RESUMO
Metallocompounds have emerged as promising new anticancer agents, which can also exhibit properties to be used in photodynamic therapy. Here, we prepared two ruthenium-based compounds with a 2,2'-bipyridine ligand conjugated to an anthracenyl moiety. These compounds coded GRBA and GRPA contain 2,2'-bipyridine or 1,10-phenathroline as auxiliary ligands, respectively, which provide quite a distinct behavior. Notably, compound GRPA exhibited remarkably high photoproduction of singlet oxygen even in water (ÏΔ = 0.96), almost twice that of GRBA (ÏΔ = 0.52). On the other hand, this latter produced twice more superoxide and hydroxyl radical species than GRPA, which may be due to the modulation of their excited state. Interestingly, GRPA exhibited a modest binding to DNA (Kb = 4.51 × 104), while GRBA did not show a measurable interaction only noticed by circular dichroism measurements. Studies with bacteria showed a great antimicrobial effect, including a synergistic effect in combination with commercial antibiotics. Besides that, GRBA showed very low or no cytotoxicity against four mammalian cells, including a hard-to-treat MDA-MB-231, triple-negative human breast cancer. Potent activities were measured for GRBA upon blue light irradiation, where IC50 of 43 and 13 nmol L-1 were seen against hard-to-treat triple-negative human breast cancer (MDA-MB-231) and ovarian cancer cells (A2780), respectively. These promising results are an interesting case of a simple modification with expressive enhancement of biological activity that deserves further biological studies.
Assuntos
Antibacterianos , Antineoplásicos , Humanos , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/síntese química , Estrutura Molecular , Testes de Sensibilidade Microbiana , Ensaios de Seleção de Medicamentos Antitumorais , Compostos de Rutênio/farmacologia , Compostos de Rutênio/química , Complexos de Coordenação/farmacologia , Complexos de Coordenação/química , Complexos de Coordenação/síntese química , Linhagem Celular Tumoral , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/síntese química , Rutênio/química , Rutênio/farmacologia , Proliferação de Células/efeitos dos fármacos , Farmacorresistência Bacteriana/efeitos dos fármacos , Processos Fotoquímicos , Sobrevivência Celular/efeitos dos fármacos , FotoquimioterapiaRESUMO
For more than 70 years, sodium nitroprusside (SNP) has been used to treat severe hypertension in hospital emergency settings. During this time, a few other clinical uses have also emerged such as in the treatment of acute heart failure as well as improving mitral incompetence and in the intra- and perioperative management during heart surgery. This drug functions by releasing nitric oxide (NO), which modulates several biological processes with many potential therapeutic applications. However, this small molecule has a short lifetime, and it has been administered through the use of NO donor molecules such as SNP. On the other hand, SNP also has some setbacks such as the release of cyanide ions, high water solubility, and very fast NO release kinetics. Currently, there are many drug delivery strategies that can be applied to overcome many of these limitations, providing novel opportunities for the use of old drugs, including SNP. This Perspective describes some nitroprusside properties and highlights new potential therapeutic uses arising from the use of drug delivery systems, mainly silica-based nanoparticles. There is a series of great opportunities to further explore SNP in many medical issues as reviewed, which deserves a closer look by the scientific community.
Assuntos
Nanopartículas , Doadores de Óxido Nítrico , Nitroprussiato , Doadores de Óxido Nítrico/uso terapêutico , Óxido Nítrico , Sistemas de Liberação de MedicamentosRESUMO
A pharmacophore design approach, based on the coordination chemistry of an intimate molecular hybrid of active metabolites of pro-drugs, known to release active species upon enzymatic oxidative activation, is devised. This is exemplified by combining two anti-mycobacterial drugs: pyrazinamide (first line) and delamanid (third line) whose active metabolites are pyrazinoic acid (PyzCOOH) and likely nitroxyl (HNO (or NO.)), respectively. Aiming to generate those active species, a hybrid compound was envisaged by coordination of pyrazine-2-hydroxamic acid (PyzCONHOH) with a Na3[FeII(CN)5] moiety. The corresponding pentacyanoferrate(II) complex Na4[FeII(CN)5(PyzCONHO-)] was synthesized and characterized by several spectroscopic techniques, cyclic voltammetry, and DFT calculations. Chemical oxidation of this complex with H2O2 was shown to induce the release of the metabolite PyzCOOH, without the need of the Mycobacterium tuberculosis (Mtb) pyrazinamidase enzyme (PncA). Control experiments show that both H2O2- and N-coordinated pyrazine FeII species are required, ruling out a direct hydrolysis of the hydroxamic acid or an alternative oxidative route through chelation of a metal center by a hydroxamic group. The release of HNO was observed using EPR spectroscopy in the presence of a spin trapping agent. The devised iron metal complex of pyrazine-2-hydroxamic acid was found inactive against an actively growing/non-resistant Mtb strain; however, it showed a strong dose-dependent and reversible vasodilatory activity with mostly lesser toxic effects than the reference drug sodium nitroprussiate, unveiling thus a potential indication for acute or chronic cardiovascular pathology. This is a priori a further indirect evidence of HNO release from this metal complex, standing as a possible pharmacophore model for an alternative vasodilator drug.
