Radical production by hydrogen peroxide/bicarbonate and copper uptake in mammalian cells: modulation by Cu(II) complexes

The presence of the bicarbonate/carbon dioxide pair is known to accelerate the transition metal ion-catalysed oxidation of various biotargets. It has been shown that stable Cu(II) complexes formed with imine ligands that allow redox cycling between Cu(I) and Cu(II) display diverse apoptotic effects on cell cultures. It is also reported that Cu(II)-tetraglycine can form a stable Cu(III) complex. In the present study, radical generation from H₂O₂ and H₂O₂/HCO₃⁻ in the presence of these two different classes of Cu(II) complexes was evaluated by monitoring the oxidation of dihydrorhodamine 123 and NADH and by the quantitative determination of thiobarbituric acid reactive substances (TBARs method). Cu(II)-imine complexes produced low levels of reactive species whereas Cu(II)-Gly-derived complexes, as well as the free Cu(II) ion, produced oxygen-derived radicals in significantly larger amounts. The effects of these two classes of complexes on mammalian tumour cell viability were equally distinct, in that Cu(II)-imine complexes caused apoptosis, entered in cell and remained almost unaffected in high levels whilst, at the same concentrations, Cu(II)-Gly peptide complexes and Cu(II) sulphate stimulated cell proliferation, with the cell managing copper efficiently. Taken together, these results highlight the different biological effects of Cu(II) complexes, some of which have been recently studied as anti-tumour drugs and radical system generators, and also update the effects of reactive oxygen species generation on cell cycle control.     

Signal transduction by members of the transforming growth factor-β superfamily

Transforming growth factor-β (TGFβ) superfamily members exert their diverse biological effects through their interaction with heteromeric receptor complexes of transmembrane serine/threonine kinases. Both components of the receptor complex, known as receptor I and receptor II are essential for signal transduction. The composition of these complexes can vary significantly due to the promiscuous nature of the ligands and the receptors, and this diversity of interactions can yield a variety of biological responses. Several receptor interacting proteins and potential mediators of signal transduction have now been identified. Recent advances, particularly in our understanding of the function of Mothers against dpp-related (MADR) proteins, are providing new insights into how the TGFβ superfamily signals its diverse biological activities.     

Medicinal applications of vanadium complexes with Schiff bases

Many transition metal complexes have been explored for their therapeutic properties after the discovery of cisplatin. Schiff bases have an efficient complexation tendency with the transition metals and several medicinal properties have been reported. However, fewer studies have reported the medicinal utility of vanadium and its Schiff base complexes. This paper provides a comprehensive overview of vanadium complexes with Schiff bases along with their mechanistic insight. Vanadium complexes in + 4 and + 5 oxidation states have exhibited well-defined geometry and found to be thermodynamically stable. The studies have reported the G0/G1 phase cell cycle arrest and decreased delta psi m, inducing mitochondrial membrane depolarization in cancer cell lines along with the alterations in the metabolism of the cancer cells upon dosing with the vanadium complexes. Cancer cell invasion and growth are also found to be markedly reduced by peroxo complexes of vanadium. The studies included in the review paper have been taken from leading indexing databases and focus was laid on recent reports in literature. The biological potential of vanadium complexes of Schiff bases opens new horizons for future interdisciplinary studies and investigation focussed on understanding the biochemistry of these complexes, along with designing new complexes which have better bioavailability, solubility and low or non-toxicity.     

Cyclohexane 1,3-diones and their inhibition of mutant SOD1-dependent protein aggregation and toxicity in PC12 cells

