometric data, are near to that of desferrioxamine, which is, by now, the only iron-chelating agent for clinical use.

Curcumin derivatives: Molecular basis of their anti-cancer activity

Curcumin, a phenolic compound from the plant Curcuma longa L., has shown a wide-spectrum of chemopreventive, antioxidant and antitumor properties. Although its promising chemotherapeutic activity, preclinical and clinical studies highlight Curcumin limited therapeutic application due to its instability in physiological conditions. To improve its stability and activity, many derivatives have been synthesized and studied, among which bis-DemethoxyCurcumin (bDMC) and diAcetylCurcumin (DAC). In this report, we show that both bDMC and DAC are more stable than Curcumin in physiological medium. To explore the mechanism of their chemotherapeutic effect, we studied their role in proliferation in the HCT116 human colon cancer cells. We correlated kinetic stability and cellular uptake data to their biological effects. Both bDMC and DAC impair correct spindles formation and induce a p53- and p21(CIP1/WAF1)-independent mitotic arrest, which is more stable and long-lasting for bDMC. A subsequent p53/p21(CIP1/WAF1)-dependent inhibition of G1 to S transition is triggered by Curcumin and DAC as a consequence of the mitotic slippage, preventing post-mitotic cells from re-entering the cell cycle. Conversely, the G1/S arrest induced by bDMC is a direct effect of the drug and concomitant to the mitotic block. Finally, we demonstrate that bDMC induces rapid DNA double-strand breaks, moving for its possible development in anti-cancer clinical applications.

Synthesis, cytotoxic and combined cDDP activity of new stable curcumin derivatives

New curcumin derivatives are synthesized in order to improve chemical properties of curcumin. The aromatic ring glycosylation of curcumin provides more water-soluble compounds with a greater kinetic stability which is a fundamental feature for drug bioavailability. The glycosylation reaction is quite simple, low cost, with high yield and minimum waste. NMR data show that the ability of curcumin to coordinate metal ion, in particular Ga(III), is maintained in the synthesized products. Although the binding of glucose to curcumin reduces the cytotoxicity of the derivatives towards cisplatin (cDDP)-sensitive and -resistant human ovarian carcinoma cell lines, the compounds display a good selectivity since they are much less toxic against non-tumourigenic Vero cells. The combination of cDDP with the most active glycosyl-curcuminoid drug against both cDDP-sensitive and -resistant as well as against Vero cell lines is tested. The results show an improvement of cDDP efficacy with higher selectivity towards cancer cells than non-cancer cells. These studies indicate the need for developing new valid components of drug treatment protocols to cDDP-resistant cells as well.

Newly synthesized curcumin derivatives: crosstalk between chemico-physical properties and biological activity.

New curcumin analogues (ester and acid series) were synthesized with the aim to improve the chemical stability in physiological conditions and potential anticancer activity. Cytotoxicity against different tumorigenic cell lines (human ovarian carcinoma cells -2008, A2780, C13*, and A2780/CP, and human colon carcinoma cells HCT116 and LoVo) was tested to evaluate cellular specificity and activity. Physico-chemical properties such as acidity, lipophilicity, kinetic stability, and free radical scavenging activity were investigated to shed light on the structure-activity relationship and provide new attractive candidates for drug development. Most of ester derivatives show IC(50) values lower than curcumin and exhibit selectivity against colon carcinoma cells. Especially they are extremely active after 24 h exposure showing enhanced inhibitory effect on cell viability. The best performances of ester curcuminoids could be ascribed to their high lipophilicity that favors a greater and faster cellular uptake overcoming their apparently higher instability in physiological condition.

bis-Dehydroxy-Curcumin Triggers Mitochondrial-Associated Cell Death in Human Colon Cancer Cells through ER-Stress Induced Autophagy