Assuntos
Antituberculosos/síntese química , Complexos de Coordenação/síntese química , Compostos Ferrosos/síntese química , Ácidos Hidroxâmicos/química , Ferro/química , Mycobacterium tuberculosis/efeitos dos fármacos , Óxidos de Nitrogênio/química , Amidoidrolases/metabolismo , Antituberculosos/farmacologia , Complexos de Coordenação/farmacologia , Descoberta de Drogas , Espectroscopia de Ressonância de Spin Eletrônica , Peróxido de Hidrogênio/química , Ligantes , Óxidos de Nitrogênio/metabolismo , Oxirredução , Pirazinamida/análogos & derivados , Pirazinamida/química , VasodilataçãoRESUMO
The aim of this study was to investigate the antibacterial activity, antibiotic-associated synergy, and anti-biofilm activity of the ruthenium complex, cis-[RuCl2 (dppb) (bqdi)]2+ (RuNN). RuNN exhibited antimicrobial activity against Gram-positive bacteria with minimum inhibitory concentration (MIC) values ranging from 15.6 to 62.5 µg ml-1 and minimum bactericidal concentration (MBC) values ranging from 62.5 to 125 µg ml-1. A synergistic effect against Staphylococcus spp. was observed when RuNN was combined with ampicillin, and the range of associated fractional inhibitory concentration index (FICI) values was 0.187 to 0.312. A time-kill curve indicated the bactericidal activity of RuNN in the first 1-5 h. In general, RuNN inhibited biofilm formation and disrupted mature biofilms. Furthermore, RuNN altered the cellular morphology of S. aureus biofilms. Further, RuNN did not cause hemolysis of erythrocytes. The results of this study provide evidence that RuNN is a novel therapeutic candidate to treat bacterial infections caused by Staphylococcus biofilms.
Assuntos
Antibacterianos , Biofilmes , Rutênio , Staphylococcus aureus , Testes de Sensibilidade MicrobianaRESUMO
Ruthenium polypyridine complexes have shown promise as agents for photodynamic therapy (PDT) and tools for molecular biology (chromophore-assisted light inactivation). To accomplish these tasks, it is important to have at least target selectivity and great reactive oxygen species (ROS) photogeneration: two properties that are not easily found in the same molecule. To prepare such new agents, we synthesized two new ruthenium complexes that combine an efficient DNA binding moiety (dppz ligand) together with naphthyl-modified (1) and anthracenyl-modified (2) bipyridine as a strong ROS generator bound to a ruthenium complex. The compounds were fully characterized and their photophysical and photochemical properties investigated. Compound 2 showed one of the highest quantum yields for singlet oxygen production ever reported (ΦΔ= 0.96), along with very high DNA binding (log Kb = 6.78). Such photochemical behavior could be ascribed to the lower triplet state involving the anthracenyl-modified bipyridine, which is associated with easier oxygen quenching. In addition, the compounds exhibited moderate selectivity toward G-quadruplex DNA and binding to the minor groove of DNA, most likely driven by the pendant ligands. Interestingly, they also showed DNA photocleavage activity even upon exposure to a yellow light-emitting diode (LED). Regarding their biological activity, the compounds exhibited an exciting antibacterial action, particularly against Gram-positive bacteria, which was enhanced upon blue LED irradiation. Altogether, these results showed that our strategy succeeded in producing light-triggered DNA binding agents with pharmacological and biotechnological potential.
