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Platinum complexes are used in chemotherapy, primarily as antineoplastic agents. In this study, we assessed the cytotoxic and cytostatic properties of a set of osmium(II), ruthenium(II), iridium(III) and rhodium(III) half-sandwich-type complexes with bidentate monosaccharide ligands. We identified 5 compounds with moderate to negligible acute cytotoxicity but with potent long-term cytostatic activity. These structure-activity relationship studies revealed that: (1) osmium(II) p-cymene complexes were active in all models, while rhodium(III) and iridium(III) Cp* complexes proved largely inactive; (2) the biological effect was influenced by the nature of the central azole ring of the ligands—1,2,3-triazole was the most effective, followed by 1,3,4-oxadiazole, while the isomeric 1,2,4-oxadiazole abolished the cytostatic activity; (3) we found a correlation between the hydrophobic character of the complexes and their cytostatic activity: compounds with O-benzoyl protective groups on the carbohydrate moiety were active, compared to O-deprotected ones. The best compound, an osmium(II) complex, had an IC50 value of 0.70 µM. Furthermore, the steepness of the inhibitory curve of the active complexes suggested cooperative binding; cooperative molecules were better inhibitors than non-cooperative ones. The cytostatic activity of the active complexes was abolished by a lipid-soluble antioxidant, vitamin E, suggesting that oxidative stress plays a major role in the biological activity of the complexes. The complexes were active on ovarian cancer, pancreatic adenocarcinoma, osteosarcoma and Hodgkin’s lymphoma cells, but were inactive on primary, non-transformed human fibroblasts, indicating their applicability as potential anticancer agents.

The interaction with red blood cells (RBC) of vanadium(V) complexes formed by 3,5-difluoropicolinic acid (HpicFF), 3-hydroxypicolinic acid (H2hypic) and pyrazinecarboxylic acid (Hprz) has been examined. Electron Paramagnetic Resonance (EPR) spectroscopy results suggest that V(V) is reduced in all cases to V(IV) inside the erythrocytes. The thermodynamic stability of V(V) and corresponding V(IV) complexes has been related with the species detected in the cellular environment; when the ligands form unstable complexes with both VVO2+ and VIVO2+ ions (picFF and prz), the species formed inside RBC are similar to those observed when starting with vanadate(V) alone, that is VIVO2+ –formed upon V(V) reduction– distributing among the cellular bioligands forming different types of complexes. When in the ligand molecule other groups able to form chelated complexes are present (hypic), more stable species are formed inside the RBC. The amount of complex able to enter the RBC depends on the ligand structure which could influence the metal uptake. The interaction of different VIVO2+ complexes formed by picolinate (picFF, pic = picolinate, picCN = 5-cyanopicolinate) and pyrazinecarboxylate (prz and przNH2 = 3-aminopyrazine-2-carboxylate) derivatives with hemoglobin (Hb), which is the main candidate to bind the VIVO2+ ion in the cytosol, was also investigated to rationalize the results. The ligands which form at physiological pH relatively stable complexes with VIVO2+ (przNH2 and pic) can give inside the erythrocytes mixed species after the replacement of an equatorial water molecule by an imidazole nitrogen of a histidine residue of the protein, while the ligands which form with VIVO2+ unstable complexes (picCN, picFF, prz) yield the same species observed in the binary system VIVO2+–Hb.

