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Dive into the research topics where Adam McCluskey is active.

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Featured researches published by Adam McCluskey.


Journal of Medicinal Chemistry | 2009

Inhibition of dynamin mediated endocytosis by the dynoles: synthesis and functional activity of a family of indoles

Timothy A. Hill; Christopher P. Gordon; Andrew B. McGeachie; Barbara Venn-Brown; Luke R. Odell; Ngoc Chau; Annie Quan; Anna Mariana; Jennette A. Sakoff; Megan Chircop; Phillip J. Robinson; Adam McCluskey

Screening identified two bisindolylmaleimides as 100 microM inhibitors of the GTPase activity of dynamin I. Focused library approaches allowed development of indole-based dynamin inhibitors called dynoles. 100-Fold in vitro enhancement of potency was noted with the best inhibitor, 2-cyano-3-(1-(2-(dimethylamino)ethyl)-1H-indol-3-yl)-N-octylacrylamide (dynole 34-2), a 1.3 +/- 0.3 microM dynamin I inhibitor. Dynole 34-2 potently inhibited receptor mediated endocytosis (RME) internalization of Texas red-transferrin. The rank order of potency for a variety of dynole analogues on RME in U2OS cells matched their rank order for dynamin inhibition, suggesting that the mechanism of inhibition is via dynamin. Dynoles are the most active dynamin I inhibitors reported for in vitro or RME evaluations. Dynole 34-2 is 15-fold more active than dynasore against dynamin I and 6-fold more active against dynamin mediated RME (IC(50) approximately 15 microM; RME IC(50) approximately 80 microM). The dynoles represent a new series of tools to better probe endocytosis and dynamin-mediated trafficking events in a variety of cells.


Journal of Biological Chemistry | 2011

Dynamin Inhibition Blocks Botulinum Neurotoxin Type A Endocytosis in Neurons and Delays Botulism

Callista B. Harper; Sally Martin; Tam Nguyen; Shari J. Daniels; Nickolas A. Lavidis; Michel R. Popoff; Gordana Hadzic; Anna Mariana; Ngoc Chau; Adam McCluskey; Phillip J. Robinson; Frederic A. Meunier

The botulinum neurotoxins (BoNTs) are di-chain bacterial proteins responsible for the paralytic disease botulism. Following binding to the plasma membrane of cholinergic motor nerve terminals, BoNTs are internalized into an endocytic compartment. Although several endocytic pathways have been characterized in neurons, the molecular mechanism underpinning the uptake of BoNTs at the presynaptic nerve terminal is still unclear. Here, a recombinant BoNT/A heavy chain binding domain (Hc) was used to unravel the internalization pathway by fluorescence and electron microscopy. BoNT/A-Hc initially enters cultured hippocampal neurons in an activity-dependent manner into synaptic vesicles and clathrin-coated vesicles before also entering endosomal structures and multivesicular bodies. We found that inhibiting dynamin with the novel potent Dynasore analog, Dyngo-4aTM, was sufficient to abolish BoNT/A-Hc internalization and BoNT/A-induced SNAP25 cleavage in hippocampal neurons. Dyngo-4a also interfered with BoNT/A-Hc internalization into motor nerve terminals. Furthermore, Dyngo-4a afforded protection against BoNT/A-induced paralysis at the rat hemidiaphragm. A significant delay of >30% in the onset of botulism was observed in mice injected with Dyngo-4a. Dynamin inhibition therefore provides a therapeutic avenue for the treatment of botulism and other diseases caused by pathogens sharing dynamin-dependent uptake mechanisms.


Green Chemistry | 2009

Activity and thermal stability of lysozyme in alkylammonium formate ionic liquids—influence of cation modification

Jason P. Mann; Adam McCluskey; Rob Atkin

The stability and activity of hens egg white lysozyme in the presence of four protic room temperature ionic liquids (ethylammonium formate (EAF), propylammonium formate (PAF), 2-methoxyethylammonium formate (MOEAF) and ethanolammonium formate (EtAF)) have been investigated. Near UV CD experiments have been used to determine protein structure in aqueous solutions of 25 wt%, 50 wt% and 75 wt% ionic liquid, and to assess the proteins ability to refold after heating to 90 °C. It was determined that EAF and MOEAF are similarly effective refolding additives, while PAF is more effective at promoting refolding at concentrations up to ∼62.5 wt%, but at higher PAF concentrations the protein spontaneously denatures. Both of these effects are attributed to the increased hydrophobicity of the cation. EtAF is shown to stabilise lysozyme against unfolding at high temperature, and renaturing appears to be near complete upon cooling. Studies of enzyme kinetics reveal increased protein activity in the presence of all ionic liquids examined, but the most significant increase is noted for EtAF, where rates are six times higher than those determined for lysozyme in buffered water.


