Parkinson's disease: Alterations in iron and redox biology as a key to unlock therapeutic strategies.

A plethora of studies indicate that iron metabolism is dysregulated in Parkinson's disease (PD). The literature reveals well-documented alterations consistent with established dogma, but also intriguing paradoxical observations requiring mechanistic dissection. An important fact is the iron loading in dopaminergic neurons of the substantia nigra pars compacta (SNpc), which are the cells primarily affected in PD. Assessment of these changes reveal increased expression of proteins critical for iron uptake, namely transferrin receptor 1 and the divalent metal transporter 1 (DMT1), and decreased expression of the iron exporter, ferroportin-1 (FPN1). Consistent with this is the activation of iron regulator protein (IRP) RNA-binding activity, which is an important regulator of iron homeostasis, with its activation indicating cytosolic iron deficiency. In fact, IRPs bind to iron-responsive elements (IREs) in the 3ꞌ untranslated region (UTR) of certain mRNAs to stabilize their half-life, while binding to the 5ꞌ UTR prevents translation. Iron loading of dopaminergic neurons in PD may occur through these mechanisms, leading to increased neuronal iron and iron-mediated reactive oxygen species (ROS) generation. The "gold standard" histological marker of PD, Lewy bodies, are mainly composed of α-synuclein, the expression of which is markedly increased in PD. Of note, an atypical IRE exists in the α-synuclein 5ꞌ UTR that may explain its up-regulation by increased iron. This dysregulation could be impacted by the unique autonomous pacemaking of dopaminergic neurons of the SNpc that engages L-type Ca+2 channels, which imparts a bioenergetic energy deficit and mitochondrial redox stress. This dysfunction could then drive alterations in iron trafficking that attempt to rescue energy deficits such as the increased iron uptake to provide iron for key electron transport proteins. Considering the increased iron-loading in PD brains, therapies utilizing limited iron chelation have shown success. Greater therapeutic advancements should be possible once the exact molecular pathways of iron processing are dissected.

Antiplasmodial activity of the natural product compounds alstonine and himbeline.

Malaria, caused by Plasmodium parasites, continues to be a devastating global health issue. Despite a decline in malaria related deaths over the last decade, overall progress has plateaued. Key challenges to malaria prevention and control include the lack of a broadly effective vaccine and parasite drug resistance, including to the current gold standard artemisinin combination therapies (ACTs). New drugs with unique modes of action are therefore a priority for both the treatment and prevention of malaria. Unlike treatment drugs which need to kill parasites quickly to reduce or prevent clinical symptoms, compounds that kill parasites more slowly may be an option for malaria prevention. Natural products and natural product derived compounds have historically been an excellent source of antimalarial drugs, including the artemisinin component of ACTs. In this study, 424 natural product derived compounds were screened for in vitro activity against P. falciparum in assays designed to detect slow action activity, with 46 hit compounds identified as having >50% inhibition at 10 µM. Dose response assays revealed nine compounds with submicromolar activity, with slow action activity confirmed for two compounds, alstonine and himbeline (50% inhibitory concentration (IC50) 0.17 and 0.58 µM, respectively). Both compounds displayed >140-fold better activity against P. falciparum versus two human cell lines (Selectivity Index (SI) >1,111 and > 144, respectively). Importantly, P. falciparum multi-drug resistant lines showed no cross-resistance to alstonine or himbeline, with some resistant lines being more sensitive to these two compounds compared to the drug sensitive line. In addition, alstonine displayed cross-species activity against the zoonotic species, P. knowelsi (IC50 ~1 µM). Outcomes of this study provide a starting point for further investigations into these compounds as antiplasmodial drug candidates and the investigation of their molecular targets.

Evaluation of fermentation conditions triggering increased antibacterial activity from a near-shore marine intertidal environment-associated Streptomyces species.

