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Analgesic Component of Venom (ACV1) from Cone Snails :
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31 December, 2005
Comment on use of Ziconotide for palliative care
Narayana, A.K. (2005) Elan: ziconotide review focused on off-label uses. Am J Hosp Palliat Care. 22 (6): 408.
[No abstract available. Comment on Prommer, E.E. (2005) Ziconotide: can we use it in palliative care? Am J Hosp Palliat Care. 22(5):369-374. ]
25 December, 2005. HAPPY CHRISTMAS ALL !!
VISAYA Vol. 5 Published. New Conus species described.
VISAYA, 5 [2005], 95pp., 54col.pls. Euro 50.00-55.00 [indication]
Abstract: The 5th issue of Visaya contains 16 articles, which were written by M.Chiapponi, F. Déniz, W. Engl, K. Fraussen, J.M. Hernandez, F. Lorenz, D. Massiglia, S. Patamakanthin, G.T. Poppe, C. Redfern, E. Rolán, R. Salisbury, D. Stratmann, F. Swinnen, S. Tagaro, Y. Terryn, R. Vanwalleghem, Y. Verhaeghe, C. Vos, J. Wolff and G. Raybaudi Massilia. The following new Conus species was described:
Conus guidopoppe
Some references to past papers on Conus available from :A.N. van der Bijl, Burgemeester van Bruggenstraat 41, 1165 NV Halfweg, Nederland +31-20-4977772 (phone); anvdbijl@xs4all.nl (e-mail)
23 December, 2005
Conantokin-G from Conus geographus: Interaction with receptors
L, Yasuda T, Lewis RJ, Dodd PR, Adams DJ. (2005) )NMDA receptor subunit-dependent modulation by conantokin-G and Ala(7)-conantokin-G. Ragnarsson. J Neurochem. 2005 Nov 29; [Epub ahead of print]
School of Molecular and Microbial Sciences, University of Queensland, Brisbane, Queensland, Australia.
Abstract: The modulation of recombinant NMDA receptors by conantokin-G (con-G) and Ala(7)-conantokin-G (Ala(7)-Con-G) was investigated in Xenopus oocytes injected with capped RNA coding for NR1 splice variants and NR2 subunits using the two-electrode voltage clamp technique. Glutamate exhibited a marginally higher apparent affinity for NR2A-containing receptors than NR2B-containing receptors, regardless of the NR1 subunit present. Conantokins were bath applied to give cumulative concentration responses in the presence of 3 and 30 mum glutamate. Both contantokins exhibited biphasic concentration-response relationships at NR2A-containing NMDA receptors, producing potentiation at low conantokin concentrations and inhibition at high concentrations. These effects were stronger with glutamate concentrations near its EC(50), and less marked at saturating concentrations. In contrast, the conantokin concentration-response relation was monophasic and inhibitory at NR2B-containing receptors. We conclude that the combinations of subunits that comprise the NMDA receptor complex influence conantokin and glutamate affinities and the nature of the responses to conantokins.
2 December, 2005
Use of Conus and other peptide toxins in the analysis of nicotinic receptors
Fruchart-Gaillard C, Menez A, Servent D. (2005) [Critical role of peptidic toxins in the functional and structural analysis of nicotinic acetylcholine receptors] . J Soc Biol. 199: 45-53. Review. French.
Departement d'lngenierie et d'Etudes des Proteines, CE Saclay, 91191 Gif, Yvette.
Abstract: Animal toxins which interact on various receptors and channels have been often used in the studies of the functional roles of these targets. Nicotinic toxins have been purified from snake and cone venoms and are characterized by high affinity and various selectivity of interactions on the different nicotinic receptors subtypes. Since 30 years they have been used as molecular probes to identify, localize and purify these receptors. Furthermore, they have played a crucial role in the better understanding of their functional properties and have been useful in their structural studies. These peptidic toxins could be chemically synthesized or recombinantly expressed and nonnatural residues could be introduced in their sequences in order to delineate their functional interaction sites. The structural modelisation of toxin-nAChR interaction allows us to understand the antagonistic property of these toxins and open the way to the design of engineered ligands with predetermined specificity, useful as pharmacological tools or therapeutic agents in the numerous diseases involving this receptor family.
Conantokin-G from Conus geographus lessens cerebral ischemia
Division of Psychiatry and Neurosciences, Walter Reed Army Institute of Research, 503 Robert Grant Ave., Silver Spring, MD 20910, USA.
Abstract: The potent NMDA receptor antagonist, Conantokin-G (CGX-1007), a snail peptide, has an 8-h therapeutic window in rat focal cerebral ischemia. We hypothesized that the mechanism of neuroprotection is the inhibition of 'secondary phase' peri-infarct depolarizations (PIDs), recently shown to recur 6--24 h post-reperfusion. Rats were implanted with intrathecal (i.t.) catheters for drug delivery and DC-compatible electrodes for continuous PID monitoring and subjected to transient (2 h) middle cerebral artery occlusion. Four groups were studied. In two groups (C(40)C and C(20)C), continuous infusion of CGX-1007 was administered over 8--24 h post-occlusion at 0.1 microg/h (0.04 nmol/h) following either a 40- or 20-nmol bolus dose at 8 h. Another group (C(40)S) received the 40-nmol bolus followed by saline infusion, and a control group received saline. Intrathecal drug treatment reduced infarct volumes relative to controls by 61%, 31%, and 10% in C(40)C, C(40)S, and C(20)C groups, respectively, but also induced dose-dependent paralysis and elevated mortality. All rats had PID rates similar to the control group prior to treatment, but following treatment secondary phase PIDs were reduced by 47--57% in each drug group compared to controls. Because several animals exhibited PID inhibition but no neuroprotection, there was no significant correlation between these endpoints across groups. However, drug-treated animals that did not exhibit secondary phase PIDs prior to treatment had significantly smaller infarcts and reduced subsequent PID activity than corresponding control rats. Results suggest that post-reperfusion PIDs play a substantial, though still undefined pathogenic role in delayed maturation of cerebral infarction and NMDA receptor-targeted neuroprotection.
