Carlotta Compari

From Ligand to Complexes. Part 2. Remarks on Human Immunodeficiency Virus type 1 Integrase Inhibition by β-Diketo Acid Metal Complexes

Previously, we synthesized a series of beta-diketo acid metal complexes as novel HIV-1 integrase (IN) inhibitors (J. Med. Chem. 2006, 46, 4248-4260). Herein, a further extension of this study is reported. First, detailed docking studies were performed in order to investigate the mode of binding in the active site of the free ligands and of their metal complexes. Second, a series of potentiometric measurements were conducted for two diketo acids chosen as model ligands, with Mn(2+) and Ca(2+), in order to outline a speciation model. Third, we designed and synthesized a new set of complexes with different stoichiometries and tested them in an in vitro assay specific for IN. Finally, we obtained the first X-ray structure of a metal complex with HIV-1 IN inhibition activity. Analysis of these results supports the hypothesis that the diketo acids could act as complexes and form complexes with the metal ions on the active site of the enzyme.

HIV-1 in strand transfer chelating inhibitors: A focus on metal binding

Most active and selective strand transfer HIV-1 integrase (IN) inhibitors contain chelating functional groups that are crucial feature for the inhibition of the catalytic activities of the enzyme. In particular, diketo acids and their derivatives can coordinate one or two metal ions within the catalytic core of the enzyme. The present work is intended as a contribution to elucidate the mechanism of action of the HIV-IN inhibitors by studying the coordinative features of H₂L¹ (L-708,906), an important member of the diketo acids family of inhibitors, and H₂L₂, a model for S-1360, another potent IN inhibitor. Magnesium(II) and manganese(II) complexes of H₂L¹ and H₂L² were isolated and fully characterized in solution and in the solid state. The crystal structures of the manganese complex [Mn(HL₂)₂(CH₃OH)₂]·2CH₃OH were solved by X-ray diffraction analysis. Moreover, the speciation models for H₂L₂ with magnesium(II) and manganese(II) ions were performed and the formation constants of the complexes were measured. M(HL₂)₂ (M = Mg²+, Mn²+) was the most abundant species in solution at physiological pH. All the synthesized compounds were tested for their anti-IN activity, showing good results both for the ligand and the corresponding complexes. From analysis of the speciation models and of the biological data we can conclude that coordination of both metal cofactors could not be strictly necessary and that inhibitors can act as complexes and not only as free ligands.

Synthesis and properties of cationic surfactants with tuned hydrophylicity

A series of pyridinium-based cationic surfactants has been synthesised and their amphiphilic properties have been studied by conductivity and surface tension measurements. The modification of the substitution pattern on the pyridinium ring by hydrophobic moieties (methyl vs. hydrogen and presence or not of condensed benzene ring) gave the opportunity to investigate structure-activity relationships. Characterization by conductivity and surface tension measurements shed light on the behaviour at the air/water interface and in the micellar environment. In particular, the tendency to form ion pairs at very low concentration was evidenced for all the surfactants substituted on the ring, but not for the simple pyridinium ones. The formation of ion pairs affects both the conductivity and the surface tension plots, showing that a series of steps is involved during the adsorption to the air/water surface. An attempt was made to qualify the single steps in the adsorption at the surface layer. Those steps were attributed to different chemical species (free surfactant ions or ion pairs) and to different arrangements of the surfactant. This work also represents a contribution of investigation at very low surfactant concentrations and high surface tension values.

Investigating the Role of Metal Chelation in HIV-1 Integrase Strand Transfer Inhibitors

HIV-1 integrase (IN) has been validated as an attractive target for the treatment of HIV/AIDS. Several studies have confirmed that the metal binding function is a crucial feature in many of the reported IN inhibitors. To provide new insights on the metal chelating mechanism of IN inhibitors, we prepared a series of metal complexes of two ligands (HL1 and HL2), designed as representative models of the clinically used compounds raltegravir and elvitegravir. Potentiometric measurements were conducted for HL2 in the presence of Mg(II), Mn(II), Co(II), and Zn(II) in order to delineate a metal speciation model. We also determined the X-ray structures of both of the ligands and of three representative metal complexes. Our results support the hypothesis that several selective strand transfer inhibitors preferentially chelate one cation in solution and that the metal complexes can interact with the active site of the enzyme.