A near-shore marine intertidal environment-associated Streptomyces isolate (USC-633) from the Sunshine Coast Region of Queensland, Australia, cultivated under a range of chemically defined and complex media to determine optimal parameters resulting in the secretion of diverse array of secondary metabolites with antimicrobial properties against various antibiotic resistant bacteria. Following extraction, fractioning and re-testing of active metabolites resulted in persistent antibacterial activity against Escherichia coli (Migula) (ATCC 13706) and subsequent Nuclear Magnetic Resonance (NMR) analysis of the active fractions confirmed the induction of metabolites different than the ones in fractions which did not display activity against the same bacterial species. Overall findings again confirmed the value of One Strain-Many Compounds (OSMAC) approach that tests a wide range of growth parameters to trigger bioactive compound secretion increasing the likelihood of finding novel therapeutic agents. The isolate was found to be adaptable to both marine and terrestrial conditions corresponding to its original near-shore marine intertidal environment. Wide variations in its morphology, sporulation and diffusible pigment production were observed when different growth media were used.

Stimulating the proliferation, migration and lamellipodia of Schwann cells using low-dose curcumin.

Transplantation of peripheral glia is being trialled for neural repair therapies, and identification of compounds that enhance the activity of glia is therefore of therapeutic interest. We have previously shown that curcumin potently stimulates the activity of olfactory glia. We have now examined the effect of curcumin on Schwann cell (SC) activities including proliferation, migration and the expression of protein markers. SCs were treated with control media and with different concentrations of curcumin (0.02-20 µM). Cell proliferation was determined by MTS assay and migration changes were determined by single live cell migration tracking. We found that small doses of curcumin (40 nM) dramatically increased the proliferation and migration in SCs within just one day. When compared with olfactory glia, curcumin stimulated SC proliferation more rapidly and at lower concentrations. Curcumin significantly increased the migration of SCs, and also increased the dynamic activity of lamellipodial waves which are essential for SC migration. Expression of the activated form of the MAP kinase p38 (p-p38) was significantly decreased in curcumin-treated SCs. These results show that curcumin's effects on SCs differ remarkably to its effects on olfactory glia, suggesting that subtypes of closely related glia can be differentially stimulated by curcumin. Overall these results demonstrate that the therapeutically beneficial activities of glia can be differentially enhanced by curcumin which could be used to improve outcomes of neural repair therapies.

Anti-staphylococcal activity of C-methyl flavanones from propolis of Australian stingless bees (Tetragonula carbonaria) and fruit resins of Corymbia torelliana (Myrtaceae).

Propolis of Australian stingless bees (Tetragonula carbonaria, Meliponini) originating from Corymbia torelliana (Myrtaceae) fruit resins was tested for its antimicrobial activities as well as its flavonoid contents. This study aimed at the isolation, structural elucidation and antibacterial testing of flavanones of C. torelliana fruit resins that are incorporated into stingless bee propolis. Flavanones of this study were elucidated by spectroscopic and spectrometric methods including UV, 1D and 2D NMR, EI-MS, ESI-MS and HR-MS. The results indicated known C-methylated flavanones namely, 1 (2S)-cryptostrobin, its regioisomer 2 (2S)- stroboponin, 3 (2S)- cryptostrobin 7-methyl ether, and 6 (2S)- desmethoxymatteucinol, and known flavanones 4 (2S)- pinostrobin and 5 (2S)- pinocembrin as markers for C. torelliana fruit resins and one propolis type. Ethanolic preparations of propolis were shown to be active against Staphylococcus aureus (ATCC 25923) and to a lesser extent against Pseudomonas aeruginosa (ATCC 27853). C. torelliana flavanones inhibited the growth of S. aureus therefore contributing to the antibacterial effects observed for Australian stingless bee propolis extracts.

Recessive congenital methaemoglobinaemia: functional characterization of the novel D239G mutation in the NADH-binding lobe of cytochrome b5 reductase.

Type I recessive congenital methaemoglobinaemia (RCM), caused by the reduced form of nicotinamide adenine dinucleotide (NADH)-cytochrome b(5) reductase (cytb(5)r) deficiency, manifests clinically as cyanosis without neurological dysfunction. Two mutations, E255- and G291D, have been identified in the NADH-binding lobe of cytb(5)r in previously reported patients, and we have detected a further novel mutation, D239G, in this lobe in two unrelated Irish families. Although one family belongs to the genetically isolated Traveller Community, which separated from the general Irish population during the 1845-48 famine, the D239G mutation was present on the same haplotype in both families. Three known cytb(5)r mutations were also identified, including the R159- mutation, which causes loss of the entire NADH-binding lobe and had previously been reported in an individual with type II RCM. Characterization of the three NADH-binding lobe mutants using a heterologous expression system revealed that all three variants retained stoichiometric levels of flavin adenine dinucleotide with spectroscopic and thermodynamic properties comparable with those of native cytb(5)r. In contrast to the E255- and G291D variants, the novel D239G mutation had no adverse impact on protein thermostability. The D239G mutation perturbed substrate binding, causing both decreased specificity for NADH and increased specificity for NADPH. Thus cytb(5)r deficient patients who are heterozygous for an NADH-binding lobe mutation can exhibit the clinically less severe type I phenotype, even in association with heterozygous deletion of the NADH-binding lobe.