Two novel, pyridinone-based chelating ligands containing separated (O,O) and (Namino,Nhet) chelating sets (Namino = secondary amine, Nhet = pyrrole N for H(L3) (1-(3-(((1H-pyrrole-2-yl)methyl)-amino)propyl)-3-hydroxy-2-methylpyridin-4(1H)-one) or pyridine N for H(L5) (3-hydroxy-2-methyl-1-(3-((pyridin-2-ylmethyl)amino)propyl)pyridin-4(1H)-one)) were synthesized via reduction of the appropriate imines. Their proton dissociation processes were explored, and the molecular structures of two synthons were assessed by X-ray crystallography. These ambidentate chelating ligands are intended to develop Co(III)/PGM (PGM = platinum group metal) heterobimetallic multitargeted complexes with anticancer potential. To explore their metal ion binding ability, the interaction with Pd(II), [(η6-p-cym)Ru]2+ and [(η5-Cp*)Rh]2+ (p-cym = 1-methyl-4-isopropylbenzene, Cp* = pentamethyl-cyclopentadienyl anion) cations was studied in aqueous solution with the combined use of pH-potentiometry, NMR and HR ESI-MS. In general, organorhodium was found to form more labile complexes over ruthenium, while complexation of the (N,N) chelating set was slower than the processes of the pyridinone unit with (O,O) coordination. Formation of the organoruthenium complexes starts at lower pH (higher thermodynamic stabilities of the corresponding complexes) than for [(η5-Cp*)Rh]2+ but, due to the higher affinity of [η6-p-cym)Ru]2+ towards hydrolysis, the complexed ligands are capable of competing with hydroxide ion in a lesser extent than for the rhodium systems. As a result, under biologically relevant conditions, the rhodium binding effectivity of the ligands becomes comparable or even slightly higher than their effectivity towards ruthenium. Our results indicate that H(L3) is a less efficient (N,N) chelator for these metal ions than H(L5). Similarly, due to the relative effectivity of the (O,O) and (N,N) chelates at a 1:1 metal-ion-to-ligand ratio, H(L5) coordinates in a (N,N) manner to both cations in the whole pH range studied while, for H(L3), the complexation starts with (O,O) coordination. At a 2:1 metal-ion-to-ligand ratio, H(L3) cannot hinder the intensive hydrolysis of the second metal ion, although a small amount of 2:1 complex with [(η5-Cp*)Rh]2+ can also be detected.

Aims: The aim of the present study was to investigate the biochemical properties of nitrosopersulfide (SSNO -), a key intermediate of the nitric oxide (NO)/sulfide cross talk. Results: We obtained corroborating evidence that SSNO - is indeed a major product of the reaction of S-nitrosothiols with hydrogen sulfide (H 2S). It was found to be relatively stable (t 1/2 *1 h at room temperature) in aqueous solution of physiological pH, stabilized by the presence of excess sulfide and resistant toward reduction by other thiols. Furthermore, we here show that SSNO - escapes the reducing power of the NADPH-driven biological reducing machineries, the thioredoxin and glutathione reductase systems. The slow decomposition of SSNO - produces inorganic polysulfide species, which effectively induce per/polysulfidation on glutathione or protein cysteine (Cys) residues. Our data also demonstrate that, in contrast to the transient activation by inorganic polysulfides, SSNO - induces long-term potentiation of TRPA1 (transient receptor potential ankyrin 1) channels, which may be due to its propensity to generate a slow flux of polysulfide in situ. Innovation: The characterized properties of SSNO - would seem to represent unique features in cell signaling by enabling sulfur and nitrogen trafficking within the reducing environment of the cytosol, with targeted release of both NO and polysulfide equivalents. Conclusion: SSNO - is a surprisingly stable bioactive product of the chemical interaction of S-nitrosothiol species and H 2S that is resistant to reduction by the thioredoxin and glutathione systems. As well as generating NO, it releases inorganic polysulfides, enabling transfer of sulfane sulfur species to peptide/protein Cys residues. The sustained activation of TRPA1 channels by SSNO - is most likely linked to all these properties. Antioxid. Redox Signal. 33, 1277–1294.

By using various techniques (pH-potentiometry, UV-Visible spectrophotometry, 1H and 17O-NMR, EPR, ESI-MS), first time in the literature, solution equilibrium study has been performed on complexes of dipeptide and tripeptide hydroxamic acids&mdash, AlaAlaNHOH, AlaAlaN(Me)OH, AlaGlyGlyNHOH, and AlaGlyGlyN(Me)OH&mdash, with 4d metals: the essential Mo(VI) and two half-sandwich type cations, [(&eta, 6-p-cym)Ru(H2O)3]2+ as well as [(&eta, 5-Cp*)Rh(H2O)3]2+, the latter two having potential importance in cancer therapy. The tripeptide derivatives have also been studied with some biologically important 3d metals, such as Fe(III), Ni(II), Cu(II), and Zn(II), in order to compare these new results with the corresponding previously obtained ones on dipeptide hydroxamic acids. Based on the outcomes, the effects of the type of metal ions, the coordination number, the number and types of donor atoms, and their relative positions to each other on the complexation have been evaluated in the present work. We hope that these collected results might be used when a new peptide-based hydroxamic acid molecule is planned with some purpose, e.g. to develop a potential metalloenzyme inhibitor.