A versatile salicyl hydrazonic ligand and its metal complexes as antiviral agents

Acylhydrazones are very versatile ligands and their coordination properties can be easily tuned, giving rise to metal complexes with different nuclearities. In the last few years, we have been looking for new pharmacophores able to coordinate simultaneously two metal ions, because many enzymes have two metal ions in the active site and their coordination can be a successful strategy to inhibit the activity of the metalloenzyme. As a part of this ongoing research, we synthesized the acylhydrazone H2L and its complexes with Mg(II), Mn(II), Co(II), Ni(II), Cu(II) and Zn(II). Their characterization, both in solution--also by means of potentiometric studies--and in the solid state, evidenced the ability of the o-vanillin hydrazone scaffold to give rise to different types of metal complexes, depending on the metal and the reaction conditions. Furthermore, we evaluated both the free ligand and its metal complexes in in vitro studies against a panel of diverse DNA- and RNA-viruses. In particular, the Mg(II), Mn(II), Ni(II) and Zn(II) complexes had EC50 values in the low micromolar range, with a pronounced activity against vaccinia virus.

Thermodynamics and biological properties of the aqueous solutions of new glucocationic surfactants.

Thermodynamic properties of aqueous solutions of newly synthesized compounds, namely, N-[2-(beta-D-glucopyranosyl)ethyl]-N,N-dimethyl-N-alkylammonium bromides with hydrophobic tails of 12 (C12DGCB) and 16 (C16DGCB) carbon atoms, determined as a function of concentration by means of direct methods, are reported here. Dilution enthalpies, densities, and sound velocities were measured at 298 K, allowing for the determination of apparent and partial molar enthalpies, volumes, and compressibilities. Changes in thermodynamic quantities upon micellization were derived using a pseudophase-transition approach. From a comparison with the corresponding acetylated compounds N-[2-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)ethyl]-N,N-dimethyl-N-dodecylammonium bromide (C12AGCB) and N-[2-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyanosyl)ethyl]-N,N-dimethyl-N-hexadecylammonium bromide (C16AGCB), the role played in the micellization process by the acetylated glycosyl moiety was inferred: it enhances the hydrophobic character of the molecule and lowers the change in enthalpy of micelle formation by about 1.5 kJ mol(-1). By comparing the volume of C12DGCB with those of DEDAB and DTAB, the volumes taken up by the (beta- d-glucopyranosyl)ethyl and beta- d-glucopyranosyl groups were found to be 133 and 99 cm3 mol(-1), respectively. Regarding the interaction with DPPC membranes, it seems that the sugar moiety of the hexadecyl deacetylated compound gives rise to hydrogen bonds with the oxygen atoms of the lipid phosphates, shifting the phase transition of DPPC from a bilayer gel to a bilayer liquid crystal to lower temperatures. C16AGCB induces significantly greater changes than C16DGCB in the structure of liposomes, suggesting the formation of domains. The interaction is strongly enhanced by the presence of water. Neither compound interacts strongly with DNA or compacts it, as shown by EMSA assays and AFM images. Only C16AGCB is able to deliver little DNA inside cells when coformulated with DOPE, as shown by the transient transfection assay. This might be related to the ability of C16AGCB to form surfactant-rich domains in the lipid structure.