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons. Currently, there is only one FDA-approved treatment for ALS (riluzole), and that drug only extends life, on average, by 2-3 months. Mutations in Cu/Zn superoxide dismutase (SOD1) are found in familial forms of the disease and have played an important role in the study of ALS pathophysiology. On the basis of their activity in a PC12-G93A-YFP high-throughput screening assay, several bioactive compounds have been identified and classified as cyclohexane-1,3-dione (CHD) derivatives. A concise and efficient synthetic route has been developed to provide diverse CHD analogs. The structural modification of the CHD scaffold led to the discovery of a more potent analog (26) with an EC(50) of 700 nM having good pharmacokinetic properties, such as high solubility, low human and mouse metabolic potential, and relatively good plasma stability. It was also found to efficiently penetrate the blood-brain barrier. However, compound 26 did not exhibit any significant life span extension in the ALS mouse model. It was found that, although 26 was active in PC12 cells, it had poor activity in other cell types, including primary cortical neurons, indicating that it can penetrate into the brain, but is not active in neuronal cells, potentially due to poor selective cell penetration. Further structural modification of the CHD scaffold was aimed at improving global cell activity as well as maintaining potency. Two new analogs (71 and 73) were synthesized, which had significantly enhanced cortical neuronal cell permeability, as well as similar potency to that of 26 in the PC12-G93A assay. These CHD analogs are being investigated further as novel therapeutic candidates for ALS.     

Complexes of salicylaldehyde acylhydrazones: cytotoxicity, QSAR and crystal structure of the sterically hindered t-butyl dimer

A series of acylhydrazones of salicylaldehyde and their transition metal complexes, predominantly copper(II), have been prepared and characterized. The crystal structure of the Cu(II) complex of the sterically hindered t-butyl derivative contains a phenolato bridged dimer with the ligand coordinated as a tridentate moiety. QSAR analyses of the cytotoxicity of the chelators and their Cu(II) complexes reveals that solubility is the dominant factor for activity. Compounds display a maximum with respect to lipophilicity, allowing optimization of the bioactivity for both the ligands and their complexes. Copper complexes are significantly more cytotoxic than the metal-free ligands and complexes of other metals: Cu > Ni > Zn = Mn > Fe = Cr > Cr > Co.     

Copper(II) complexes with sparfloxacin and nitrogen-donor heterocyclic ligands: Structure-activity relationship

Three novel neutral mononuclear copper(II) complexes of the third-generation quinolone antibacterial drug sparfloxacin in the presence of a nitrogen donor heterocyclic ligand 2,2'-bipyridine, 1,10-phenanthroline or 2,2'-dipyridylamine have been prepared and characterized physicochemically and spectroscopically. The resultant complexes are of the type Cu(sparfloxacinato)(N-donor)Cl. Copper(II) is pentacoordinate having a distorted square pyramidal geometry. Molecular modeling calculations have been performed in order to propose the lowest energy model structure of the complexes. The interaction of the complexes with calf-thymus DNA has been investigated with diverse spectroscopic techniques and has shown that the complexes can bind to calf-thymus DNA by the intercalative mode. The antimicrobial activity of the complexes has been tested on three different microorganisms. The Cu(sparfloxacinato)(N-donor)Cl complexes are among the most active ones against Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus, when compared to the other corresponding copper-quinolone complexes studied by our group and their antimicrobial activity is increased in the order bipyam相似文献   

Extracellular vesicles and pancreatitis: mechanisms,status and perspectives

Comprehensive reviews and large population-based cohort studies have played an important role in the diagnosis and treatment of pancreatitis and its sequelae. The incidence and mortality of pancreatitis have been reduced significantly due to substantial advancements in the pathophysiological mechanisms and clinically effective treatments. The study of extracellular vesicles (EVs) has the potential to identify cell-to-cell communication in diseases such as pancreatitis. Exosomes are a subset of EVs with an average diameter of 50~150 nm. Their diverse and unique constituents include nucleic acids, proteins, and lipids, which can be transferred to trigger phenotypic changes of recipient cells. In recent years, many reports have indicated the role of EVs in pancreatitis, including acute pancreatitis, chronic pancreatitis and autoimmune pancreatitis, suggesting their potential influence on the development and progression of pancreatitis. Plasma exosomes of acute pancreatitis can effectively reach the alveolar cavity and activate alveolar macrophages to cause acute lung injury. Furthermore, upregulated exosomal miRNAs can be used as biomarkers for acute pancreatitis. Here, we summarized the current understanding of EVs in pancreatitis with an emphasis on their biological roles and their potential use as diagnostic biomarkers and therapeutic agents for this disease.     