In this work the speciation in aqueous solution of the compound [V IV O(oda)(H 2 O) 2 ], where oda is oxydiacetate dianion (OOCCH 2 ) 2 O, which shows in vitro anticancer activity, was studied. Its interaction with the two serum bioligands with highest affinity for V IV , lactic (Hlact) and citric (H 3 citr) acid, and with the two proteins candidate to participate to the transport of V IV O compounds in the organism, transferrin (hTf) and albumin (HSA), was also examined. The study was carried out with the combined application of spectroscopic (Electron Paramagnetic Resonance, EPR), analytical (pH-potentiometry) and computational (Density Functional Theory, DFT) methods. The results showed that in aqueous solution [VO(oda)(H 2 O) 2 ] undergoes hydrolysis above pH 4–5 with formation of the EPR-active species [(VO) 2 (oda) 2 (OH) 2 ] 2− around pH 6 and of [(VO) 2 (OH) 5 ] − at physiological pH. DFT calculations suggested that the most stable isomers of 1:1 species are the hexa-coordinated OC -6-23 with a mer arrangement of oda – similar to that observed in the solid state – and the penta-coordinated SPY -5-14, whereas for 1:2 species the fac arrangement of oda is favored. Citrate is able to displace completely the oda ligand in [VO(oda)(H 2 O) 2 ] and only the dinuclear species [(VO) 2 (citrH -1 ) 2 ] 4− was detected at pH 7.4, while with lactate the formation of a mixed complex V IV O–oda–lact was observed. [VO(oda)(H 2 O) 2 ] interacts with apo-transferrin forming a mixed complex (VO)(hTf)(oda) where vanadium is bound in the iron sites and oda behaves as a synergistic anion, while with albumin no interaction was revealed. Model calculations suggest that when [VO(oda)(H 2 O) 2 ] is administered orally (concentration ca . 1–10 μM) or by injection (concentration approximately in the range 10–100 μM), (VO)(hTf) and (VO) 2 (hTf) should be formed; these species could reach the target organs and be recognized by the hTf receptors of the cells, favoring the vanadium uptake.

Complex formation equilibria between [(η5-Cp*)RhIII(H2O)3]2+ and aminohydroxamic acids (L-2-amino-N-hydroxyacetamide (α-alahaH), 3-amino-N-hydroxypropanamide (β-alahaH) and 4-amino-N-hydroxybutanamide (GABAha, γ-abhaH)) having the primary amino group in different chelatable positions relative to the hydroxamic function were studied using pH-potentiometric, 1H NMR and ESI-MS methods and the formation constants of the complexes present in aqueous solution are reported. The relative order of the pH-dependent conditional stability of the hydroxamate type (O,O) and (Namino,Nhydroxamato) chelates was found to determine to a great extent the coordination modes both in the mono- and various dinuclear species formed. While with α-alaha−, a 5-membered (N,N) chelated mononuclear complex predominates, with β-alaha− in a wide pH-range, very stable dinuclear cluster ions exist. With γ-abha−, in the most stable complexes, two ligands (in reverse variation) link two half-sandwich cations, coordinating each ligand via the hydroxamate chelate to one metal centre, while via the amino-N to the other one. This arrangement seems to be further stabilized by a hydrogen bond as DFT calculations support the extra stabilization effect of internal H-bonding in [{(η5-Cp*)RhIII}2H−1(γ-abha)2]+. The synthesis, spectral (NMR and IR) and MS characterization of a novel complex with an iridium analogue, [(η5-Cp*)IrIII(α-alaha)Br] (1) is also described. This complex was tested for its in vitro cytotoxicity using human-derived cancer cell lines (A2780, HeLa, DU-145, A549, and MCF-7) and showed insignificant anti-proliferative activity in the micromolar concentration range.