Assuntos
Complexos de Coordenação/farmacologia , DNA/química , Substâncias Intercalantes/farmacologia , Rutênio/química , 2,2'-Dipiridil/síntese química , 2,2'-Dipiridil/química , 2,2'-Dipiridil/farmacologia , 2,2'-Dipiridil/efeitos da radiação , Antracenos/síntese química , Antracenos/química , Antracenos/farmacologia , Antracenos/efeitos da radiação , Antibacterianos/síntese química , Antibacterianos/química , Antibacterianos/farmacologia , Antibacterianos/efeitos da radiação , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Complexos de Coordenação/efeitos da radiação , Dano ao DNA , Etídio/farmacologia , Bactérias Gram-Positivas/efeitos dos fármacos , Substâncias Intercalantes/síntese química , Substâncias Intercalantes/química , Substâncias Intercalantes/efeitos da radiação , Ligantes , Luz , Oxigênio/química , Fármacos Fotossensibilizantes/síntese química , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/efeitos da radiação , Espécies Reativas de Oxigênio/síntese químicaRESUMO
Antimicrobial resistance has become a global threat to human health, which is coupled with the lack of novel drugs. Metallocompounds have emerged as promising diverse scaffolds for the development of new antibiotics. Herein, we prepared some metal compounds mainly focusing on cis-[Ru(bpy)(dppz)(SO3)(NO)](PF6) (PR02, bpy = 2,2'-bipyridine, dppz = dipyrido[3,2-a:2',3'-c]phenazine), in which phenazinic and nitric oxide ligands along with sulfite conferred some key properties. This compound exhibited a redox potential for bound NO+/0 of -0.252 V (vs. Ag|AgCl) and a high pH for nitrosyl-nitro conversion of 9.16, making the nitrosyl ligand the major species. These compounds were still able to bind to DNA structures. Interestingly, reduced glutathione (GSH) was unable to promote significant NO/HNO release, an uncommon feature of many similar systems. However, this reducing agent was essential to generate superoxide radicals. Antimicrobial studies were carried out using six bacterial strains, where none or very low activity was observed for Gram-negative bacteria. However, PR02 and PR (cis-[Ru(bpy)(dppz)Cl2]) showed high antibacterial activity in some Gram-positive strains (MBC for S. aureus up to 4.9 µmol L-1), where the activity of PR02 was similar to or at least 4-fold better than that of PR. Besides, PR02 showed capacity to inhibit bacterial biofilm formation, a major health issue leading to bacterial tolerance to antibiotics. Interestingly, we also showed that PR02 can function in synergism with the known antibiotic ampicillin, improving their action up to 4-fold even against resistant strains. Altogether, these results showed that PR02 is a promising antimicrobial nitrosyl ruthenium compound combining features beyond its killing action, which deserves further biological studies.
Assuntos
Antibacterianos , Biofilmes , Complexos de Coordenação , Testes de Sensibilidade Microbiana , Fenazinas , Rutênio , Fenazinas/química , Fenazinas/farmacologia , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/síntese química , Rutênio/química , Rutênio/farmacologia , Biofilmes/efeitos dos fármacos , Complexos de Coordenação/química , Complexos de Coordenação/farmacologia , Complexos de Coordenação/síntese química , Sinergismo Farmacológico , Staphylococcus aureus/efeitos dos fármacosRESUMO
Nitrosyl ruthenium complexes are promising platforms for nitric oxide (NO) and nitroxyl (HNO) release, which exert their therapeutic application. In this context, we developed two polypyridinic compounds with the general formula cis-[Ru(NO)(bpy)2(L)]n+, where L is an imidazole derivative. These species were characterized by spectroscopic and electrochemical techniques, including XANES/EXAFS experiments, and further supported by DFT calculations. Interestingly, assays using selective probes evidenced that both complexes can release HNO on reaction with thiols. This finding was biologically validated by HIF-1α detection. The latter protein is related to angiogenesis and inflammation processes under hypoxic conditions, which is selectively destabilized by nitroxyl. These metal complexes also presented vasodilating properties using isolated rat aorta rings and demonstrated antioxidant properties in free radical scavenging experiments. Based on these results, the new nitrosyl ruthenium compounds showed promising characteristics as potential therapeutic agents for the treatment of cardiovascular conditions such as atherosclerosis, deserving further investigation.
Assuntos
Complexos de Coordenação , Rutênio , Animais , Ratos , Óxido Nítrico/química , Óxidos de Nitrogênio/química , Rutênio/química , Compostos de Sulfidrila/química , Doenças CardiovascularesRESUMO
For over a decade, tuberculosis (TB) has been the leading cause of death among infectious diseases. Since the 1950s, isoniazid has been used as a front-line drug in the treatment of TB; however, resistant TB strains have limited its use. The major route of isoniazid resistance relies on KatG enzyme disruption, which does not promote an electron transfer reaction. Here, we investigated the reactivity of isoniazid metal complexes as prototypes for novel self-activating metallodrugs against TB with the aim to overcome resistance. Reactivity studies were conducted with hydrogen peroxide, hexacyanoferrate(III), and aquopentacyanoferrate(III). The latter species showed a preference for the inner-sphere electron transfer reaction pathway. Additionally, electron transfer reaction performed with either free isoniazid or (isoniazid)pentacyanoferrate(II) complex resulted in similar oxidized isoniazid derivatives as observed when the KatG enzyme was used. However, upon metal coordination, a significant enhancement in the formation of isonicotinic acid was observed compared with that of isonicotinamide. These results suggest that the pathway of a carbonyl-centered radical might be favored upon coordination to the Fe(II) owing to the π-back-bonding effect promoted by this metal center; therefore, the isoniazid metal complex could serve as a potential metallodrug. Enzymatic inhibition assays conducted with InhA showed that the cyanoferrate moiety is not the major player involved in this inhibition but the presence of isoniazid is required in this process. Other isoniazid metal complexes, [Ru(CN)(5)(izd)](3-) and [Ru(NH(3))(5)(izd)](2+) (where izd is isoniazid), were also unable to inhibit InhA, supporting our proposed self-activating mechanism of action. We propose that isoniazid reactivity can be rationally modulated by metal coordination chemistry, leading to the development of novel anti-TB metallodrugs.