Conantokin G, an antagonist of NMDA (NR1/2A/2B and NR1/2B) receptors, inhibits p38 phosphorylation.
Waxman, E.A. and Lynch, D.R. (2005). N-methyl-D-aspartate receptor subtype mediated bidirectional control of p38 mitogen-activated protein kinase. J Biol Chem. 280: 29322-29333..
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA.
Abstract: N-methyl-d-aspartate receptor (NMDAR) stimulation activates many downstream mechanisms involved in both cell survival and cell death. The manner in which the NMDAR regulates one of these pathways, the p38 mitogen-activated protein kinase (p38) pathway, is currently unknown. In the present study, we have defined a developmental-, concentration-, and time-dependent phosphorylation and subsequent dephosphorylation of p38. In cultured hippocampal neurons 7-8 days in vitro (DIV7-8), NMDAR stimulation leads to a concentration-dependent increase in p38 phosphorylation (phospho-p38). However, in more mature neurons (>DIV17) application of NMDA produces concentration-dependent effects, such that low concentrations result in sustained increases in phospho-p38 levels, and high concentrations dephosphorylate p38 within 5 min. Conantokin G, an antagonist of NR1/2A/2B and NR1/2B receptors, inhibits p38 phosphorylation, while NR1/2B-specific antagonists prevent the rapid dephosphorylation of p38 without affecting p38 activation. Furthermore, inhibition of calcineurin prevents the activation of p38, whereas inhibition of phosphoinositide 3-kinase (PI3K) prevents the rapid dephosphorylation of p38. Our results support the presence of subtype-dependent pathways regulating p38 activation and deactivation: one involves NR1/2A/2B receptors activating calcineurin and resulting in p38 phosphorylation, and the other utilizes NR1/2B receptors binding to and activating PI3K and leading to the dephosphorylation of p38 in a manner involving both NR1/2A/2B receptor activation and tyrosine phosphorylation of NR2B. The ability of NMDAR subtype-specific mechanisms to regulate p38 has implications for NMDAR-mediated synaptic plasticity, gene regulation, and excitotoxicity.
Acrorhagin II, a novel peptide toxin in the sea anemone, Actinia equina, resembles a cone snail neurotoxin
Honma, T., Minagawa, S., Nagai, H., Ishida, M., Nagashima, Y. and Shiomi K. (2005). Novel peptide toxins from acrorhagi, aggressive organs of the sea anemone Actinia equina. Toxicon. 46: 768-774..
Department of Food Science and Technology, Tokyo University of Marine Science and Technology, Konan-4, Minato-ku, Tokyo 108-8477, Japan.
Abstract: Two peptide toxins, acrorhagin I (50 residues) and II (44 residues), were isolated from special aggressive organs (acrorhagi) of the sea anemone Actinia equina by gel filtration on Sephadex G-50 and reverse-phase HPLC on TSKgel ODS-120T. The LD50 against crabs of acrorhagin I and II were estimated to be 520 and 80 microg/kg, respectively. 3'- and 5'-RACE established the amino acid sequences of the acrorhagin precursors. The precursor of acrorhagin I is composed of both signal and mature peptides and that of acrorhagin II has an additional sequence (propart) between signal and mature peptides. Acrorhagin I has no sequence homologies with any toxins, while acrorhagin II is somewhat similar to spider neurotoxins (hainantoxin-I from Selenocosmia hainana and Tx 3-2 from Phoneutria nigriventer) and cone snail neurotoxin (omega-conotoxin MVIIB from Conus magus). In addition, analogous peptides (acrorhagin Ia and IIa) were also cloned during RT-PCR experiments performed to confirm the nucleotide sequences of acrorhagins. This is the first to demonstrate the existence of novel peptide toxins in the sea anemone acrorhagi.
Marine neurotoxins: recognition and treatments. Review
Watters, M.R. and Stommel, E.W. (2005). Marine Neurotoxins: Envenomations and Contact Toxins. Curr Treat Options Neurol. 6: 115-123 .
Department of Medicine, Division of Neurology, University of Hawaii, 1356 Lusitana Street, 7th Floor, Honolulu, HI 96813, USA. mwatters@hawaii.edu
Abstract: Familiarity with the appearance and habitat of venomous sea creatures, the location of their stinging apparatus, and surveillance of population concentrations within recreational waters are essential in avoiding envenomations. Compared with the thermo-stable low molecular weighted ingestible seafood toxins, venomous toxins are often large molecular weight proteins and many are heat labile, which provides opportunity for therapeutic intervention. Heat therapy may denature the toxins, and provide immediate relief of pain in coelenterate and venomous fish envenomations. Injections of local anesthetic agents may also be used. First aid measures at the seashore may limit the spread of venom, and include immobilization of the affected sites, compression bandaging, and venous-lymphatic occlusive bandages. Measures to limit continued envenomation by attached stinging cells include topical vinegar for jellyfish tentacles and irrigation with debridment for spines of venomous fish. Antivenins are of limited availability and may be used for envenomations with sea snakes, Chironex box jellyfish, and some venomous fish. Sea snakes bites may be treated with antivenin from land snakes or with hemodialysis when antivenin is not available. Neuromuscular paralysis occurs with bites by sea snakes, cone snails, blue octopuses, and some jellyfish. Supportive treatment includes attention to cardiopulmonary resuscitation and intubation. Exposure to Pfeisteria may result in cognitive and behavioral abnormalities. Treatment with cholestyramine may be helpful in binding the toxin and improve recovery.
1 December, 2005
View these Real Top Seashells
Philippe Quiquandon and Olivier Santini of Real Top Seashells have some tempting examples of Conus on display at their 'Shells Passion.com' site.
On this web site you can see reference shells classified in TOP & World Record Specimens (WRS) sections. With respect to Conus species, there are already 61 WRS and 523 TOP specimens including 7 New Additions of which 3 are WRS (C. bandanus nigrescens; C. betulinus rifoluteus; C. cordigera) and 4 are TOP (C. fischolderi; C. nocturnus deburgliae; C. suratensis and C. terryni). Read more about this site here.
A regular update (daily or weekly) will keep you informed of additions to both sections. In all, over 4000 shell images in the image bank (currently 3459 TOP pictures). Subscriptions available for 6 months and 12 months access. (Click here for Details of membership).