Traffic | 2013

Building a Better Dynasore: The Dyngo Compounds Potently Inhibit Dynamin and Endocytosis†

Adam McCluskey; James A. Daniel; Gordana Hadzic; Ngoc Chau; Emma L. Clayton; Anna Mariana; Ainslie Whiting; Nick N. Gorgani; Jonathan Lloyd; Annie Quan; Lia Moshkanbaryans; Sai Krishnan; Swetha Perera; Megan Chircop; Lisa von Kleist; Andrew B. McGeachie; Mark T. Howes; Robert G. Parton; Michael Campbell; Jennette A. Sakoff; Xuefeng Wang; JianYuan Sun; Mark J. Robertson; Fiona M. Deane; Tam Nguyen; Frederic A. Meunier; Michael A. Cousin; Phillip J. Robinson

Dynamin GTPase activity increases when it oligomerizes either into helices in the presence of lipid templates or into rings in the presence of SH3 domain proteins. Dynasore is a dynamin inhibitor of moderate potency (IC50 ˜ 15 μM in vitro). We show that dynasore binds stoichiometrically to detergents used for in vitro drug screening, drastically reducing its potency (IC50 = 479 μM) and research tool utility. We synthesized a focused set of dihydroxyl and trihydroxyl dynasore analogs called the Dyngo™ compounds, five of which had improved potency, reduced detergent binding and reduced cytotoxicity, conferred by changes in the position and/or number of hydroxyl substituents. The Dyngo compound 4a was the most potent compound, exhibiting a 37‐fold improvement in potency over dynasore for liposome‐stimulated helical dynamin activity. In contrast, while dynasore about equally inhibited dynamin assembled in its helical or ring states, 4a and 6a exhibited >36‐fold reduced activity against rings, suggesting that they can discriminate between helical or ring oligomerization states. 4a and 6a inhibited dynamin‐dependent endocytosis of transferrin in multiple cell types (IC50 of 5.7 and 5.8 μM, respectively), at least sixfold more potently than dynasore, but had no effect on dynamin‐independent endocytosis of cholera toxin. 4a also reduced synaptic vesicle endocytosis and activity‐dependent bulk endocytosis in cultured neurons and synaptosomes. Overall, 4a and 6a are improved and versatile helical dynamin and endocytosis inhibitors in terms of potency, non‐specific binding and cytotoxicity. The data further suggest that the ring oligomerization state of dynamin is not required for clathrin‐mediated endocytosis.


Molecular Pharmacology | 2007

Myristyl Trimethyl Ammonium Bromide and Octadecyl Trimethyl Ammonium Bromide Are Surface-Active Small Molecule Dynamin Inhibitors that Block Endocytosis Mediated by Dynamin I or Dynamin II

Annie Quan; Andrew B. McGeachie; Damien J. Keating; Ellen M. van Dam; Jenny Rusak; Ngoc Chau; Chandra S. Malladi; Chen Chen; Adam McCluskey; Michael A. Cousin; Phillip J. Robinson

Dynamin is a GTPase enzyme involved in membrane constriction and fission during endocytosis. Phospholipid binding via its pleckstrin homology domain maximally stimulates dynamin activity. We developed a series of surface-active small-molecule inhibitors, such as myristyl trimethyl ammonium bromide (MiTMAB) and octadecyltrimethyl ammonium bromide (OcTMAB), and we now show MiTMAB targets the dynamin-phospholipid interaction. MiTMAB inhibited dynamin GTPase activity, with a Ki of 940 ± 25 nM. It potently inhibited receptor-mediated endocytosis (RME) of transferrin or epidermal growth factor (EGF) in a range of cells without blocking EGF binding, receptor number, or autophosphorylation. RME inhibition was rapidly reversed after washout. The rank order of potency for a variety of MiTMAB analogs on RME matched the rank order for dynamin inhibition, suggesting dynamin recruitment to the membrane is a primary cellular target. MiTMAB also inhibited synaptic vesicle endocytosis in rat brain nerve terminals (synaptosomes) without inducing depolarization or morphological defects. Therefore, the drug rapidly and reversibly blocks multiple forms of endocytosis with no acute cellular damage. The unique mechanism of action of MiTMAB provides an important tool to better understand dynamin-mediated membrane trafficking events in a variety of cells.