Assuntos
Antituberculosos/química , Antituberculosos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Desenho de Fármacos , Isoniazida/química , Isoniazida/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Oxirredutases/antagonistas & inibidores , Farmacorresistência Bacteriana , Compostos Férricos/química , Humanos , Mycobacterium tuberculosis/enzimologia , Oxirredução , Tuberculose/tratamento farmacológico , Tuberculose/enzimologiaRESUMO
This study aimed to investigate the synthesis and potential vasodilator effect of a novel ruthenium complex, cis-[Ru(bpy)2(2-MIM)(NO2)]PF6 (bpy = 2,2'-bipyridine and 2-MIM = 2-methylimidazole) (FOR711A), containing an imidazole derivative via an in silico molecular docking model using ß1 H-NOX (Heme-nitric oxide/oxygen binding) domain proteins of reduced and oxidized soluble guanylate cyclase (sGC). In addition, pharmacokinetic properties in the human organism were predicted through computational simulations and the potential for acute irritation of FOR711A was also investigated in vitro using the hen's egg chorioallantoic membrane (HET-CAM). FOR711A interacted with sites of the ß1 H-NOX domain of reduced and oxidized sGC, demonstrating shorter bond distances to several residues and negative values of total energy. The predictive study revealed molar refractivity (RM): 127.65; Log Po/w = 1.29; topological polar surface area (TPSA): 86.26 Å2; molar mass (MM) = 541.55 g/mol; low solubility, high unsaturation index, high gastrointestinal absorption; toxicity class 4; failure to cross the blood-brain barrier and to react with cytochrome P450 (CYP) enzymes CYP1A2, CYP2C19, CYP2C9, CYP2D6 and CYP3A4. After the HET-CAM assay, the FOR711A complex was classified as non-irritant (N.I.) and its vasodilator effect was confirmed through greater evidence of blood vessels after the administration and ending of the observation period of 5 min. These results suggest that FOR711A presented a potential stimulator/activator effect of sGC via NO/sGC/cGMP. However, results indicate it needs a vehicle for oral administration.
Assuntos
Complexos de Coordenação/química , Óxido Nítrico/química , Rutênio/química , Vasodilatadores/química , Vasodilatadores/farmacologia , Animais , Galinhas , Membrana Corioalantoide/metabolismo , Heme/química , Humanos , Imidazóis/química , Simulação de Acoplamento Molecular/métodos , Óxido Nítrico/metabolismo , Oxigênio/química , Domínios Proteicos , Guanilil Ciclase Solúvel/química , Guanilil Ciclase Solúvel/metabolismoRESUMO
Nitric oxide (NO) has emerged as a promising antibacterial agent, where NO donor compounds have been explored. Here, we investigated the role of a silica nanoparticle containing nitroprusside (MPSi-NP) as a NO donor agent against methicillin-sensitive (ATCC 25,923 and ATCC 12228) and methicillin-resistant (ATCC 700,698 and ATCC 35984) Staphylococcus strains. Biofilm inhibition was studied along with antibiotic activity in combination with standard antibiotics (ampicillin and tetracycline). MPSi-NP exhibited thermal release of 63% of NO within 24 h, while free nitroprusside released only 18% during a dialysis assay, indicating an assisted release of NO mediated by the nanoparticles. This nanomaterial showed only a moderate activity in blocking biofilm production, but exhibited a significant decrease in the number of viable bacterial cells (over 600-fold for Staphylococcus aureus ATCC 700,698 and Staphylococcus epidermidis ATCC 35984). Remarkably, even using MPSi-NP at concentrations below any antibacterial action, its combination with ampicillin promoted a significant decrease in MIC for resistant strains of S. aureus ATCC 700,698 (2-fold) and S. epidermidis ATCC 35,984 (4-fold). A carbopol-based gel formulation with MPSi-NP (0.5% w/w) was prepared and showed a zone of inhibition of 7.7 ± 0.6 mm for S. epidermidis ATCC 35984. Topical use of MPSi-NP in combination with antibiotics might be a manageable strategy to prevent and eventually treat complicated resistant bacterial infections.