30 November, 2005
REVIEW : Posttranslational modifications of conotoxins
Buczek, O., Bulaj, G. and Olivera, B.M. (2005) Conotoxins and the posttranslational modification of secreted gene products. Cellular and Molecular Life Sciences (CMLS) [REVIEW] Published online: 28 November 2005.
Department of Biology, University of Utah, Salt Lake City, Utah 84112, USA
Abstract: The venoms of predatory cone snails (genus Conus) have yielded a complex library of about 50-100,000 bioactive peptides, each believed to have a specific physiological target (although peptides from different species may overlap in their target specificity). Conus has evolved the equivalent of a drug development strategy that combines the accelerated evolution of toxin sequences with an unprecedented degree of posttranslational modification. Some Conus venom peptide families are the most highly posttranslationally modified classes of gene products known. We review the variety and complexity of posttranslational modifications documented in Conus peptides so far, and explore the potential of Conus venom peptides as a model system for a more general understanding of which secreted gene products may have modified amino acids. Although the database of modified conotoxins is growing rapidly, there are far more questions raised than answers provided about possible mechanisms and functions of posttranslational modifications in Conus.
29 November, 2005
Conus spurius reveals a hydrophobic T-1-conotoxin
Abstract: A major, very hydrophobic peptide, sr5a, was purified from the venom duct of Conus spurius specimens collected in the Yucatan Channel, Mexico. Its amino acid sequence (IINWCCLIFYQCC; calculated monoisotopic mass assuming two disulfide bridges 1616.68Da) was determined by automatic Edman degradation after reduction and alkylation, and confirmed by mass spectrometry (ESI monoisotopic mass, 1616.60; MALDI monoisotopic mass 1616.42Da). The primary structure of sr5a showed the pattern that characterizes the family of the T-1-conotoxins, which belong to the T-superfamily of conotoxins. The disulfide bonds were determined by partial reduction and alkylation with N-ethylmaleimide, followed by total reduction and alkylation with 4-vinylpyridine, and automatic Edman sequencing. The connectivity of the Cys residues (I-III, II-IV) is the same as that found in the T-1-conotoxin family. When injected intracranially (2.0nmol) into mice, peptide sr5a caused depressed behavioral activity.
See also:
Aguilar, M.B., Lopez-Vera, E., Imperial, J.S., Falcon, A., Olivera, B.M. and de la Cotera,
E.P. (2005) Putative gamma-conotoxins in vermivorous cone snails: the case of Conus
delessertii. Peptides. 26: 23-27.
20 November, 2005
Conus magus and other marine creatures : a source of Nature's Drugs
Synopsis: Scientists tend to prefer the lab to the mess and complication of living beings. Now they realize that forests and oceans hold a bounty of useful chemicals. Swimmers in the coral reefs of the Philippines know to stay away from Conus magus. The sea snail may be small—just a few inches long—but it's deadly mean. One dose of its venom can paralyze the passing fish that make up its diet. To drugmakers, though, the potency of its toxin is sheer poetry. Scientists who recently broke down the poison discovered—and copied exactly—a chemical compound that blocked nerve cells from sending signals to the brain. Result: Prialt, a new painkilling drug 1,000 times more powerful than morphine, the most potent analgesic now available to medicine.
Evolution of a Scientist
He had planned to enter the ministry, but his discoveries on a fateful voyage 170 years ago shook his faith and changed our conception of the origins of life. See article about Charles Darwin by Jerry Adler in Newsweek Technology and Science
Chi, S.W., Lee, S.H., Kim, D.H., Kim, J.S., Olivera, B.M., McIntosh, J.M. and Han, K.H. (2005) Solution structure of alpha-conotoxin PIA, a novel antagonist of alpha6 subunit containing nicotinic acetylcholine receptors. Biochem Biophys Res Commun. 338:1990-1997.
Protein Analysis and Design Laboratory, Division of Drug Discovery, Korea Research Institute of Bioscience and Biotechnology, Yusong P. O. Box 115, Daejon, Republic of Korea.
Abstract: alpha-Conotoxin PIA is a novel nicotinic acetylcholine receptor (nAChR) antagonist isolated from Conus purpurascens that targets nAChR subtypes containing alpha6 and alpha3 subunits. alpha-conotoxin PIA displays 75-fold higher affinity for rat alpha6/alpha3beta2beta3 nAChRs than for rat alpha3beta2 nAChRs. We have determined the three-dimensional structure of alpha-conotoxin PIA by nuclear magnetic resonance spectroscopy. The alpha-conotoxin PIA has an "omega-shaped" overall topology as other alpha4/7 subfamily conotoxins. Yet, unlike other neuronally targeted alpha4/7-conotoxins, its N-terminal tail Arg(1)-Asp(2)-Pro(3) protrudes out of its main molecular body because Asp(2)-Pro(3)-Cys(4)-Cys(5) forms a stable type I beta-turn. In addition, a kink introduced by Pro(15) in the second loop of this toxin provides a distinct steric and electrostatic environment from those in alpha-conotoxins MII and GIC. By comparing the structure of alpha-conotoxin PIA with other functionally related alpha-conotoxins we suggest structural features in alpha-conotoxin PIA that may be associated with its unique receptor recognition profile.
Mordvintsev, D.Y., Polyak, Y.L., Levtsova, O.V., Tourleigh, Y.V., Kasheverov, I.E., Shaitan, K.V., Utkin, Y.N., Tsetlin, V.I. (2005) A model for short alpha-neurotoxin bound to nicotinic acetylcholine receptor from Torpedo californica: Comparison with long-chain alpha-neurotoxins and alpha-conotoxins. Comput Biol Chem. 29: 398-411.
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, 117997, Miklukho-Maklaya str., 16/10, GSP-7, Moscow, Russia.