Bis(thiosemicarbazone) copper complexes: mechanism of intracellular accumulation

The molecular basis of Alzheimer’s disease has not been clearly established, but disruption of brain metal ion homeostasis, particularly copper and zinc, might be closely involved in the pathogenesis of this disease and its characteristic β-amyloid neuropathological features. The use of complexes of copper with bis(thiosemicarbazones) ([Cu(btsc)]) has been proposed for the treatment of Alzheimer’s disease. Their mode of action could involve modulation of the concentration of copper or zinc, and it has been suggested that the compounds can modulate the production of β-amyloid peptide at the neuron level. Furthermore, it has been reported that [Cu(btsc)] complexes can be reduced inside the cells. However, to our knowledge the intracellular reduction of these compounds has never been demonstrated. Thus, the goal of our study was to increase understanding of the mechanism of intracellular accumulation of [Cu(btsc)] complexes. Our results reveal that the intracellular concentration of copper inside the cells is very high and that these compounds are not P-glycoprotein substrates. This protein is a key element of the low permeability properties of the blood–brain barrier. Furthermore, no intracellular reduction of cupric ions was detected. Finally, once inside the cells, the complexes undergo aggregation, strongly suggesting that aggregation of complexes is the driving force responsible for their intracellular accumulation.     

Subcellular localization of a fluorescent derivative of CuII(atsm) offers insight into the neuroprotective action of CuII(atsm)

Copper complexes of bis(thiosemicarbazone) (Cu(II)(btsc)s) have been studied as potential anti-cancer agents and hypoxia imaging agents. More recently, Cu(II)(btsc)s have been identified as possessing potent neuroprotective properties in cell and animal models of neurodegenerative disease. Despite their broad range of pharmacological activity little is known about how cells traffic Cu(II)(btsc)s and how this relates to potential anti-cancer or neuroprotective outcomes. One method of investigating sub-cellular localization of metal complexes is through confocal fluorescence imaging of the compounds in cells. Previously we harnessed the fluorescence of a pyrene group attached to diacetyl-bis(N4-methylthiosemicarbazonato)copper(ii)) (Cu(II)(atsm)), (Cu(II)L(1)). We demonstrated that Cu(II)L(1) was partially localized to lysosomes in HeLa cancer epithelial cells. Here we extend these studies to map the sub-cellular localization of Cu(II)L(1) in M17 neuroblastoma cells. Treatment of M17 or HeLa cells led to rapid association of the Cu-complex into distinct punctate structures that partially co-localized with lysosomes as assessed by co-localization with Lysotracker and acridine orange. No localization to early or late endosomes, the nucleus or mitochondria was observed. We also found evidence for a limited association of Cu(II)L(1) with autophagic structures, however, this did not account for the majority of the punctate localization of Cu(II)L(1). In addition, Cu(II)L(1) revealed partial localization with ER Tracker and was found to inhibit ER stress induced by tunicamycin. This is the first report to comprehensively characterize the sub-cellular localization of a Cu(II)(atsm) derivative in cells of a neuronal origin and the partial association with lysosome/autophagic structures and the ER may have a potential role in neuroprotection.     

Attaching histidine-tagged peptides and proteins to lipid-based carriers through use of metal-ion-chelating lipids

The therapeutic potential of selected peptides and proteins is enormous, with applications ranging from use as therapeutic vaccines, as modulators of intracellular signaling pathways and as highly selective agents capable of recognizing unique extracellular targets. We have been pursuing development of hybrid lipid-based carrier formulations designed to take advantage of the therapeutic benefits of peptides selected for their ability to act in a complementary fashion with the carrier system. In this regard, it is critical to have simple and versatile methods to promote and control the binding of diverse peptides to a broad range of carrier formulations. As demonstrated here, recombinant proteins and synthetic peptides containing poly-histidine residues (4 to 10) can be specifically bound to liposomes containing a metal-ion-chelating lipid, DOGS-NTA-Ni. The potential of this approach is demonstrated using two functional peptides, AntpHD-Cw3 (applications for vaccine production) and AHNP (specificity for Her-2 expressing cells).     

Cytotoxicity of salicylaldehyde benzoylhydrazone analogs and their transition metal complexes: quantitative structure-activity relationships

A series of salicylaldehyde benzoylhydrazone derivatives, their copper(II) complexes and a range of transition metal complexes of the unsubstituted ligand has been synthesized and evaluated for cytotoxicity against a human adenocarcinoma cell line. A QSAR analysis revealed ligand cytotoxicity is strongly correlated with electronic and transport factors and can be modeled by treating each 'half' of the molecule as an isolated unit. Activity increases when substituents in the benzoyl ring were electron withdrawing whereas, for the salicylaldehyde ring, electron donation was required. The cytotoxicity of the Cu(II) complexes was greater than, and paralleled the ligands. Activity for the transition metal complexes of the unsubstituted ligand mirrored charge density on the metal.     