Assuntos
Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Antibacterianos/farmacologia , Biofilmes , Humanos , Testes de Sensibilidade Microbiana , Doadores de Óxido Nítrico/farmacologia , Diálise Renal , Staphylococcus aureusRESUMO
We aimed at evaluating the anti-asthmatic effect of cis-[Ru(bpy)2(2-MIM)(NO)](PF6)3 (FOR811A), a nitrosyl-ruthenium compound, in a murine model of allergic asthma. The anti-asthmatic effects were analyzed by measuring the mechanical lung and morphometrical parameters in female Swiss mice allocated in the following groups: untreated control (Ctl+Sal) and control treated with FOR811A (Ctl+FOR), along asthmatic groups untreated (Ast+Sal) and treated with FOR811A (Ast+FOR). The drug-protein interaction was evaluated by in-silico assay using molecular docking. The results showed that the use of FOR811A in experimental asthma (Ast+FOR) decreased the pressure-volume curve, hysteresis, tissue elastance, tissue resistance, and airway resistance, similar to the control groups (Ctl+Sal; Ctl+FOR). However, it differed from the untreated asthmatic group (Ast+Sal, p<0.05), indicating that FOR811A corrected the lung parenchyma and relaxed the smooth muscles of the bronchi. Similar to control groups (Ctl+Sal; Ctl+FOR), FOR811A increased the inspiratory capacity and static compliance in asthmatic animals (Ast+Sal, p<0.05), showing that this metallodrug improved the capacity of inspiration during asthma. The morphometric parameters showed that FOR811A decreased the alveolar collapse and kept the bronchoconstriction during asthma. Beyond that, the molecular docking using FOR811A showed a strong interaction in the distal portion of the heme group of the soluble guanylate cyclase, particularly with cysteine residue (Cys141). In summary, FOR811A relaxed bronchial smooth muscles and improved respiratory mechanics during asthma, providing a protective effect and promising use for the development of an anti-asthmatic drug.
Assuntos
Antiasmáticos , Asma , Doadores de Óxido Nítrico , Compostos Organometálicos , Mecânica Respiratória/efeitos dos fármacos , Rutênio , Animais , Antiasmáticos/química , Antiasmáticos/farmacologia , Asma/tratamento farmacológico , Asma/fisiopatologia , Feminino , Camundongos , Doadores de Óxido Nítrico/química , Doadores de Óxido Nítrico/farmacologia , Compostos Organometálicos/química , Compostos Organometálicos/farmacologia , Rutênio/química , Rutênio/farmacologiaRESUMO
The aim of this study was to investigate the antihypertensive properties of cis-[Ru(bpy)2ImN(NO)]3+ (FOR0811) in normotensive and in Nω-nitro-L-arginine methyl ester (L-NAME)-induced hypertensive rats. Vasorelaxant effects were analyzed by performing concentration response curve to FOR0811 in rat aortic rings in the absence or presence of 1H-[1,2,4]-oxadiazolo-[4,3,-a]quinoxalin-1-one (ODQ), L-cysteine or hydroxocobalamin. Normotensive and L-NAME-hypertensive rats were treated with FOR0811 and the effects in blood pressure and heart rate variability in the frequency domain (HRV) were followed. FOR0811 induced relaxation in rat aortic rings. Neither endothelium removal nor L-cysteine altered the FOR0811 effects. However, the incubation with ODQ and hydroxocobalamin completely blunted FOR0811 effects. FOR0811 administered intravenously by bolus infusion (0.01-1 mg/bolus) or chronically by using subcutaneous implanted osmotic pumps significantly reduced the mean arterial blood pressure. The effect was long lasting and did not induce reflex tachycardia. FOR0811 prevented both LF and VLF increases in L-NAME hypertensive rats and has antihypertensive properties. This new ruthenium complex compound might be a promising nitric oxide donor to treat cardiovascular diseases.