Abstract: Short-chain alpha-neurotoxins from snakes are highly selective antagonists of the muscle-type nicotinic acetylcholine receptors (nAChR). Although their spatial structures are known and abundant information on topology of binding to nAChR is obtained by labeling and mutagenesis studies, the accurate structure of the complex is not yet known. Here, we present a model for a short alpha-neurotoxin, neurotoxin II from Naja oxiana (NTII), bound to Torpedo californica nAChR. It was built by comparative modeling, docking and molecular dynamics using (1)H NMR structure of NTII, cross-linking and mutagenesis data, cryoelectron microscopy structure of Torpedo marmorata nAChR [Unwin, N., 2005. Refined structure of the nicotinic acetylcholine receptor at 4A resolution. J. Mol. Biol. 346, 967-989] and X-ray structures of acetylcholine-binding protein (AChBP) with agonists [Celie, P.H., van Rossum-Fikkert, S.E., van Dijk, W.J., Brejc, K., Smit, A.B., Sixma, T.K., 2004. Nicotine and carbamylcholine binding to nicotinic acetylcholine receptors as studied in AChBP crystal structures. Neuron 41 (6), 907-914] and antagonists: alpha-cobratoxin, a long-chain alpha-neurotoxin [Bourne, Y., Talley, T.T., Hansen, S.B., Taylor, P., Marchot, P., 2005. Crystal structure of Cbtx-AChBP complex reveals essential interactions between snake alpha-neurotoxins and nicotinic receptors. EMBO J. 24 (8), 1512-1522] and alpha-conotoxin [Celie, P.H., Kasheverov, I.E., Mordvintsev, D.Y., Hogg, R.C., van Nierop, P., van Elk, R., van Rossum-Fikkert, S.E., Zhmak, M.N., Bertrand, D., Tsetlin, V., Sixma, T.K., Smit, A.B., 2005. Crystal structure of nicotinic acetylcholine receptor homolog AChBP in complex with an alpha-conotoxin PnIA variant. Nat. Struct. Mol. Biol. 12 (7), 582-588]. In complex with the receptor, NTII was located at about 30A from the membrane surface, the tip of its loop II plunges into the ligand-binding pocket between the alpha/gamma or alpha/delta nAChR subunits, while the loops I and III contact nAChR by their tips only in a 'surface-touch' manner. The toxin structure undergoes some changes during the final complex formation (for 1.45rmsd in 15-25ps according to AMBER'99 molecular dynamics simulation), which correlates with NMR data. The data on the mobility and accessibility of spin- and fluorescence labels in free and bound NTII were used in MD simulations. The binding process is dependent on spontaneous outward movement of the C-loop earlier found in the AChBP complexes with alpha-cobratoxin and alpha-conotoxin. Among common features in binding of short- and long alpha-neurotoxins is the rearrangement of aromatic residues in the binding pocket not observed for alpha-conotoxin binding. Being in general very similar, the binding modes of short- and long alpha-neurotoxins differ in the ways of loop II entry into nAChR.
18 November, 2005
Novel conotoxins from Conus floridanus floridensis and Conus villepinii,
Möller, C., Rahmankhah, S., Lauer-Fields, J., Bubis, J., Fields, G.B. and Marí, F. (2005). A novel conotoxin framework with a helix-loop-helix (Cs alpha/alpha) fold. Biochemistry (in-press)
Department of Chemistry and Biochemistry and Center of Excellence in Biomedical and Marine Biotechnology, Florida Atlantic University, Boca Raton, Florida 33431, and Departamento de Biología Celular, Universidad Simón Bolívar, Sartenejas 1080, Venezuela.
Abstract: Venomous predatory animals, such as snakes, spiders, scorpions, sea anemones, and cone snails, produce a variety of highly stable cystine-constrained peptide scaffolds as part of their neurochemical strategy for capturing prey. Here we report a new family of four-cystine, three-loop conotoxins (designated framework 14). Three peptides of this family (flf14a-c) were isolated from the venom of Conus floridanus floridensis, and one (vil14a) was isolated from the venom of Conus villepinii, two worm-hunting Western Atlantic cone snail species. The primary structure for these peptides was determined using Edman degradation sequencing, and their cystine pairing was assessed by limited hydrolysis with a combination of CNBr and chymotrypsin under nonreducing, nonalkylating conditions in combination with MALDI-TOF MS analysis of the resulting peptidic fragments. CD spectra and nanoNMR spectroscopy of these conotoxins directly isolated from the cone snails revealed a highly helical secondary structure for the four conotoxins. Sequence-specific nanoNMR analysis at room temperature revealed a well-defined helix-loop-helix tertiary structure that resembles that of the Cs alpha/alpha scorpion toxins kappa-hefutoxin, kappa-KTx1.3, and Om-toxins, which adopt a stable three-dimensional fold where the two alpha-helices are linked by the two disulfide bridges. One of these conotoxins (vil14a) has a Lys/Tyr dyad, separated by approximately 6Å, which is a conserved structural feature in K+ channel blockers. The presence of this framework in scorpions and in cone snails indicates a common molecular imprint in the venom of apparently unrelated predatory animals and suggests a common ancestral genetic origin.
Interactions of fluorescent conotoxin GI with the nicotinic acetylcholine receptor
Schreiter, C., Gjoni, M., Hovius, R., Martinez, K.L., Segura, J.M., Vogel, H. (2005). Reversible Sequential-Binding Probe Receptor-Ligand Interactions in Single Cells. Chembiochem. 2005 Nov 4; [Epub ahead of print] PMID:
Ecole Polytechnique Federale de Lausanne (EPFL) Laboratoire de Chimie Physique des Polymeres et Membranes 1015 Lausanne, Switzerland, Fax: (+41) 21-693-6190.
Molluscan nicotinic acetylcholine receptors
van Nierop, P., Keramidas, A., Bertrand, S., van Minnen, J., Gouwenberg, Y., Bertrand, D. and Smit, A.B. (2005) Identification of molluscan nicotinic acetylcholine receptor (nAChR) subunits involved in formation of cation- and anion-selective nAChRs. J Neurosci. 25:10617-10626.
Department of Molecular and Cellular Neurobiology, Center for Neurogenomics and Cognitive Research, Faculty of Earth and Life Sciences, Vrije Universiteit, 1081 HV Amsterdam, The Netherlands, and Department of Neuroscience, University Medical Centre, 1211 Geneva 4, Switzerland.