Heterobimetallic complexes with the evolutionary, well-preserved, histidyl-alanyl-valinyl (HAV) sequence for cadherin targeting, an organometallic Ru core with anticancer activity and a radioactive moiety for imaging may hold potential as theranostic agents for cancer. Visible-light irradiation of the HAVAY-NH2 pentapeptide in the presence of [(η(5)-Cp)Ru(η(6)-naphthalene)](+) resulted in the formation of a full sandwich type complex, (η(6)-Tyr-RuCp)-HAVAY-NH2 in aqueous solution, where the metal ion is connected to the Tyr (Y) unit of the peptide. Conjugation of this complex to 2,2'-(7-(1-carboxy-4-((4-isothiocyanatobenzyl)amino)-4-oxobutyl)-1,4,7-triazonane-1,4-diyl)diacetic acid (NODA-GA) and subsequent metalation of the resulting product with stable ((nat)Ga) and radioactive ((67)Ga) isotope yielded (nat)Ga/(67)Ga-NODA-GA-[(η(6)-Tyr-RuCp)-HAVAY-NH2]. The non-radioactive compounds were characterized by NMR spectroscopy and Mass Spectrometry. The cellular uptake and cytotoxicity of the radioactive and non-radioactive complexes, respectively, were evaluated in various human cancer cell lines characterized by different levels of N- or E-cadherins expression. Results from these studies indicate moderate cellular uptake of the radioactive complexes. However, the inhibition of the cell proliferation was not relevant.

In order to model the metal ion binding capabilities of high molecular mass components of blood the interaction between [(η6-p-cym)Ru(H2O)3]2+ and terminally protected oligopeptides containing three histidyl moieties (Ac-HHH-NH2, Ac-HAHH-NH2, Ac-HAHAH-NH2 and Ac-H*AH*AH*-NH2, where A = L-alanyl, H = L-histidyl, H* = N3-methyl-L-histidyl) were studied by pH-potentiometric, ESI-TOF-MS, circular dichroism and NMR methods at an ionic strength of 0.20 M KCl or KNO3 as well as using density functional theory (DFT) calculations. Protonation constants of the novel peptides are reported. Although for Ac-HHH-NH2 the immediate formation of precipitation with [(η6-p-cym)Ru(H2O)3]2+ hindered any further solution investigations results of the detailed NMR and MS studies revealed that the other three ligands coordinate to the metal ion in rather slow processes via the imidazole moieties forming [(η6-p-cym)RuL]2+ (L = oligopeptide) type species in the slightly acidic, neutral pH-range. At pH ∼7.5 identical binding mode of Ac-HAHH-NH2 and Ac-HAHAH-NH2 in the [(η6-p-cym)RuL]2+ via three imidazole nitrogens was found hindering completely the hydrolysis of the metal ion even at 1:1 metal ion to ligand ratio. At elevated pH MS evidences support the involvement of amide-N donor(s) in metal ion binding too beside partial hydrolysis. 0.20 M KCl medium was found to hinder effectively the hydrolytic processes of the metal ion in the basic pH-range without altering the coordination of the imidazole side chains. Both NMR and DFT results support the imidazole-N1 (“far” or “τ”) over the N3 (“near” or “π”) coordination of the histidyl side chains of all these oligopeptides to the organometallic ruthenium(II) cation.

In order to model the biotransformation reactions of half-sandwich type platinum metal complexes with potential antiproliferative activity in the serum the interaction between [(η 6 - p -cym)Ru(H 2 O) 3 ] 2+ and N-methylimidazole was studied in aqueous 0.20 M KNO 3 or KCl medium with the combined use of pH-potentiometric, NMR, UV–Vis and ESI-TOF-MS techniques. The results indicate that up to three ligands can coordinate to the metal ion in rather slow processes resulting in mononuclear species. In the absence of chloride ions various mixed hydroxido complexes are also identified under basic conditions. Chloride was found to take the free coordination site(s) of the metal ion in the 1:1 and 1:2 species and to hinder effectively the hydrolytic processes. Stability constants of the species are reported. The results indicate that even this monodentate model ligand, N-methylimidazole, forms stable complexes with the metal ion under physiologically relevant conditions at 1:3 metal to ligand ratio and prevents it from complete hydrolysis at pH = 7.4.