Assuntos
Anti-Hipertensivos/farmacologia , Hipertensão/tratamento farmacológico , Doadores de Óxido Nítrico/farmacologia , Compostos Organometálicos/farmacologia , Rutênio/farmacologia , Vasodilatadores/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , Hipertensão/induzido quimicamente , Masculino , NG-Nitroarginina Metil Éster/administração & dosagem , Ratos , Ratos WistarRESUMO
Inflammation is a physiological process triggered in response to tissue damage, and involves events related to cell recruitment, cytokines release and reactive oxygen species (ROS) production. Failing to control the process duration lead to chronification and may be associated with the development of various pathologies, including autoimmune diseases and cancer. Considering the pharmacological potential of metal-based compounds, two new ruthenium complexes were synthesized: cis-[Ru(NO2)(bpy)2(5NIM)]PF6 (1) and cis-[RuCl(bpy)2(MTZ)]PF6 (2), where bpyâ¯=â¯2,2'-bipyridine, 5NIMâ¯=â¯5-nitroimidazole and MTZâ¯=â¯metronidazole. Both products were characterized by spectroscopic techniques, followed by Density Functional Theory (DFT) calculations in order to support experimental findings. Afterwards, their in vitro cytotoxic, antioxidant and anti-inflammatory activities were investigated. Compounds 1 and 2 presented expressive in vitro antioxidant activity, reducing lipid peroxidation and decreasing intracellular ROS levels with comparable effectiveness to the standard steroidal drug dexamethasone or α-tocopherol. These complexes showed no noticeable cytotoxicity on the tested cancer cell lines. Bactericidal assay against metronidazole-resistant Helicobacter pylori, a microorganism able to disrupt oxidative balance, unraveled compound 1 moderate activity over that strain. Besides this, it was able to inhibit interleukin-6 (IL-6) and tumor necrosis factor-α (TNF- α) production as well as interleukin-1ß (IL-1ß) and cyclooxygenase-2 (COX-2) expression in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. This latter activity is remarkable, which has not been reported for other ruthenium-based complexes. Altogether, these results suggest cis-[Ru(NO2)(bpy)2(5NIM)]PF6 complex has potential pharmacological application as an anti-inflammatory agent that deserve further biological investigation.
Assuntos
Antibacterianos/farmacologia , Anti-Inflamatórios/farmacologia , Antineoplásicos/farmacologia , Antioxidantes/farmacologia , Complexos de Coordenação/farmacologia , Imidazóis/química , Rutênio/química , Células A549 , Animais , Antibacterianos/química , Anti-Inflamatórios/química , Antineoplásicos/química , Antioxidantes/química , Bactérias/efeitos dos fármacos , Proliferação de Células , Complexos de Coordenação/química , Humanos , Peroxidação de Lipídeos , Células MCF-7 , Camundongos , Estrutura Molecular , Células RAW 264.7 , Superóxidos/metabolismoRESUMO
Nitric oxide (NO) and nitroxyl (HNO) have gained broad attention due to their roles in several physiological and pathophysiological processes. Remarkably, these sibling species can exhibit opposing effects including the promotion of angiogenic activity by NO compared to HNO, which blocks neovascularization. While many NO donors have been developed over the years, interest in HNO has led to the recent emergence of new donors. However, in both cases there is an expressive lack of iron-based compounds. Herein, we explored the novel chemical reactivity and stability of the trans-[Fe(cyclam)(NO)Cl]Cl2 (cyclam = 1,4,8,11-tetraazacyclotetradecane) complex. Interestingly, the half-life (t1/2) for NO release was 1.8 min upon light irradiation, vs 5.4 h upon thermal activation at 37 °C. Importantly, spectroscopic evidence supported the generation of HNO rather than NO induced by glutathione. Moreover, we observed significant inhibition of NO donor- or hypoxia-induced HIF-1α (hypoxia-inducible factor 1α) accumulation in breast cancer cells, as well as reduced vascular tube formation by endothelial cells pretreated with the trans-[Fe(cyclam)(NO)Cl]Cl2 complex. Together, these studies provide the first example of an iron-nitrosyl complex with anti-angiogenic activity as well as the potential dual activity of this compound as a NO/HNO releasing agent, which warrants further pharmacological investigation.
Assuntos
Inibidores da Angiogênese/farmacologia , Complexos de Coordenação/farmacologia , Doadores de Óxido Nítrico/farmacologia , Inibidores da Angiogênese/síntese química , Inibidores da Angiogênese/efeitos da radiação , Animais , Linhagem Celular Tumoral , Complexos de Coordenação/síntese química , Complexos de Coordenação/efeitos da radiação , Glutationa/química , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Ferro/química , Ferro/efeitos da radiação , Camundongos , Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/síntese química , Doadores de Óxido Nítrico/efeitos da radiação , Óxidos de Nitrogênio/metabolismo , Ratos , Temperatura , Raios Ultravioleta , Vasodilatadores/síntese química , Vasodilatadores/farmacologia , Vasodilatadores/efeitos da radiaçãoRESUMO
Leishmaniasis is a parasitic disease caused by protozoa of the genus Leishmania. There are many complications presented by the current treatment, as high toxicity, high cost and parasite resistance, making the development of new therapeutic agents indispensable. The present study aims to evaluate the leishmanicidal potential of ruthenium nitrosyl complex cis-[Ru(bpy)2(SO3)(NO)](PF6) against Leishmania (Viannia) braziliensis. The effect of this metal complex on parasite-host interaction was evaluated by in vitro efficacy test in dermal fibrobast cells in the presence of different concentrations (1, 10, 50 and 100 µM) and by in vivo efficacy tests performed in the presence of two different concentrations of complex (100 µg/kg/day or 300 µg/kg/day) evaluating its effect on the size of the lesion and the number of parasites present in the draining lymph nodes in hamsters. Even at the lowest concentration of 1 µM of ruthenium complex, it was observed a significant decrease of the infected cells, after 24 h exposure in vitro, with total reduction at 50 µM of the ruthenium complex. In the in vivo cutaneous infection model, administration of daily doses of 300 µg/kg/day of complex reduced significantly lesion size by 51% (p < 0.05), with a 99.9% elimination of the parasites found in the lymph nodes (p < 0.001). The results suggest a promising leishmanicidal effect by that ruthenium nitrosyl complex against L. (V.) braziliensis.