Background The activation of autophagy has been extensively described as a pro-survival strategy, which helps to keep cells alive following deprivation of nutrients/growth factors and other stressful cellular conditions. In addition to cytoprotective effects, autophagy can accompany cell death. Autophagic vacuoles can be observed before or during cell death, but the role of autophagy in the death process is still controversial. A complex interplay between autophagy and apoptosis has come to light, taking into account that numerous genes, such as p53 and Bcl-2 family members, are shared between these two pathways. Methodology/Principal Findings In this study we showed a potent and irreversible cytotoxic activity of the stable Curcumin derivative bis-DeHydroxyCurcumin (bDHC) on human colon cancer cells, but not on human normal cells. Autophagy is elicited by bDHC before cell death as demonstrated by increased autophagosome formation -measured by electron microscopy, fluorescent LC3 puncta and LC3 lipidation- and autophagic flux -measured by interfering LC3-II turnover. The accumulation of poly-ubiquitinated proteins and ER-stress occurred upstream of autophagy induction and resulted in cell death. Cell cycle and Western blot analyses highlighted the activation of a mitochondrial-dependent apoptosis, which involves caspase 7, 8, 9 and Cytochrome C release. Using pharmacological inhibitions and RNAi experiments, we showed that ER-stress induced autophagy has a major role in triggering bDHC-cell death. Conclusion/Significance Our findings describe the mechanism through which bDHC promotes tumor selective inhibition of proliferation, providing unequivocal evidence of the role of autophagy in contrasting the proliferation of colon cancer cells.

Synthesis and Characterization of 68Ga-Labeled Curcumin and Curcuminoid Complexes as Potential Radiotracers for Imaging of Cancer and Alzheimer’s Disease

Curcumin (CUR) and curcuminoids complexes labeled with fluorine-18 or technetium-99m have recently shown their potential as diagnostic tools for Alzheimers disease. Gallium-68 is a positron-emitting, generator-produced radionuclide, and its properties can be exploited in situ in medical facilities without a cyclotron. Moreover, CUR showed a higher uptake in tumor cells compared to normal cells, suggesting potential diagnostic applications in this field. In spite of this, no studies using labeled CUR have been performed in this direction, so far. Herein, (68)Ga-labeled complexes with CUR and two curcuminoids, namely diacetyl-curcumin (DAC) and bis(dehydroxy)curcumin (bDHC), were synthesized and characterized by means of experimental and theoretical approaches. Moreover, a first evaluation of their affinity to synthetic β-amyloid fibrils and uptake by A549 lung cancer cells was performed to show the potential application of these new labeled curcuminoids in these diagnostic fields. The radiotracers were prepared by reacting (68)Ga(3+) obtained from a (68)Ge/(68)Ga generator with 1 mg/mL curcuminoids solutions. Reaction parameters (precursor amount, reaction temperature, and pH) were optimized to obtain high and reproducible radiochemical yield and purity. Stoichiometry and formation of the curcuminoid complexes were investigated by matrix-assisted laser desorption ionization time-of-flight mass spectrometry, NMR, ultraviolet-visible, and fluorescence spectroscopy on the equivalent (nat)Ga-curcuminoids (nat = natural) complexes, and their structure was computed by theoretical density functional theory calculations. The analyses evidenced that CUR, DAC, and bDHC were predominantly in the keto-enol form and attested to Ga(L)2(+) species formation. Identity of the (68)Ga(L)2(+) complexes was confirmed by coelution with the equivalent (nat)Ga(L)2(+) complexes in ultrahigh-performance liquid chromatography analyses.(68)Ga(CUR)2(+), (68)Ga(DAC)2(+), and (68)Ga(bDHC)2(+) were highly (87 ± 4, 90 ± 1%) and moderately (48 ± 2%), respectively, retained by synthetic β-amyloid fibrils in vitro. All the Ga-curcuminoid complexes showed an uptake in A549 lung cancer cells, at least equivalent to the respective free curcuminoids, confirming potential applications as cancer-detecting radiotracers.

Sugar complexes with metal2+ ions: thermodynamic parameters of associations of Ca2+, Mg2+ and Zn2+ with galactaric acid

Abstract A solution study on the complex ability of galactaric acid (GalaH 2 ) for complexation with Ca 2+ , Mg 2+ and Zn 2+ ions is reported. The stability constants of the complex species are determined by means of potentiometric measurements. From the dependence of stability constants on temperature, the Δ H ° and Δ S ° values are also determined. The formation of the complex species is an endothermic process and Δ H ° and Δ S ° values suggest a chelate coordination mode of galactaric acid involving carboxylic oxygen and α-hydroxylic group. The prevailing species at acidic or neutral pH is [MGala] (M=Ca 2+ , Mg 2+ , Zn 2+ ) which is also isolated in the solid state and characterised by means of IR spectroscopy. On increasing pH, the [MGalaH −1 ] − species is also formed, where the coordinated OH group undergoes deprotonation.