To explore the role of the side chain alcoholic OH group in metal ion binding the interaction between [Ru(η6-p-cym)(H2O)3]2+ (p-cym = 1-isopropyl-4-methylbenzene) and dl -serine (serH) or dl -isoserine (iseH) was studied by means of pH-potentiometry, 1H NMR and ESI-MS techniques in aqueous solution. The results indicate that, unlike simple amino acids with no donor atom in the side chain, ser and especially ise are potent ruthenium ion binders and are capable of preventing the metal ion from significant hydrolysis at pH = 7.4 due to the metal ion assisted deprotonation and coordination of the alcoholic OH function and forming stable joined chelates with [NH2, COO−, O−] donor set. Comparison of the speciations obtained for the two systems indicates that ise with (6 + 5) membered joined chelates of the [NH2, COO−, O−] donor set forms more stable complexes than ser which is suitable for (5 + 6) [NH2, COO−, O−] coordination. Analysis of the effect of the replacement of the amino group in the [NH2, COO−, O−] donor set of ise by a COO− (citrate) reveals that the latter is more effective in the acidic pH range; the two donor sets show comparable metal ion strengths under physiologically relevant conditions and ise is found to be a better metal ion binder in the basic pH range than citrate due to the basicity difference of the –NH2 and –COO− groups in ise and citrate ligands, respectively. [Ru(η6-p-cym)ise](CF3SO3)·0.5H2O (1) with [NH2, COO−, OH] coordination of the ligand was also synthesized and characterized in the solid state by various analytical techniques.

The biotransformation in the blood serum of the two anti-diabetic agents [VO(ema) 2 ] – or BEOV – and [VO(koj) 2 ] formed by ethylmaltol (Hema) and kojic acid (Hkoj) was studied with EPR spectroscopy, pH-potentiometry and DFT calculations. For comparison, the behavior of the systems with tropolone (Htrop) was also analyzed. The interaction of [VO(ema) 2 ] and [VO(koj) 2 ] with the most important bioligands of the serum, lactic (Hlact) and citric acid (H 3 citr), human serum transferrin (hTf), human serum albumin (HSA) and immunoglobulin G (IgG) was examined and discussed. Among the several mixed species observed, cis -VO( carrier ) 2 (hTf), cis -VO( carrier ) 2 (HSA) and cis -VO( carrier ) 2 (IgG), where carrier is ethylmaltolate or kojate, with a His-N of the protein coordinated in the equatorial position, are plausible candidates for the transport processes of the drug toward the target organs. The values of the logβ are in the range 19.6–19.8 for the species formed by ethylmaltol and 17.4–17.6 for those formed by kojic acid. The formation of such species was confirmed through pH-titrations of the model systems VO 2 + / carrier /1-MeIm and VO 2 + / carrier /Ac-his, where 1-MeIm and Ac-his are 1-methylimidazole and N -acetylhistamine, and DFT calculations of 51 V A z of the model species cis -[VO( carrier ) 2 (1-MeIm)] and cis -[VO( carrier ) 2 (Ac-his)]. The values of the stability constants for the mixed species observed were used to predict the biodistribution of VO 2 + ion between the blood serum components for concentrations of 1, 10 and 50 μM.

The interaction of the potent anti-diabetic agent bis(maltolato)oxidovanadium(IV) (BMOV) with some proteins of blood serum was studied by EPR spectroscopy, pH-potentiometry and DFT calculations. The formation of cis-VO(ma)(2)(hTf), cis-VO(ma)(2)(HSA) and cis-VO(ma)(2)(IgG), their role in the biotransformation in vivo and the mechanism of transport of BMOV in blood are discussed.

Three new peptidehydroxamic acids ( l -alanyl- l -histidinehydroxamic acid, l -Ala- l -HisNHOH, l -alanyl- l -alanyl- l -histidinehydroxamic acid, l -Ala- l -Ala- l -HisNHOH and l -histidyl- l -alaninehydroxamic acid, l -His- l -AlaNHOH) were synthesized and their complexation with Cu(II), Ni(II) and Zn(II) were studied by pH-potentiometric, UV–Vis, CD, 1 H NMR, EPR and ESI-MS methods. Each of the studied peptide derivatives involves one side-chain imidazole unit and the effect of this group on the metal binding of the hydroxamic moiety is evaluated in the paper. The obtained results are compared to those of the complexes of some histidine-containing di- or tripeptides and also to those of hydroxamic derivatives of aliphatic peptides. A competition between the hydroxamate and imidazole functions occurs in all systems, but the extent differs from metal to metal, from ligand to ligand and depends very much on the pH. The imidazole was found to play the most determinant role in the Cu(II) complexes, somewhat less in the Ni(II)-containing ones, while (except the case of l -Ala- l -HisNHOH) negligible role was found in the Zn(II)-complexes. Common feature of the Ni(II)- and especially Cu(II)-containing systems is that if an imidazole-N is displaced by a hydroxamate, imidazole-bridged di- and polynuclear complexes are formed.