Assuntos
Leishmania braziliensis/efeitos dos fármacos , Compostos de Rutênio/farmacologia , Animais , Cricetinae , Relação Dose-Resposta a Droga , Interações Hospedeiro-Parasita , PeleRESUMO
Heme-based sensors are a recently discovered functional class of heme proteins that serve to detect physiological fluctuations in oxygen (O(2)), carbon monoxide (CO), or nitric oxide (NO). Many of these modular sensors detect heme ligands by coupling a histidine-protein kinase to a heme-binding domain. They typically bind O2, CO, and NO but respond only to one of these ligands. Usually, they are active in the ferrous unliganded state but are switched off by saturation with O2. The heme-binding domains of these kinases are quite varied. They may feature a PAS fold, as in the Bradyrhizobium japonicum and Sinorhizobium melitoti FixL proteins, or a GAF fold, as in the Mycobacterium tuberculosis DevS and DosT proteins. Alternative folds, such as HNOB (also H-NOX), have also been noted for such signal-transducing kinases, although these classes are less well studied. Histidine-protein kinases function in partnership with cognate response-regulator substrate(s): usually transcription factors that they activate by phosphorylation. For example, FixL proteins specifically phosphorylate their FixJ partners, and DevS and DosT proteins phosphorylate DevR in response to hypoxia. We present methods for purifying these sensors and their protein substrates, verifying the quality of the preparations, determining the K(d) values for binding of ligand and preparing sensors of known saturation, and measuring the rates of turnover (k(cat)) of the protein substrate by sensors of known heme status.
Assuntos
Oxigênio/análise , Proteínas Quinases/análise , Proteínas Quinases/metabolismo , Proteínas de Bactérias/análise , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Hemeproteínas/análise , Hemeproteínas/fisiologia , Histidina Quinase , Ligantes , Oxigênio/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Fosforilação , Ligação Proteica , Proteínas Quinases/isolamento & purificação , Controle de Qualidade , Titulometria/métodosRESUMO
The emergence of multidrug-resistant strains of Mycobacterium tuberculosis (MTB) represents a major threat to global health. Isoniazid (INH) is a prodrug used in the first-line treatment of tuberculosis. It undergoes oxidation by a catalase-peroxidase KatG, leading to generation of an isonicotinoyl radical that reacts with NAD(H) forming the INH-NADH adduct as the active metabolite. A redox-mediated activation of isoniazid using an iron metal complex was previously proposed as a strategy to overcome isoniazid resistance due to KatG mutations. Here, we have prepared a series of iron metal complexes with isoniazid and analogues, containing alkyl substituents at the hydrazide moiety, and also with pyrazinamide derivatives. These complexes were activated by H2O2 and studied by ESR and LC-MS. For the first time, the formation of the oxidized INH-NAD adduct from the pentacyano(isoniazid)ferrate(II) complex was detected by LC-MS, supporting a redox-mediated activation, for which a mechanistic proposition is reported. ESR data showed all alkylated hydrazides, in contrast to non-substituted hydrazides, only generated alkyl-based radicals. The structural modifications did not improve minimal inhibitory concentration (MIC) against MTB in comparison to isoniazid iron complex, providing support to isonicotinoyl radical formation as a requirement for activity. Nonetheless, the pyrazinoic acid hydrazide iron complex showed redox-mediated activation using H2O2 with generation of a pyrazinoyl radical intermediate and production of pyrazinoic acid, which is in fact the active metabolite of pyrazinamide prodrug. Thereby, this strategy can also unveil new opportunities for activation of this type of drug.