A systemic insight into astrocytoma biology across different grades

Astrocytomas are the most common type of brain tumors, which originate from glial cells, and are classified into specific grades based on their histopathological behavior. To develop precise therapeutic strategies for the disease, it is important to identify the molecular signatures specific to each grade as well as the key factors responsible for the transition from one grade to the next. In this study, we have taken a systems approach to investigate the gene expression profiles of each grades of the disease by mapping shortest paths of gene interaction in each grade and also between one grade and the next. Module core genes govern the topology of these networks and serve as important functional players in each grade as well as help in grade transition events. Shortlisted among these module core genes are well-characterized players of glioma as well as novel molecules (32 grade-specific genes and 15 grade transition–specific genes), which influence important prooncogenic functions but have not been linked to glioma biology yet. These module core genes provide interesting insight into the biology of astrocytic tumors and are potential therapeutic targets for astrocytoma.     

ADAM-17: the enzyme that does it all

This review focuses on the role of ADAM-17 in disease. Since its debut as the tumor necrosis factor converting enzyme (TACE), ADAM-17 has been reported to be an indispensible regulator of almost every cellular event from proliferation to migration. The central role of ADAM-17 in cell regulation is rooted in its diverse array of substrates: cytokines, growth factors, and their receptors as well as adhesion molecules are activated or inactivated by their cleavage with ADAM-17. It is therefore not surprising that ADAM-17 is implicated in numerous human diseases including cancer, heart disease, diabetes, rheumatoid arthritis, kidney fibrosis, Alzheimer’s disease, and is a promising target for future treatments. The specific role of ADAM-17 in the pathophysiology of these diseases is very complex and depends on the cellular context. To exploit the therapeutic potential of ADAM-17, it is important to understand how its activity is regulated and how specific organs and cells can be targeted to inactivate or activate the enzyme.     

Postsynaptic signaling at glutamatergic synapses as therapeutic targets

Dysregulation of glutamatergic synapses plays an important role in the pathogenesis of neurological diseases. In addition to mediating excitatory synaptic transmission, postsynaptic glutamate receptors interact with various membrane and intracellular proteins. They form structural and/or signaling synaptic protein complexes and thereby play diverse postsynaptic functions. Recently, several postsynaptic protein complexes have been associated with various neurological diseases and hence, have been characterized as important therapeutic targets. Moreover, novel small molecules and therapeutic peptides targeting and modulating the activities of these protein complexes have been discovered, some of which have advanced through preclinical translational research and/or clinical studies. This article describes the recent investigation of eight key protein complexes associated with the postsynaptic ionotropic and metabotropic glutamate receptors as therapeutic targets for central nervous system diseases.     

Towards the treatment of saphenous vein bypass graft failure--a perspective of the Bristol Heart Institute

Coronary artery bypass graft surgery (CABG) is widely used for the treatment of atheromatous stenosis of coronary arteries. However, as many as 50% of grafts fail within 10 years after CABG due to neointima (NI) formation, a process involving the proliferation and migration of vascular smooth muscle cells (VSMCs). Superimposed on neointima formation is accelerated atherogenesis which ultimately results in late vein graft failure. To date no therapeutic intervention has proved successful in treating late vein graft failure and as such is a matter of some urgency. However, in recent years, several diverse approaches aimed at preventing neointimal formation have been devised which have yielded promising results. These include the use of external stents, gene therapy as well as conventional pharmacological interventions. The objective of this article, therefore, is to review these recent approaches and their potential clinical applications in the treatment of vein graft disease.     