Solution equilibrium studies on Cu(II)-, Ni(II)- and Zn(II)-N-Me-β-Alaninehydroxamic acid (N-Me-β-Alaha), -N-Me-α-alaninehydroxamic acid (N-Me-α-Alaha), -Imidazole-4-carbohydroxamic acid (Im-4-Cha), -N-Me-imidazole-4-carbohydroxamic acid (N-Me-Im-4-Cha) and -Imidazole-4-acetohydroxamic acid (Im-4-Aha) systems have been performed by pH-potentiometry, UV–Vis spectrophotometry, EPR, CD, ESI-MS and 1H NMR methods. According to the results: (i) the amino-N atoms are more basic in N-Me-α-Alaha and N-Me-β-Alaha than the hydroxamate function, but the trend is just the opposite between the imidazole-N(3) and hydroxamate. (ii) The metal ion anchor is always the hydroxamate part in the amino acid derivatives, while it is always the imidazole-N(3) in the studied imidazolehydroxamic acids. (iii) The three studied N-Me derivatives do not form metallacrowns. Only hydroxamate type chelate is formed with N-Me-β-Alaha, but with N-Me-α-Alaha a new type of coordination mode (via amino-N and hydroxamate-O) also exists. N-Me-Im-4-Cha also forms a dinuclear complex, [M2L3], with Cu(II) and Ni(II) (but not with Zn(II)). In this complex, one of the three ligands might bridge the two metal ions (five-membered hydroxamate-(O,O) plus five-membered (Nim, Ocarb) bridging bis-chelating mode), while each of the additional two ligands binds to one metal. (iv) The two studied N–H derivatives, having dissociable proton on the hydroxamic-N, are able to form metallacrown species. A pentanuclear complex, [M5L4H−4], is exclusively formed above pH 4 between Cu(II) and Im-4-Aha. Interestingly, this 12-metallacrown-4 type complex, although together with various mononuclear binding isomers, appears also with Ni(II) and Zn(II). Unfortunately, the complexes of Im-4-Cha are not soluble in water at physiological pH at all.

The interaction between oxovanadium(IV) and primary hydroxamic acids (aceto- (HAha), benzohydroxamic acid (HBha) or N -hydroxy- N ′-[3-( N -hydroxy-carbamoyl)-propyl]-heptane-1,7-dicarboxamide (H 2 Diha)) as well as the corresponding N -methyl substituted hydroxamic acids (HMeAha, HMeBha, H 2 MeDiha) was studied by pH-potentiometric and pH-stat methods in aqueous solution. Stoichiometry and stability constants of the VO(IV) complexes formed in the pH range 2.0–5.0 were determined and the effect of the substituents on the C or N atoms of the hydroxamic group is discussed. The dihydroxamic acids are more effective oxovanadium(IV) binders than the monohydroxamic ones. Above pH = 4.5–5.0 (depending upon ligand excess), slow redox reaction was found in all the systems studied. During this process, beside the oxidation of VO(IV) to the appropriate V(V)–hydroxamate complex, formation of amides as reduction products was detected under anaerobic conditions. The same reaction can also take place when stirring sparingly soluble VOA 2 type complexes (A: Bha or MeBha) in water in the absence of oxygen. The exact stoichiometry of the redox reaction was determined by pH-stat measurements and found to be 2VO(R 1 CON(R 2 )O) 2 + R 1 CON(R 2 )OH + H 2 O = 2[VO 2 (R 1 CON(R 2 )O) 2 ] − + 2H + + R 1 CON(R 2 )H (R 1 : CH 3 , C 6 H 5 ; R 2 : H, CH 3 ) for the monohydroxamates. The N -methyl hydroxamic acids tend to oxidise VO 2+ significantly slower than the corresponding primary ones.