Assuntos
Antituberculosos/farmacologia , Complexos de Coordenação/farmacologia , Compostos Ferrosos/farmacologia , Isoniazida/análogos & derivados , Isoniazida/farmacologia , Antituberculosos/síntese química , Antituberculosos/química , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Compostos Ferrosos/síntese química , Compostos Ferrosos/química , Isoniazida/síntese química , Isoniazida/química , Testes de Sensibilidade Microbiana , Modelos Químicos , Mycobacterium tuberculosis/efeitos dos fármacos , OxirreduçãoRESUMO
Exposure of Mycobacterium tuberculosis to hypoxia is known to alter the expression of many genes, including ones thought to be involved in latency, via the transcription factor DevR (also called DosR). Two sensory kinases, DosT and DevS (also called DosS), control the activity of DevR. We show that, like DevS, DosT contains a heme cofactor within an N-terminal GAF domain. For full-length DosT and DevS, we determined the ligand-binding parameters and the rates of ATP reaction with the liganded and unliganded states. In both proteins, the heme state was coupled to the kinase such that the unliganded, CO-bound, and NO-bound forms were active, but the O(2)-bound form was inactive. Oxygen-bound DosT was unusually inert to oxidation to the ferric state (half life in air >60 h). Though the kinase activity of DosT was unaffected by NO, this ligand bound 5000 times more avidly than O(2) to DosT (K(d) [NO] approximately 5 nM versus K(d) [O(2)] = 26 microM). These results demonstrate direct and specific O(2) sensing by proteins in M. tuberculosis and identify for the first time a signal ligand for a sensory kinase from this organism. They also explain why exposure of M. tuberculosis to NO donors under aerobic conditions can give results identical to hypoxia, i.e., NO saturates DosT, preventing O(2) binding and yielding an active kinase.
Assuntos
Proteínas de Bactérias/química , Mycobacterium tuberculosis/enzimologia , Oxigênio/metabolismo , Protamina Quinase/química , Proteínas Quinases/química , Trifosfato de Adenosina/química , Trifosfato de Adenosina/metabolismo , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Monóxido de Carbono/química , Monóxido de Carbono/metabolismo , Cátions Bivalentes/química , Cátions Bivalentes/metabolismo , Heme/química , Heme/metabolismo , Cinética , Ligantes , Modelos Biológicos , Óxido Nítrico/química , Óxido Nítrico/metabolismo , Oxirredução , Protamina Quinase/isolamento & purificação , Protamina Quinase/metabolismo , Proteínas Quinases/isolamento & purificação , Proteínas Quinases/metabolismo , Estrutura Terciária de ProteínaRESUMO
In oxygen-sensing PAS domains, a conserved polar residue on the proximal side of the heme cofactor, usually arginine or histidine, interacts alternately with the protein in the "on-state" or the heme edge in the "off-state" but does not contact the bound ligand directly. We assessed the contributions of this residue in Bradyrhizobium japonicum FixL by determining the effects of an R206A substitution on the heme-PAS structure, ligand affinity, and regulatory capacity. The crystal structures of the unliganded forms of the R206A and wild-type BjFixL heme-PAS domains were similar, except for a more ruffled porphyrin ring in R206A BjFixL and a relaxation of the H214 residue and heme propionate 7 due to their lost interactions. The oxygen affinity of R206A BjFixL (Kd approximately 350 microM) was 2.5 times lower than that of BjFixL, and this was due to a higher off-rate constant for the R206A variant. The enzymatic activities of the unliganded "on-state" forms, either deoxy or met-R206A BjFixL, were comparable to each other and slightly lower (twofold less) than those of the corresponding BjFixL species. The most striking difference between the two proteins was in the enzymatic activities of the liganded "off-state" forms. In particular, saturation with a regulatory ligand (the Fe(III) form with cyanide) caused a >2000-fold inhibition of the BjFixL phosphorylation of BjFixJ, but a 140-fold inhibition of this catalytic activity in R206A BjFixL. Thus, in oxygen-sensing PAS domains, the interactions of polar residues with the heme edge couple the heme-binding domain to a transmitter during signal transduction.
Assuntos
Arginina/química , Proteínas de Bactérias/química , Bradyrhizobium/metabolismo , Hemeproteínas/química , Oxigênio/metabolismo , Sequência de Aminoácidos , Escherichia coli/metabolismo , Histidina/química , Histidina Quinase , Cinética , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Homologia de Sequência de Aminoácidos , Transdução de SinaisRESUMO
Heme-based sensors have emerged during the last 20years as being a large family of proteins that occur in all kingdoms of life. A myriad of biological adaptations are associated with these sensors, which include vasodilation, bacterial virulence, dormancy, chemotaxis, biofilm formation, among others. Due to the key activities regulated by these proteins along with many other systems that use similar output domains, there is a growing interest in developing small molecules as their regulators. Here, we review the development of potential activators and inhibitors for many of these systems, including human soluble guanylate cyclase, c-di-GMP-related enzymes, Mycobacterium tuberculosis DevR/DevS/DosT (differentially expressed in virulent strain response regulator/sensor/dormancysurvival sensorT), the Rev-erb-α and ß nuclear receptor, among others. The possible roles of these molecules as biochemical tools, therapeutic agents, and novel antibiotics are critically examined.