Structure, properties, and decomposition in biological systems of a new nitrosyl iron complex with 2-methoxythiophenolyls, promising for the treatment of cardiovascular diseases.

A new promising binuclear tetranitrosyl iron complex with 2-methoxythiophenolyl of the composition [Fe2(C7H7OS)2(NO)4] (complex 1), which acts on the therapeutic targets of cardiovascular diseases, guanylate and adenylate cyclase, has been synthesized. X-ray diffraction data show the presence of two isoforms of complex 1; according to quantum chemical calculations, the structure of only the trans isomer is stable in solutions. The processes of transformation of complex 1 in DMSO, in aqueous solutions, as well as in the presence of bovine serum albumin, reduced glutathione, and mucin were studied. DMSO promotes the decomposition of the original complex 1 into mononuclear products. In biological systems, the mechanisms of decomposition of the complex 1 differ from aqueous solutions. In albumin solution, a gradual formation of a high-molecular-weight dinitrosyl complex is observed, obtained by coordinating the [Fe(NO)2]+ fragment with the amino acid groups of the protein. In the presence of mucin, an EPR signal from stable mononitrosyl products is observed. The introduction of glutathione into the system of the complex 1 leads to the replacement of one initial thioligand with glutathione. In the model systems under study, a more efficient and prolonged generation of NO groups is observed compared to a buffer solution. The obtained data on the influence of the environment on the properties of the complex 1 in combination with a study of their effect on enzymes allow us to recommend it for further study as a potential drug with vasodilator, antianginal, and hypotensive properties.

Effects of Nitrosyl Iron Complexes with Thiol, Phosphate, and Thiosulfate Ligands on Hemoglobin.

Nitrosyl iron complexes are remarkably multifactorial pharmacological agents. These compounds have been proven to be particularly effective in treating cardiovascular and oncological diseases. We evaluated and compared the antioxidant activity of tetranitrosyl iron complexes (TNICs) with thiosulfate ligands and dinitrosyl iron complexes (DNICs) with glutathione (DNIC-GS) or phosphate (DNIC-PO4-) ligands in hemoglobin-containing systems. The studied effects included the production of free radical intermediates during hemoglobin (Hb) oxidation by tert-butyl hydroperoxide, oxidative modification of Hb, and antioxidant properties of nitrosyl iron complexes. Measuring luminol chemiluminescence revealed that the antioxidant effect of TNICs was higher compared to DNIC-PO4-. DNIC-GS either did not exhibit antioxidant activity or exerted prooxidant effects at certain concentrations, which might have resulted from thiyl radical formation. TNICs and DNIC-PO4- efficiently protected the Hb heme group from decomposition by organic hydroperoxides. DNIC-GS did not exert any protective effects on the heme group; however, it abolished oxoferrylHb generation. TNICs inhibited the formation of Hb multimeric forms more efficiently than DNICs. Thus, TNICs had more pronounced antioxidant activity than DNICs in Hb-containing systems.

Effect of solvents and glutathione on the decomposition of the nitrosyl iron complex with N-ethylthiourea ligands: An experimental and theoretical study.

Dinitrosyl iron complexes (DNICs) are a depot and potential source of free NO in organisms. Their synthetic analog, N-ethylthiourea DNIC [Fe(SC(NH2)(NHC2H5))2(NO)2]+Cl-∙[Fe(SC(NH2)(NHC2H5))Cl(NO)2]0 (complex 1), as cardioprotective and cytostatic agent is a promising prodrug for the treatment of socially relevant diseases. In this work, transformation mechanism of complex 1 has been studied in anaerobic aqueous solution (pH = 7.0), DMSO, and ethanol. It was shown that the solvent has a significant effect on the decomposition of complex. According to EPR-spectroscopy, only cationic part of complex is found upon its dissolution in water; only neutral part is retained in DMSO, and both fragments are present in ethanol. Effective generation of NO occurs in an aqueous solution. The structures of the decomposition products were proposed for all solvents, their UV-spectra and rate constants were calculated. From the experimental and theoretical data obtained, it follows that complex 1 is most stable in DMSO. Solutions of complex in a DMSO-water mixture can be used to improve its bioavailability in further in vitro and in vivo studies. Also, we have analyzed its interaction with glutathione (GSH), which can participate in the metabolism of this compound. This study shows that complex 1 reacts with GSH to form a new binuclear DNIC with two GS--ligands. It was found that the resulting complex is a more prolonged NO-donor than the initial one: k = 6.1∙10-3·s-1 in buffer, k = 6.4∙10-5 s-1 with GSH. This reaction may prevent S-glutathionylation of the essential enzyme systems and is important for metabolism of complex, associated with its antitumor activity.