Investigational New Drugs | 2002

Anticancer Activity and Protein Phosphatase 1 and 2A Inhibition of a New Generation of Cantharidin Analogues

Jennette A. Sakoff; Stephen P. Ackland; Monique L. Baldwin; Mirella A. Keane; Adam McCluskey

Cantharidin (Spanish Fly) is a naturaltoxin and an inhibitor of proteinphosphatases 1 (PP1) and 2A (PP2A), whichhave key roles in cell cycle progression.We have synthesised two series ofdemethylated cantharidin analogues, onedisplaying an open-ring lactoneconfiguration in solution (Novo-1 toNovo-5) similar to cantharidin, the othershowing a closed-ring lactone configuration(Novo-6 to Novo-10). In the present study,these ten agents were screened for invitro PP1 and PP2A inhibition and cellularcytotoxicity in nine cancer cell lines ofhaematopoietic (L1210, HL60), ovarian(A2780, ADDP), osteo (143B), and colon(HCT116, HT29, WiDr, SW480) origin and onenormal colon cell line (CCD-018).The open-ring series (IC50,PP1=2.0−4.8 μM, PP2A=0.2−0.5 μM)maintained the PP2A selectivity ofcantharidin (IC50, PP1=1.8 μM,PP2A=0.2 μM), although some were lesspotent. The closed-ring series (IC50,PP1 = 12.5->1000 μM,PP2A=5->1000 μM) were considerablyless potent inhibitors, confirming the needof ring opening for inhibition. Thecytotoxicity (IC50, 72 h, MTT assay) ofcantharidin ranged from 6−15 μM, whilethe new analogues ranged from 14 to>1000 μM. Cytotoxicity of the agentsdid not consistently parallel the invitro potency of protein phosphataseinhibition. A number of analogues showedcolon cancer selectivity, particularlyNovo-6, where the cytotoxicity ranged from14−88 μM in the colon cancer cells and275−680 μM in all other cell linesincluding normal colon cells. The reasonfor this selectivity was not apparent andmay involve additional intracellulartargets. Cell cycle analysis showedcantharidin to enhance cell cycleprogression as evident from an increasedS-phase population and enhanced DNAsynthesis, culminating in G2/M arrestand apoptosis. With Novo-1 and Novo-6, thecell cycle changes paralleled thecytotoxicity responses, with thepredominant effect of G2/M cell cyclearrest followed by cell death. Inconclusion, we have synthesised newanticancer agents that show selectivecytotoxicity in colon cancer cells whileremaining inactive in normal colon cells,and which mediate their effects viathe G2/M phase of the cell cycle.


Bioorganic & Medicinal Chemistry Letters | 2000

Anhydride modified cantharidin analogues: synthesis, inhibition of protein phosphatases 1 and 2A and anticancer activity

Adam McCluskey; Michael C. Bowyer; Elizabeth Collins; Alistair T. R. Sim; Jennette A. Sakoff; Monique L. Baldwin

Two series of anhydride modified cantharidin analogues were synthesised and screened for their phosphatase inhibition (PP1 and PP2A) and cytotoxicity in various cancer cell lines (Ovarian A2780, ADDP; Osteosarcoma 143B; and Colon HCT116 and HT29). One series was synthesised by a novel, high yielding one-pot hydrogenation-ring-opening-esterification procedure, the other by acid catalysed acetal formation. Analogues 5-7 and 9 displayed moderate PP2A selectivity (ca. 5- to 20-fold) and inhibition typically in the low microM range (comparable, in some cases to cantharidin). The anticancer activity of these analogues varied with the cell line under study; however, many of them showed selective cytotoxicity for the colon tumour cell lines.