Diketoacid chelating ligands as dual inhibitors of HIV-1 integration process

HIV-1 Integrase (IN) represents a very attractive pharmacological target for the development of new and more efficient drugs. Recently, an allosteric inhibitory approach also emerged, that targets the interaction between IN and cellular cofactors, such as LEDGF/p75. Small molecules based on the diketoacid pharmachophore were studied for their ability to inhibit at the same time integration and IN-LEDGF/p75 interaction (dual inhibitors): in this study, we evaluated three indole diketoacid derivatives and their magnesium(II) complexes for their ability to act as dual inhibitors. Effectively, the metal complexes exhibited IN inhibition potency in low nanomolar/micromolar concentration range; both the complexes and the free ligands are also able to inhibit the IN-LEDGF/p75 interaction at low μM values. Moreover, these magnesium compounds showed good antiviral activity, suggesting the possibility to exploit metal coordination for the design of new antivirals.

Host-guest interactions between β-cyclodextrin and piroxicam

Abstract In order to obtain experimental evidence for the complexation between β-cyclodextrin and piroxicam (4-hydroxy-2-methyl-N-2-pyridyl-2H-1,2-benzothiazine-3-carboxamide-1,1-dioxide) and to investigate the thermodynamics of this interaction, a flow microcalorimetric study of this system has been undertaken. In fact, the mixing of ,β-cyclodextrin and piroxicam in ratio 1:1 in given experimental conditions gives rise to a system showing better pharmacological properties than piroxicam alone. The results confirm the formation of a complex between β-cyclodextrin and piroxicam and allow the evaluation of the equilibrium constant for the process, assuming the formation of a 1: 1 complex. It results in log K ′ = 3.54, where K ′ is the apparent formation constant at pH 9.00, in good agreement with literature data for similar compounds. The values of ΔH and ΔS are also reported.

Isobaric heat capacity and structure of water and heavy water in the liquid state

Abstract The isobaric heat capacity of liquid H2O, Cp, as a function of temperature, decreases between 0° and about 35°C and then increases up to 100°C. Analogous behaviour is shown by liquid D2O. A statistical thermodynamic model has been applied to the experimental heat capacity data. The behaviour is explained by assuming that an equilibrium A + B = AB is established between clusters A and AB of water of different composition. The total heat capacity is considered as the sum of three terms Cp = (1 − α)Cp,0,AB + αCp,0,A + ΔCp,app. The term ΔCp,app depends explicitly on the reaction enthalpy. In H2O, the enthalpy ΔH = − 18.4 kJ mol−1 for the dissociation reaction and the heat capacity Cp,B = 47.8 J K−1 mol−1 for free water molecules are calculated. Analogous calculations performed for D2O yield the enthalpy, ΔH = − 1.64 kJ mol−1 and the heat capacity, Cp,B = 49.18 J K−1 mol−1.

Solubility of inert gases and liquid hydrocarbons in water

The thermodynamic statistical model based on the distribution of molecular populations among energy levels has been employed for the analysis of the solubility of hydrocarbons and other inert gases or liquids in water at different temperatures. The statistical distribution is described by a convoluted partition function ZG·ζs. The product of a grand canonical partition function ZG represents the distribution of the species in the reaction while the canonical partition function ZG represents the properties of the solvent. The first derivative of the logarithm of the partition function with respect to 1/T is the apparent enthalpy which is the result of the contributions of the separate partition functions, {ΔHaap}T=ΔHo+nwCp,wT, where {ΔHapp}T refers to ZG, nwCp,wT=−ΔHw to ζs, and ΔHo is the change in enthalpy of hydrocarbon-water reaction. The plot {ΔHapp}T vs/ T results in a straight line with slope nw at constant Cp,w. The apparent enthalpy is obtained from the coefficients of the polynomial fitting of the solubility data, as a function of 1/T. Alternatively, the apparent enthalpy can be determined calorimetrically. The enthalpy thus obtained is a linear function of the Kelvin temperature. The values of nw range from 1.6, 1.9, 5.6 to 5.8 for helium, hydorgen, butane and hexane, respectively. For fluorocompounds the range of nw is 10.1 to 11.1 indicating that nw is a function of the number of water molecules expelled from the cage of solvent to form a cavity to host the solute molecule. The analysis of several sets of calorimetric or solubility data with the present molecular thermodynamic model yields values of ΔHo and nw consistent with the size of the dissolved molecules.