Albumin as a prospective carrier of the nitrosyl iron complex with thiourea and thiosulfate ligands under aerobic conditions.

High-molecular-weight dinitrosyl iron complexes (DNICs) are formed in living systems and are a stable depot of nitrogen monoxide (NO). In this work, using experimental and theoretical methods, we investigated the interaction of their synthetic analog, a promising cardiotropic complex of the composition [Fe(SC(NH2)2)2(NO)2]2[Fe2(S2O3)2(NO)4], with bovine serum albumin (BSA) in aqueous aerobic solutions. We suggested that, under these conditions, the decomposition product of the initial complex with oxygen, the [Fe(NO)(NO2)]+ fragment, can bind in the hydrophobic pocket of the protein. As a result of this interaction, high-molecular-weight Fe(Cys34)(His39)(NO)(NO2) is formed. The binding constant of the complex with protein measured by the quenching of intrinsic fluorescence of BSA is 7.2 × 105 M-1. According to EPR and UV-spectroscopy data, the interaction of the complex with the protein leads to its significant stabilization. In addition to coordination binding, the studied complex can be adsorbed onto the protein surface due to weak intermolecular interactions, resulting in the prolonged generation of NO.

Effect of albumin on the transformation of dinitrosyl iron complexes with thiourea ligands.

Interaction and transformation of the mononuclear cationic dinitrosyl iron complex [Fe(SC(NH2)2)2(NO)2]+ (complex 1) upon binding with bovine serum albumin (BSA) have been explored using kinetic measurements, UV-Vis and fluorescence spectroscopy, and computational molecular modeling. BSA was found to bind up to five molecules of complex 1 per one protein molecule; as a result, the rate of NO release by complex 1 into solution decreases by a factor of 10. The binding constant of complex 1 with BSA measured by the quenching of intrinsic fluorescence of BSA is 5 × 105 М-1. Molecular docking calculations at pH = 7 have determined five-six low-energy binding sites for complex 1 at subunits I and II of BSA. The most stable protein-ligand complexes are located at the protein pockets near Cys34. The spectroscopic measurements and docking calculations have shown that the decomposition product of complex 1, the Fe(NO)2+ fragment, can form an adduct Fe(Cys34)(His39)(NO)2 (complex 2) with the coordination bonds of Fe with atoms S of Cys34 and ND of His39. The structure of complex 2 was supported by the density functional calculations of the absorption spectrum. Decomposition of complex 2 leads to nitrosylation of BSA at atom S of Cys34. Complexes 1 (bound with BSA), 2 and the nitrosylated BSA can serve as NO depot in plasma.

Decomposition of water-soluble mononitrosyl iron complexes with dithiocarbamates and of dinitrosyl iron complexes with thiol ligands in animal organisms.