Bioorganic Chemistry | 2003

Cantharidin analogues: synthesis and evaluation of growth inhibition in a panel of selected tumour cell lines

Adam McCluskey; Stephen P. Ackland; Michael C. Bowyer; Monique L. Baldwin; James Garner; Cecilia Walkom; Jennette A. Sakoff

Diels-Alder addition of furans (furan, furfuryl alcohol, and 3-bromofuran) to maelic anhydride yields three distinct 5,6-dehydronorcantharidins. Hydrogenation of (4,10-dioxatricyclo[5.2.1.0]decane-3,5-dione) (4a), in dry ethanol affords the monoester (7-oxabicyclo[2.2.1]heptane-2,3-dicarboxylic aid monoethyl ester) (6). Subsequent transesterification affords a series of monoesters (7-oxabicyclo[2.2.1]heptane-2,3-dicarboxylic acid monomethyl ester (7)), 7-oxabicyclo[2.2.1]heptane-2,3-dicarboxylic acid monopropyl ester (8), (7-oxabicyclo[2.2.1]heptane-2,3-dicarboxylic acid monohexyl ester (9)) and differentially substituted diesters (7-oxabicyclo[2.2.1]heptane-2,3-dicarboxylic acid 2-ethyl ester 3-isopropyl ester) (10), and (7-oxabicyclo[2.2.1]heptane-2,3-dicarboxylic acid 2-ethyl ester 3-phenyl ester) (11). Analogues were firstly screened for their ability to inhibit protein phosphatases 1 (PP1) and 2A (PP2A) as the lead compounds cantharidin (1) and norcantharidin (2) are known PP1 and PP2A inhibitors. Only analogues 4a, 6-8 displayed good PP1 and PP2A inhibition (PP1 IC(50)s=2.0, 2.96, 4.71, and 4.82 microM, respectively; PP2A IC(50)s=0.2, 0.45, 0.41, and 0.47 microM, respectively). All analogues were also screened for their anti-cancer potential against a panel of tumour cell lines, HL60, L1210, SW480, WiDr, HT29, HCT116, A2780, ADDP, and 143B, producing GI(50) values ranging from 6 microM to >1000 microM. Analogues possessing good PP1 and/or PP2A inhibition also returned moderate to good anti-cancer activity. Analogues with substituents directly attached to the intact bicyclo[2.2.1]heptane skeleton were poor to moderate anti-cancer agents. This correlates well with their lack of PP1 or PP2A activity. Analogues capable of undergoing a facile ring opening of the anhydride or with a single carboxylate were good PP1 and PP2A inhibitors, largely correlating to the observed anti-cancer activity in all cases, except 11. Analogue 11, whist neither a PP1 nor a PP2A inhibitor shows anti-cancer activity comparable to 1 and 2. We believe that intracellular esterases generate the corresponding dicarboxylate, which is a potent PP1 and PP2A inhibitor, and that it is this species which is responsible for the observed anti-cancer activity.


Green Chemistry | 2004

‘Green’ leaching: recyclable and selective leaching of gold-bearing ore in an ionic liquid

Jacqueline A. Whitehead; Geoffrey A. Lawrance; Adam McCluskey

The recovery of gold and silver from ore in an ionic liquid is reported for the first time. The 1-butyl-3-methyl-imidazolium hydrogen sulfate ionic liquid (bmim+HSO4−) was employed, with iron(III) sulfate oxidant and thiourea added. Selective extraction of gold (≥85%) and silver (≥60%) from powdered ore (of dominantly chalcopyrite/pyrite/pyrrhotite/sphalerite mineralogy) was achieved at room temperature in 50 h, with other lower-value metals present in the ore (Cu, Zn, Pb, Fe) extracted to only low percentages. Gold extraction was similar to that achieved in aqueous H2SO4/thiourea/Fe2(SO4)3, and silver extraction was significantly better. Moreover, the ionic liquid can be recycled following selective stripping of gold and silver on activated charcoal, with reuse in at least four successive treatments leading to neither ionic liquid degradation nor any loss in extraction efficiency.


Organic and Biomolecular Chemistry | 2007

Molecularly imprinted polymers (MIPs): sensing, an explosive new opportunity?

Adam McCluskey; Clovia I. Holdsworth; Michael C. Bowyer

Our group is currently developing in-field detection systems alongside the Australian Federal Police Forensic Services utilising molecularly imprinted polymers as the recognition elements. This review looks at MIP synthesis and our perceptions of future directions from an Australian and forensic perspective.

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Jennette A. Sakoff

Mater Misericordiae Hospital

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Phillip J. Robinson

Children's Medical Research Institute

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Ngoc Chau

Children's Medical Research Institute

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