Solvent effect on keto–enol tautomerism in a new β-diketone: a comparison between experimental data and different theoretical approaches

The novel β-diketo compound (3-acetyl-4-oxopentanoic acid) OPAA is here synthesized and completely characterized in the solid state by means of X-ray crystallography and in solution by potentiometry and 1H and 13C NMR spectroscopy. In the solid state, OPAA exhibits the di-keto (DK) structure, however, in solution, we can observe a strong solvent dependent tautomeric equilibrium. Theoretical ab initio calculations employing DFT at the B3LYP/6-311G** level, and different methods of theoretical model chemistry (CBS-4M, G3MP2, CBS-QB3) are used to extensively investigate the tautomeric equilibrium and compare it with experimental data. Solvent effects are evaluated using a CPCM continuum solvation method; among all applied methods, CBS-4M is the one that better predicts experimental data and is able to qualitatively describe tautomeric equilibrium in solution, allowing thermodynamic calculations of pKa. Furthermore a supermolecular solvent approach is used to better analyze solvent–solute interactions in order to predict chemical properties.

Curcumin derivatives as metal-chelating agents with potential multifunctional activity for pharmaceutical applications.

Curcuminoids represent new perspectives for the development of novel therapeutics for Alzheimers disease (AD), one probable mechanism of action is related to their metal complexing ability. In this work we examined the metal complexing ability of substituted curcuminoids to propose new chelating molecules with biological properties comparable with curcumin but with improved stability as new potential AD therapeutic agents. The K2T derivatives originate from the insertion of a -CH2COOC(CH3)3 group on the central atom of the diketonic moiety of curcumin. They retain the diketo-ketoenol tautomerism which is solvent dependent. In aqueous solution the prevalent form is the diketo one but the addition of metal ion (Ga(3+), Cu(2+)) causes the dissociation of the enolic proton creating chelate complexes and shifting the tautomeric equilibrium towards the keto-enol form. The formation of metal complexes is followed by both NMR and UV-vis spectroscopy. The density functional theory (DFT) calculations on K2T21 complexes with Ga(3+) and Cu(2+) are performed and compared with those on curcumin complexes. [Ga(K2T21)2(H2O)2](+) was found more stable than curcumin one. Good agreement is detected between calculated and experimental (1)H and (13)C NMR data. The calculated OH bond dissociation energy (BDE) and the OH proton dissociation enthalpy (PDE), allowed to predict the radical scavenging ability of the metal ion complexed with K2T21, while the calculated electronic affinity (EA) and ionization potential (IP) represent yardsticks of antioxidant properties. Eventually theoretical calculations suggest that the proton-transfer-associated superoxide-scavenging activity is enhanced after binding metal ions, and that Ga(3+) complexes display possible superoxide dismutase (SOD)-like activity.

Metal binding ability of curcumin derivatives: a theoretical vs. experimental approach

Theoretical calculations employing DFT at the B3LYP/6-311G++** level are used to investigate the tautomeric equilibrium in curcumin derivatives. The solvent effect is evaluated using the CPCM continuum solvation method. The results are compared with experimental data obtained from the X-ray crystal structure of K2A23 and UV-vis data. The KE tautomer is more stable in a vacuum and in the solid state, while in water the DK tautomer reaches a population of 90%. In agreement with spectroscopic data, theoretical calculations predict a slight prevalence of the DK form in non-aqueous solvent systems. The ability to chelate metal ions [Fe(3+), Ga(3+) and Cu(2+)] is then explored by means of (1)H, (13)C NMR and UV-Vis spectroscopy. From the calculation of the overall stability constants of metal complexes and (1)H NMR titrations with Ga(3+), it is clear that the more stable species has a 1 : 2 M/L molar ratio. The curcuminoid coordinates the metal ion through the keto-enol function in the dissociated form; in addition 2D (1)H (13)C NMR experiments suggest the involvement of carboxylic oxygen in metal coordination it was found in the solid state for the complex [Ga(K2A33)2]PF6. The rate of the complexation reaction is strongly influenced by the type of substituent on the aromatic ring of the curcuminoid (K2A33 ≈ K2A23 ≫ K2A21). In addition DPPH assay evidences how antioxidant ability of curcumin derivatives is mainly due to the presence of a phenolic group and metal coordination by a keto-enolic moiety does not affect it, especially for K2A21.