EPR studies have shown that water-soluble mononitrosyl iron complexes with N-methyl-d-glucamine dithiocarbamate (MNIC-MGD) (3 micromol) injected to intact mice were decomposed virtually completely within 1h. The total content of MNIC-MGD in animal urine did not exceed 30 nmol/ml. In the liver, a small amount of MNIC-MGD were converted into dinitrosyl iron complexes (30 nmol/g of liver tissue). The same was observed in intact rabbits in which MNIC-MGD formation was induced by endogenous or exogenous NO binding to NO traps, viz., iron complexes with MGD. In mice, the content of MNIC-MGD in urine samples did not change after bacterial lipopolysaccharide-induced expression of iNOS. It was supposed that MNIC-MGD decomposition in intact animals was largely due to the release of NO from the complexes and its further transfer to other specific acceptors. In mice with iNOS expression, the main contribution to MNIC-MGD decomposition was made by superoxide ions whose destructive effect is mediated by an oxidative mechanism. This effect could fully compensate the augmented synthesis of MNIC-MGD involving endogenous NO whose production was supported by iNOS. Water-soluble dinitrosyl iron complexes (DNIC) with various thiol-containing ligands and thiosulfate injected to intact mice were also decomposed; however, in this case the effect was less pronounced than in the case of MNIC-MGD. It was concluded that DNIC decomposition was largely due to the oxidative effect of superoxide ions on these complexes.

Dinitrosyl-iron complexes with thiol-containing ligands: spatial and electronic structures.

Parameters of the EPR signals of monomeric dinitrosyl-iron complexes with 1H-1,2,4-triazole-3-thiol (DNIC-MT), obtained by treating MT+ferrous iron in DMSO solution with gaseous NO, have been compared with those of the crystalline monomeric DNIC-MT with tetrahedral structure. Dissolved DNIC-MT were characterized by the isotropic EPR signal centered at g=2.03 with half-width of 0.7 mT and quintet hyperfine structure when recorded at ambient temperature or the anisotropic EPR signal with g( perpendicular)=2.045, g( parallel)=2.014 from frozen solution at 77 kappa, Cyrillic. DNIC-MT in crystalline state showed the structure-less symmetrical singlet EPR signal centered at g=2.03 and half-width of 1.7 mT at both room and liquid nitrogen temperature. The Lorentz shape of this signal indicates the strong exchange interaction between these complexes in the DNIC-MT crystal. Being dissolved in DMSO the crystalline sample of DNIC-MT demonstrated the EPR signal typical for DNIC-MT, obtained by treating MT+ferrous iron in DMSO solution with gaseous NO. Low spin (S=1/2) d(9) electron configuration of DNIC-MT with tetrahedral structure (formula [(MT-S(.))(2)Fe(-1)(NO(+))(2)](+)) was suggested to be responsible for the signal of DNIC-MT in crystalline state. Dissolving of the crystals of DNIC-MT may result in the change of their spatial and electronic structure, namely, tetrahedral structure of the complexes characterized by low spin d(9) electronic configuration transforms into a plane-square structure with d(7) electronic configuration and low spin S=1/2 state (formula [(MT- S(-))(2)Fe(+)(NO(+))(2)](+)). The latter was suggested to be characteristic of other DNICs with various thiol-containing ligands in the solutions. The proposed mechanism of these DNICs formation from ferrous iron, thiol and NO shows that the process could be accompanied by the ionization of NO molecules to NO(+) and NO(-) ions in the complexes. Detailed analysis of the shape of the EPR signals of these complexes provided additional information about the exchange interaction typical for DNIC-MT in crystals.

Protein-bound dinitrosyl-iron complexes appearing in blood of rabbit added with a low-molecular dinitrosyl-iron complex: EPR studies.

The formation of protein-bound dinitrosyl-iron complexes (DNIC) in blood plasma and packed red cell fraction has been demonstrated by the EPR method in the experiments on rabbits which were i/v injected with the low-molecular DNIC with thiosulphate. This formation was ensured by transfer of Fe(+)(NO(+))(2) moieties from low-molecular DNIC onto serum albumin or hemoglobin molecules. Protein-bound DNICs appeared immediately after low-molecular DNIC injection followed with gradually decreasing their amounts. The complexes could be detected by EPR technique during more than two days. The addition of water-soluble NO scavenger, the iron complex with N-methyl-d-glucamine dithiocarbamate (MGD) resulted in decomposition of a part of protein-bound DNICs and in effective excretion of secondary products (mainly mononitrosyl-iron complexes with MGD) from the blood flow.