Attitudes of general practitioners to their interactions with pharmaceutical companies: a qualitative study.

OBJECTIVES: To obtain the attitudes of a sample of General Practitioners to their interactions with pharmaceutical companies. DESIGN: Semi-structured face to face interviews. SETTING: General Practices in the north-west of Ireland. RESULTS: General Practitioners do not value their interactions with pharmaceutical representatives. They regard it as promotional, not educational activity and believe they are presented with biased information. Positive aspects (social, and receiving information) do not compensate. The content of educational meetings should be decided by G.Ps. alone. More directly promotional meetings are valued less, except when fairly lavish, in which case they are harder to resist. Material received through the post is not valued at all by G.Ps. CONCLUSIONS: Pharmaceutical companies in Ireland have a good relationship with G.Ps. It is in jeopardy. To rescue it, companies need to provide G.Ps. with assistance (information and other types) which is directly helpful to G.Ps. caring for their patients. Companies need to row back on the deluge of promotional material that G.Ps. are faced with. G.Ps. need to be trained to learn how to demand more helpful material from companies, and to refuse the promotional tidal wave.

Cheilanthane sesterterpenes, protein kinase inhibitors, from a marine sponge of the genus Ircinia.

Four cheilanthane sesterterpenoids, 25-hydroxy-13(24),15,17-cheilanthatrien-19,25-olide (1), 13,16-epoxy-25-hydroxy-17-cheilanthen-19,25-olide (2), 25-hydroxy-13(24),17-cheilanthadien-16,19-olide (3), and 16,25-dihydroxy-13(24),17-cheilanthadien-19,25-olide (4), were isolated from the marine sponge Irciniasp. Compounds 1, 3, and 4 are new natural products. The four compounds inhibit MSK1 (mitogen and stress activated kinase) and MAPKAPK-2 (mitogen activated protein kinase activated protein kinase), two protein kinases involved in mitogen and stress signal transduction.

High-pressure synthesis of enantiomerically pure C-6 substituted pyrazol.

The synthesis of enantiomerically pure C-6 substituted pyrazolo[3,4-d]pyrimidines has been performed by aromatic nucleophilic substitution of 4-amino-6-chloro-1-phenylpyrazolo[3,4-rd]pyrimidine under conditions of high pressure at ambient temperature. Conventional synthetic conditions (reflux at atmospheric pressure) were unsuccessful. The S enantiomer 11 displayed higher affinity and selectivity for the adenosine A1 receptor than the R enantiomer 12.

A study of the binding requirements of calyculin A and dephosphonocalyculin A with PP1, development of a molecular recognition model for the binding interactions of the okadaic acid class of compounds with PP1.

The interactions of the okadaic acid class of compounds, with special emphasis on the solution structures of calyculin A and dephosphonocalyculin A with PP1 are reported. After examination of the interactions of all docked structures, a receptor based pharmacophore model for the interactions of the protein phosphatase inhibitors has been developed. Calyculin A or dephosphonocalyculin A can interact with the enzyme in either a manner similar to the reported crystal structure, or in an extended form. The inhibitors require two essential regions interacting with the hydrophobic region and the central metal binding regions of the enzyme. This simplified model is consistent with previously published models of the okadaic acid class of compounds with PP1.

A benzylisoquinoline alkaloid from Doryphora sassafras.

Chemical investigation of the Australian rainforest plant Doryphora sassafras has resulted in the isolation of a new natural product, 2-methyl-1-(p-methoxybenzyl)-6,7-methylenedioxyisoquinolinium chloride (1). The iodide salt of compound 1 has previously been synthesized but only partially characterized. This paper reports the full spectroscopic characterization of 1 by MS, IR, UV, and NMR data.

Anhydride modified cantharidin analogues. Is ring opening important in the inhibition of protein phosphatase 2A?