Abstract: Acetylcholine (ACh) is a neurotransmitter commonly found in all animal species. It was shown to mediate fast excitatory and inhibitory neurotransmission in the molluscan CNS. Since early intracellular recordings, it was shown that the receptors mediating these currents belong to the family of neuronal nicotinic acetylcholine receptors and that they can be distinguished on the basis of their pharmacology. We previously identified 12 Lymnaea cDNAs that were predicted to encode ion channel subunits of the family of the neuronal nicotinic acetylcholine receptors. These Lymnaea nAChRs can be subdivided in groups according to the residues supposedly contributing to the selectivity of ion conductance. Functional analysis in Xenopus oocytes revealed that two types of subunits with predicted distinct ion selectivities form homopentameric nicotinic ACh receptor (nAChR) subtypes conducting either cations or anions. Phylogenetic analysis of the nAChR gene sequences suggests that molluscan anionic nAChRs probably evolved from cationic ancestors through amino acid substitutions in the ion channel pore, a mechanism different from acetylcholine-gated channels in other invertebrates.
Effects of alpha-conotoxin GI with
Barik, J. and Wonnacott, S. (2005) Indirect modulation by {alpha}7 nicotinic acetylcholine receptors of noradrenaline release in rat hippocampal slices: interaction with glutamate and GABA systems and effect of nicotine withdrawal. Mol Pharmacol. 2005 Nov 3; [Epub ahead of print]
University of BATH, UK.
Abstract: Nicotinic acetylcholine receptors (nAChR) can modulate transmitter release. Striatal [(3)H]dopamine ([(3)H]DA) release is regulated by presynaptic nAChR on dopaminergic terminals and alpha7 nAChR on neighbouring glutamatergic afferents. Here, we explored the role of alpha7 nAChR in the modulation of [(3)H]noradrenaline ([(3)H]NA) release from rat hippocampal slices. The nicotinic agonist anatoxin-a (AnTx) evoked monophasic [(3)H]NA release (EC50=1.2 microM) that was unaffected by alpha-conotoxin-MII or dihydro-beta-erythroidine, antagonists of alpha3/alpha6beta2* and beta2* nAChR respectively. In contrast AnTx-evoked striatal [(3)H]DA release was biphasic (EC50=138.9 nM; 7.1 microM) and blocked by these antagonists. At a high AnTx concentration (25 microM), alpha7 nAChR antagonists (methyllycaconitine, alpha-conotoxin-ImI) and glutamate receptor (GluR) antagonists (kynurenic acid, DNQX) partially inhibited [(3)H]NA release . The alpha7 nAChR-selective agonist choline evoked [(3)H]NA release (Emax=33% of that of AnTx) that was blocked by GluR antagonists, supporting a model in which alpha7 nAChR trigger glutamate release that subsequently stimulates [(3)H]NA release. A GABAergic component was also revealed: choline-evoked [(3)H]NA release was partially blocked by the GABAA receptor antagonist bicuculline, and co-application of bicuculline and DNQX fully abolished this response. These findings support alpha7 nAChR on GABAergic neurones that can promote GABA release that, in turn, leads to [(3)H]NA release, probably by disinhibition. To investigate the impact of chronic nicotine exposure on this model, rats were exposed for 14 days to nicotine (4 mg/kg/day) with or without 3 or 7 days withdrawal. alpha7 nAChR responses were selectively and transiently up-regulated after 3 days withdrawal. This functional up-regulation could contribute to the withdrawal effects of nicotine.
Mondal, S., Vijayan, R., Shichina, K., Babu, R.M., Ramakumar, S. (2005). I-superfamily conotoxins: sequence and structure analysis. In Silico Biol. 5 (4):0050 [Epub ahead of print]
Department of Physics, Indian Institute of Science, Bangalore 560 012, India; Email: ramak@physics.iisc.ernet.in.
Abstract: I-superfamily conotoxins have four-disulfide bonds with cysteine arrangement C-C-CC-CC-C-C, and they inhibit or modify ion channels of nerve cells. They have been characterized only recently and are relatively less well studied compared to other superfamily conotoxins. We have detected selective and sensitive sequence pattern for I-superfamily conotoxins. The availability of sequence pattern should be useful in protein family classification and functional annotation. We have built by homology modeling, a theoretical structural 3D model of ViTx from Conus virgo, a typical member of I-superfamily conotoxins. The modeling was based on the available 3D structure of Janus-atracotoxin-Hv1c of Janus-atracotoxin family whose members have been suggested as possible biopesticides. A study comparing the theoretically modeled structure of ViTx, with experimentally determined structures of other toxins, which share functional similarity with ViTx, reveals the crucial role of C-terminal region of ViTx in blocking therapeutically important voltage-gated potassium channels.
Wada, T., Imanishi, T., Kawaguchi, A., Mori, M.X., Mori, Y., Imoto, K. and Ichida, S. (2005). Effects of Calmodulin and Ca(2+) Channel Blockers on omega-conotoxin GVI A Binding to Crude Membranes from alpha(1B) Subunit (Ca(v)2.2) Expressed BHK Cells and Mice Brain Lacking the alpha(1B) Subunits. Neurochem Res. 30:1045-1054.
Departments of Biological Chemistry, School of Pharmaceutical Sciences, Kinki University , Kowakae 3-4-1, 577-8502, Higashiosaka , Japan, seiji@phar.kindai.ac.jp.