Parameters determining the relative efficacy of hydroxy-naphthoquinone inhibitors of the cytochrome bc1 complex

Hydroxy-naphthoquinones are competitive inhibitors of the cytochrome bc(1) complex that bind to the ubiquinol oxidation site between cytochrome b and the iron-sulfur protein and presumably mimic a transition state in the ubiquinol oxidation reaction catalyzed by the enzyme. The parameters that affect efficacy of binding of these inhibitors to the bc(1) complex are not well understood. Atovaquone, a hydroxy-naphthoquinone, has been used therapeutically to treat Pneumocystis carinii and Plasmodium infections. As the pathogens have developed resistance to this drug, it is important to understand the molecular basis of the drug resistance and to develop new drugs that can circumvent the drug resistance. We previously developed the yeast and bovine bc(1) complexes as surrogates to model the interaction of atovaquone with the bc(1) complexes of the target pathogens and human host. As a first step to identify new cytochrome bc(1) complex inhibitors with therapeutic potential and to better understand the determinants of inhibitor binding, we have screened a library of 2-hydroxy-naphthoquinones with aromatic, cyclic, and non-cyclic alkyl side-chain substitutions at carbon-3 on the hydroxy-quinone ring. We found a group of compounds with alkyl side-chains that effectively inhibit the yeast bc(1) complex. Molecular modeling of these into the crystal structure of the yeast cytochrome bc(1) complex provides structural and quantitative explanations for their binding efficacy to the target enzyme. In addition we also identified a 2-hydroxy-naphthoquinone with a branched side-chain that has potential for development as an anti-fungal and anti-parasitic therapeutic.     

Synthesis and characterization of metal binding pseudotripeptides.

Metal complexes with peptide or pseudopeptide type ligands can serve as good model compounds for a deeper understanding of enzymatic catalysis, but ligands with a high selectivity for different transition metal cations are hard to find due to the rather flexible nature of peptides. Since such ligands would be the sine qua non condition for the synthesis of heterodinuclear peptide metal complexes with catalytic activity, the search for small, affine and selective metal chelating sequences is of interest. Using four different amino acids (His, Lys, Asp, Glu) a set of 16 pseudotripeptides of the common structure Bz-AS1-Sar-AS2-NH2 has been synthesized, purified and characterized by mass spectrometry and 1H-NMR. Their ability to form metal complexes has been investigated leading to short motifs capable of selectively binding only one or two transition metal cations with high affinity. As expected, the complexation of transition metal cations by pseudotripeptides is strongly dependent not only on the amino acid composition, but also on the sequence with regard to the stability of the resulting complexes, as well as the selectivity of the ligands towards Cu2+, Co2+, Ni2+, Zn2+ and Mn2+.     

Do the Copper Complexes of Histamine,Histidine and of Two H2-Antagonists React with O−2?

The effects of cimetidine, ranitidine, histamine and histidine. as well as of their copper complexes, have been examined in an enzymic and chemical O^?₂ generated systems. Copper complexes like CuZnSOD inhibited both the reduction of cytochrome c and NBT²⁺ in xanthine-xanthine oxidase systems, but their inhibitory action was due to a certain extent to the copper-induced inhibition of xanthine oxidase. EDTA abolished the inhibitory effect of all copper complexes studied. Luminol chemiluminescence in NADH,-PMS system was inhibited by CuZnSOD while it was enhanced by copper complexes. The copper-accelerating effect gradually increased up to about I μM Cu and decreased, reaching the control values up to 10 μM Cu. In the presence of low copper concentrations chemiluminescence was inhibited by CuZnSOD only, while in the presence of high copper concentrations it was inhibited by catalase and mannitol. but not by CuZnSOD. The ligands however, have been ineffective in the two O^?_2; generated systems.     

Membrane-targeted strategies for modulating APP and Aβ-mediated toxicity

Alzheimer's disease (AD) is a progressive neurodegenerative disease characterized by numerous pathological features including the accumulation of neurotoxic amyloid-β (Aβ) peptide. There is currently no effective therapy for AD, but the development of therapeutic strategies that target the cell membrane is gaining increased interest. The amyloid precursor protein (APP) from which Aβ is formed is a membrane-bound protein, and Aβ production and toxicity are both membrane mediated events. This review describes the critical role of cell membranes in AD with particular emphasis on how the composition and structure of the membrane and its specialized regions may influence toxic or benign Aβ/APP pathways in AD. The putative role of copper (Cu) in AD is also discussed, and we highlight how targeting the cell membrane with Cu complexes has therapeutic potential in AD.