A series of anhydride modified cantharidin analogues have been synthesised and screened for their ability to inhibit protein phosphatase 2A. Surprisingly only analogues capable of undergoing a facile ring opening of the anhydride moiety displayed any significant inhibition. Subsequent NMR experiments indicated that 7-oxobicyclo[2.2.1]heptane-2,3-dicarboxylic acid was the major (sole) species under assay conditions. The ability of these modified anhydro-cantharidin analogues to inhibit protein phosphatase 2A varies from 4 (16) to 100% (8) at 100 microM test concentration.

1-Phenylpyrazolo[3,4-d]pyrimidines; structure-ac:tivity relationships for C6 substituents at A1 and A2A adenosine receptors.

Substitution of I-phenylpyrazolo[3,4-d]pyrimidines at C6 with N-alkyl-2-thiopropionamide groups has resulted in a series of 18 compounds which have been evaluated for binding at A1 and A2A adenosine receptors. Introduction of an N-ethyl group gave increased affinity at both A1 and A2A receptors for the amino compound 7b compared to the primary amide 7a. An additional hydrophobic pocket exists for substituents on the amide. This pocket allows an N-ethyl group for increased affinity at both A1 and A2A receptors, allows larger alkyl groups at A2A receptors but not at A1 receptors and there is an H-bond interaction requiring one H-bond donor. Molecular modeling studies have also enabled a proposal of the amino acid residues involved in ligand binding at both the A1 and A2A receptors.

10-Hydroxydarlingine, a new tropane alkaloid from the Australian proteaceous plant triunia erythrocarpa.

Triunia erythrocarpa was identified as containing alkaloids during chemical screening of Queensland Proteaceae using Dragendorff's reagent. A new tropane, 10-hydroxydarlingine (1), and the known tropane, darlingine (2), were isolated from the leaves of T. erythrocarpa. The absolute stereochemistry of 10-hydroxydarlingine (1) was assigned using the advanced Mosher method. T. erythrocarpa is only the seventh member of the Proteaceae to have been shown to produce alkaloids.

Polyoxygenated dysidea sterols that inhibit the binding of [I125] IL-8 to the human recombinant IL-8 receptor type A.

The combined CH(2)Cl(2) and MeOH crude extract of a new species of the marine sponge Dysidea, collected in Northern Australia was found to inhibit the binding of [I125] interleukin-8 [IL-8] to the human recombinant IL-8 receptor type A at 500 microg/mL. Bioassay-guided fractionation led to the isolation of three new polyoxygenated sterols 3, 4, and 5. Their structures were assigned on the basis of 1D and 2D NMR experiments, and relative stereochemistries were established by ROESY correlations and analysis of coupling constants. The IC(50) values for inhibition of IL-8Ra for sterols 3, 4, and 5 were 20, 5.5, and 4.5 microM, respectively.

Anthoptilides A-E, new Briarane diterpenes from the Australian sea pen Anthoptilum cf. kukenthali.

The Australian sea pen Anthoptilum cf. kukenthali has afforded five new briarane-type diterpenes, anthoptilides A-E. Their structures were determined on the basis of their spectroscopic data. Single-crystal X-ray determination was performed on anthoptilide A. Anthoptilides B and C inhibited the binding of [(3)H]1, 3-dipropyl-8-cyclopentylxanthine ([(3)H]DPCPX) on adenosine A(1) receptors.

Isolation of psammaplin A 11'-sulfate and bisaprasin 11'-sulfate from the marine sponge Aplysinella rhax.

Psammaplin A 11'-sulfate (3) and bisaprasin 11'-sulfate (4) have been isolated from the marine sponge Aplysinella rhax, along with the known psammaplin A (1). Their structures were determined on the basis of their spectroscopic data. Compounds 1 and 3 inhibited [(3)H]1,3-dipropyl-8-cyclopentylxanthine binding to rat-brain adenosine A(1) receptors.

The solution structures of calyculin A and dephosphonocalyculin A by NMR.

The NMR solution structure of calyculin A (1) in chloroform exhibits intramolecular interactions, resembling the original crystal structure. In methanol, calyculin A has the hydrogen bonding moieties solvent exposed. Dephosphonocalyculin A in chloroform resembles calyculin A in chloroform and the crystal structure of calyculin A. Dephosphonocalyculin A in methanol resembles calyculin A in methanol.