Abstract:Characteristics for the specific binding of (125)I-omega-CTX GVIA and (125)I-omega-CTX MVIIC to crude membranes from BHKN101 cells expressing the alpha(1B) subunits of Ca(v)2.2 channels and from mice brain lacking the alpha(1B) subunits of Ca(v)2.2 channels, particularly, the effects of CaM and various Ca(2+) channel blockers on these specific bindings were investigated. Specific binding of (125)I-omega-CTX GVIA to the crude membranes from BHKN101 cells was observed, but not from control BHK6 cells. omega-CTX GVIA, omega-CTX MVIIC and omega-CTX SVIB inhibited the specific binding of (125)I-omega-CTX GVIA to crude membranes from BHKN101 cells, and the IC(50) values for omega-CTXGVIA, omega-CTX MVIIC and omega-CTX SVIB were 0.07, 8.5 and 1.7 nM, respectively. However, omega-agatoxin IVA and calciseptine at concentrations of 10(-9)-10(-6) M did not inhibit specific binding. Specific binding was also about 80% inhibited by 20 mug protein/ml CaM. The amount of (125)I-omega-CTX GVIA (30 pM) specifically bound to membranes from brain of knockout mice lacking alpha(1B) subunits of Ca(v)2.2 channels was about 30% of that to the crude membranes from brain of wild-type. On the other hand, specific binding of (125)I-omega-CTX MVIIC (200 pM) was observed on the crude membranes of both BHKN101 and control BHK6 cells. The specific binding of (125)I-omega-CTX MVIIC (200 pM) was not inhibited by omega-CTX GVIA and omega-CTX SVIB, and also omega-Aga IVA and calciseptine at concentrations of 10(-9)-10(-7) M, although specific binding was almost completely dose dependently inhibited by non-radiolabeled omega-CTX MVIIC (IC(50) value was about 0.1 nM). 20 mug protein/ml CaM did not inhibit specific binding. Therefore, these results suggest that BHKN101 cells have a typical Ca(v)2.2 channels which are also inhibited by CaM and have not specific binding sites for omega-CTX MVIIC, although omega-CTX MVIIC is a blocker for both Ca(v)2.1 (alpha(1A; )P/Q-type) and Ca(v)2.2 channels.
Xenome Limited Launches MultiCentre Clinical Trial For Severe Cancer Pain
BRISBANE, Australia and SAN DIEGO, Nov. 15 /PRNewswire/ -- Xenome Limited today announced it had commenced a Phase I/IIa trial of Xen2174, a new class of peptide therapeutic for the treatment of severe intractable pain. (see Story from BioSpace.com)
1 November, 2005
Video Clip: Conus textile and Conus geographus
Cone shell cures ( Printable Page)
Script: When it comes to research on venom and converting it into useful drugs, studies involving exotic snakes or brightly colored frogs seem to attract the most attention. However, one of the most promising new venom-derived drugs actually comes from a very modest-looking sea snail.
Check out the numerous and varied Cone-shell Fact bites
31 October, 2005
Toxic Snail Venoms Yielding New Painkillers
See also the article about Cone Shells on the Australian Institute of Marine Science (AIMS) site "Dangerous marine animals of Northern Australia".
Kohn, A. J. and Almasi, K. N. (1993) Imposex in Australian Conus. J. Mar. Biol. Assoc. U.K.
73 (1), 241-244.
Duda, T. F. and Rolan, E. (2005) Explosive radiation of Cape Verde Conus, a marine species flock.
Molecular Ecology 14 , 267-272.
Conotoxins: essential data (Toxins List, Biological Weapons Gateway).
Ziconotide not for patients with psychiatric symptoms - a review
Wermeling, D.P. (2005) Ziconotide, an intrathecally administered N-type calcium channel antagonist for the treatment of chronic pain. Pharmacotherapy. 25:1084-1094.REVIEW.
Abstract: Ziconotide is a novel peptide that blocks the entry of calcium into neuronal N-type voltage-sensitive calcium channels, preventing the conduction of nerve signals. N-type calcium channels are present in the superficial laminae of the dorsal horn of the spinal cord. In various animal models of pain, intrathecal administration of ziconotide blocked nerve transmission and nociception. The United States Food and Drug Administration recently approved ziconotide intrathecal infusion for the management of severe chronic pain in patients who require intrathecal therapy and who are intolerant of or refractory to other treatment, such as systemic analgesics, adjunctive therapies, or intrathecal morphine. The drug has a narrow therapeutic window and a lag time for the onset and offset of analgesia and adverse events. In early clinical trials, frequent and severe psychiatric and central nervous system adverse effects were associated with rapid intrathecal infusion (0.4 microg/hr) and frequent up-titration (every 12 hrs). Therefore, patients with psychiatric symptoms are not candidates for this drug. Drug trials of external intrathecal catheters and microinfusion devices demonstrated a 3% risk of meningitis. A low initial infusion rate of 0.1 microg/hour and limiting infusion rate increases to 2-3 times/week are now recommended. Patients responsive to intrathecal ziconotide require an implanted infusion system to receive long-term therapy.
Outline: An intrathecal formulation of a synthetic version of a toxin used by a fish-eating marine snail to catch its prey has been approved as a treatment for severe, chronic pain.
The Food and Drug Administration approved ziconotide for intrathecal (IT) infusion for managing severe chronic pain “in patients for whom intrathecal therapy is warranted and who are intolerant of or refractory to other treatment, such as systemic analgesics, adjunctive therapies, or IT morphine.” It is being marketed under the trade name Prialt by Elan Pharmaceuticals Inc.
Ziconotide, which is not an opioid, is a synthetic version of a conopeptide used by a species of marine snail, Conus magus, to sting fish. In nature the toxin “is so powerful it stops the fish dead in its track, and the snail eats it,” said Mark Wallace, M.D., director of the center for pain and palliative medicine at the University of California, San Diego.
The synthetic version of this “conotoxin” is an N-type calcium channel antagonist. N-type calcium channels are located mainly in the dorsal horn cells of the spinal cord, predominantly on the superficial layers, in the area of substantia gelatinosa where pain fibers synapse, Dr. Wallace explained. Ziconotide “blocks those calcium channels at the level where these pain fibers meet up,” essentially shutting them down, he said, noting that opioids also have the same effect.
The three trials that led to the approval included patients with “really refractory” pain due to various causes, including low back pain, cancer pain, neuropathic pain, pain from nervous system injuries, and HIV-related pain, said Dr. Wallace, an investigator in the studies and a consultant to the manufacturer.
The three pivotal trials used the Visual Analog Scale of Pain Intensity (VASPI), as the primary end point.
The most recent trial was a multicenter study in 220 patients with severe chronic pain, described by most as refractory to treatments including IT morphine.
Patients were first taken off IT medications and stabilized on analgesics that included opiates and then treated with placebo or ziconotide. At 3 weeks, VASPI scores had improved by a mean of 12% from baseline vs. a 5% mean improvement for patients on placebo, a highly significant difference. During treatment, the use of non-IT opioids dropped by 24% among patients on ziconotide, compared with 17% among those on placebo.
Dr. Wallace said ziconotide treatment is not associated with addiction, withdrawal, or tolerance, and it is not a controlled substance. The most common side effects are neurologic, including neurocognitive impairment and dizziness. In ziconotide trials, dizziness, nausea, confusion, and headache were among the most common side effects, reported in at least 25% of patients.
The drug comes with a black box warning that people can develop severe psychiatric symptoms and neurologic impairment during treatment, and it is contraindicated in people with a history of psychosis.
Chen, Z.J. and Minneman, K.P. (2005) Recent progress in alpha1-adrenergic receptor research. Acta Pharmacol Sin. 26: 1281-1287.
Abstract: alpha1-Adrenergic receptors (AR) play an important role in the regulation of physiological responses mediated by norepinephrine and epinephrine, particularly in the cardiovascular system. The three cloned alpha1-AR subtypes (alpha1A, alpha1B, and alpha1D) are G protein-coupled receptors that signal through the Gq/11 signaling pathway, each showing distinct pharmacological properties and tissue distributions. However, due to the lack of highly subtype-selective drugs, the functional roles of individual subtypes are still not clear. Development of new subtype-specific drugs will greatly facilitate the identification of the functions of each subtype. Conopeptide rho-TIA has been found to be a new alpha1B-AR selective antagonist with different modes of inhibition at alpha1-AR subtypes. In addition, recent studies using genetically engineered mice have shed some light on alpha1-AR functions in vivo, especially in the cardiovascular system and brain. Several proteins have been shown to interact directly with particular alpha1-AR, and may be important in regulating receptor function. Receptor heterodimerization has been shown to be important for cell surface expression, signaling and internalization. These new observations are likely to help elucidate the functional roles of individual alpha1-AR subtypes.
21 October, 2005
Ziconotide and palliative care
Prommer, E.E. (2005) Ziconotide: can we use it in palliative care? Am J Hosp Palliat Care. 22: 369-374. Comment by Narayana, A.K. in: Am J Hosp Palliat Care. 2005 Nov-Dec;22(6):408.
Abstract: Ziconotide (PRIALT) is a new nonopioid treatment for chronic pain. It is a peptide that is the synthetic analog of the omega-conotoxin, derived from the marine snail, Conus magus. The therapeutic benefit of ziconotide derives from its potent and selective blockade of neuronal N-type voltage-sensitive calcium channels. Interference with these channels inhibits input from pain-sensing primary nociceptors. A recent clinical trial demonstrated that ziconotide has a significant analgesic effect compared to placebo in patients considered intolerant or refractory to other treatment such as systemic analgesics, adjunctive therapies, or intrathecal (IT) morphine. Thus, ziconotide is the first of a new class of agents--N-type calcium channel blockers, or NCCBs. Ziconotide may represent another option for patients with refractory pain.
Division of Hematology/Oncology, UCLA School of Medicine, Los Angeles, California, USA.
10 October, 2005
An alpha-conotoxin from Conus victoriae prevents pain
Satkunanathan, N., Livett, B., Gayler, K., Sandall, D., Down, J. and Khalil, Z. (2005) Alpha-conotoxin Vc1.1 alleviates neuropathic pain and accelerates functional recovery of injured neurones. Brain Res. 1059: 149-158.
National Ageing Research Institute, and Department of Biochemistry and Molecular Biology, University of Melbourne, Parkville Victoria 3010, Australia.
Abstract: This paper demonstrates the capacity of the neuronal nicotinic acetylcholine receptor (nAChR) antagonist alpha-conotoxin Vc1.1 to inhibit pain responses in vivo. Vc1.1 suppressed pain behaviors when tested in two models of peripheral neuropathy of the rat sciatic nerve, the chronic constriction injury (CCI) and partial nerve ligation (PNL) models. Mechanical hyperalgesia was assessed using an Ugo Basile Analgesymeter. Vc1.1 was administered by intramuscular bolus injection near the site of injury at doses of 0.036 mug, 0.36 mug and 3.6 mug in CCI rats and at a dose of 0.36 mug in PNL rats. Vc1.1 was also administered contralaterally in CCI rats at doses of 0.36 mug and 3.6 mug. Treatment started after the development of hyperalgesia and continued for 7 days. Vc1.1 significantly attenuated mechanical hyperalgesia in both CCI and PNL rats for up to a week following cessation of treatment. Vc1.1 also accelerated functional recovery of injured neurones. A blister was raised over the footpad innervated by the peripheral terminals of the injured nerve. The ability of these terminals to mount an inflammatory vascular response upon perfusion of the blister base with substance P provided a measure of functional recovery. This study shows that alpha-conotoxin Vc1.1, a neuronal nAChR antagonist, suppressed mechanical pain responses associated with peripheral neuropathy in rats in vivo and accelerated functional recovery of the injured neurones. A role for neuronal nAChRs in the analgesic activity of Vc1.1 is proposed.
Novel pain relief via marine snails
Sharp, D. Novel pain relief via marine snails. The Lancet 366 (9484)439-440, 2005.
Abstract: Most conopeptides (conotoxins and related compounds) with potential clinical applications are in the early stage of development (Table 1). The exception is ziconotide,(6) a synthetic drug but with a 25-aminoacid sequence identical to that of the parent venom constituent in C magus. These peptides have a wide range of targets. Listing eight conopeptides under development for neurological conditions, including pain, [Livett BG] et al(5) noted competitive blockade of N-type nicotinic acetylcholine receptors, N-type calcium-channel blockade, inhibition of the neuronal noradrenaline transporter, binding to the neurotensin receptor and selective inhibition of N-methyl-D-aspartate receptors. Ziconotide is a selective blocker of N-type voltage-sensitive calcium-channels, which are abundant in dorsal root ganglion cells, so the spinal transmission of pain messages is interrupted. It has no effect on opioid receptors. This drug has been approved in the USA and Europe for the management, via the intrathecal route only, of severe resistant pain; it is on the market in the USA. Animal studies did not encourage the development of ziconotide as an epilepsy drug and clinical studies of its neuroprotective properties were stopped because of a lowering of blood pressure. Severe pain, in which the patient will usually be at the limit of morphine intake, is the only approved indication.
To date only one controlled clinical trial on ziconotide has been fully published (8) however, trial data are available (9) and the results of three pivotal controlled studies are summarised in Table 2. Clinical information on other conopeptides is not abundant so far-eg, for the Amrad product, AM336, there is just one poster presentation on findings in seven patients (10). Trials have involved cautious dose-titration, and the results for ziconotide in Table 2 are those for the end of the initial titration phase. From the ten controlled longer-term open-label, and other studies of intrathecal ziconotide, in well over 1000 patients, a picture of the drug's adverse reactions profile is emerging. Serious reactions in three closely monitored trial patients were reported in detail 5 years ago (11). The most frequent reactions (recorded in 10% or more of patients) are confusion, dizziness, nystagmus, memory impairment, headache, blurred vision, nausea and vomiting, abnormal gait, and asthenia. A novel non-opioid class of drugs for the management of severe pain would be welcome. Whether synthetic conopeptides or new chemical compounds built round their structures fit the bill remains to be seen.
Reference
Novel "M"-Superfamily conotoxin from Conus vexillum
Jiang H, Wang CZ, Xu CQ, Fan CX, Dai XD, Chen JS, Chi CW (2005) A novel M-superfamily conotoxin with a unique motif from Conus vexillum. Peptides. 2005, Sept. 19 [ahead of publication]
Abstract: Cone snails are tropical marine mollusks that envenomate prey with a complex mixture of neuropharmacologically active compounds for the purpose of feeding and defence, each evolved to act in a highly specific manner on different parts of the nervous system. Here, we report the peptide purification, molecular cloning, chemical synthesis, and functional characterization of a structurally unique toxin isolated from the venom of Conus vexillum. The novel peptide, designated Vx2, was composed of 21 amino acid residues cross-linked by 3 disulfide bonds (WIDPSHYCCCGGGCTDDCVNC). Intriguingly, its mature peptide sequence shows low level of similarity with other identified conotoxins, and its unique motif (-CCCGGGC-) was not reported in other Conus peptides. However, its signal peptide sequence shares high similarity with those of the M-superfamily conotoxins. Hence, Vx2 could be classified into a new family of the M-superfamily.
Binding of alpha-conotoxin ImI from Conus imperialis to Aplysia acetylcholine binding protein
Hansen, S.B., Sulzenbacher, G., Huxford, T., Marchot, P., Taylor, P. and Bourne, Y. (2005) Structures of Aplysia AChBP complexes with nicotinic agonists and antagonists reveal distinctive binding interfaces and conformations. EMBO J. 24: 3635-3646
Abstract: Upon ligand binding at the subunit interfaces, the extracellular domain of the nicotinic acetylcholine receptor undergoes conformational changes, and agonist binding allosterically triggers opening of the ion channel. The soluble acetylcholine-binding protein (AChBP) from snail has been shown to be a structural and functional surrogate of the ligand-binding domain (LBD) of the receptor. Yet, individual AChBP species display disparate affinities for nicotinic ligands. The crystal structure of AChBP from Aplysia californica in the apo form reveals a more open loop C and distinctive positions for other surface loops, compared with previous structures. Analysis of Aplysia AChBP complexes with nicotinic ligands shows that loop C, which does not significantly change conformation upon binding of the antagonist, methyllycaconitine, further opens to accommodate the peptidic antagonist, alpha-conotoxin ImI, but wraps around the agonists lobeline and epibatidine. The structures also reveal extended and nonoverlapping interaction surfaces for the two antagonists, outside the binding loci for agonists. This comprehensive set of structures reflects a dynamic template for delineating further conformational changes of the LBD of the nicotinic receptor.
Mechanisms leading to conotoxin diversity in Conus striatus
Pi, C., Liu, Y., Peng, C., Jiang, X., Liu, J., Xu, B., Yu, X., Yu, Y., Jiang, X., Wang, L., Dong, M., Chen, S. and Xu, A.L. (2006). Analysis of expressed sequence tags from the venom ducts of Conus striatus: focusing on the expression profile of conotoxins. Biochimie 88: 131-140.
Abstract: Cone snails (genus Conus) are predatory marine gastropods that use venom peptides for interacting with prey, predators and competitors. A majority of these peptides, generally known as conotoxins demonstrate striking selectivity in targeting specific subtypes of ion channels and neurotransmitter receptors. So they are not only useful tools in neuroscience to characterize receptors and receptor subtypes, but offer great potential in new drug research and development as well. Here, a cDNA library from the venom ducts of a fish-hunting cone snail species, Conus striatus is described for the generation of expressed sequence tags (ESTs). A total of 429 ESTs were grouped into 137 clusters or singletons. Among these sequences, 221 were toxin sequences, accounting for 52.1% (corresponding to 19 clusters) of all transcripts. A-superfamily (132 ESTs) and O-superfamily conotoxins (80 ESTs) constitute the predominant toxin components. Some non-disulfide-rich Conus peptides were also found. The expression profile of conotoxins also explained to some extent the pharmacological and physiological reactions elicited by this typical piscivorous species. For the first time, a nonstop transcript of conotoxin was identified, which is suggestive that alternative polyadenylation may be a means of post-transcriptional regulation of conotoxin production. A comparison analysis of these conotoxins reveals the different variation and divergence patterns in these two superfamilies. Our investigations indicate that focal hyper-mutation, block substitution and exon shuffling are three main mechanisms leading to the conotoxin diversity in a species . The comprehensive set of Conus gene sequences allowed the identification of the representative classes of conotoxins and related components, which may lay the foundation for further research and development of conotoxins.
Production of an isotopically (15)N/(13)C-enriched conopeptide
Kumar, G.S., Ramasamy, P., Sikdar, S.K. and Sarma, S.P. (2005). Overexpression, purification, and pharmacological activity of a biosynthetically derived conopeptide. Biochem Biophys Res Commun. 335:965-972.
Abstract: A high yielding fusion protein system based on the protein cytochrome b(5) has been used for the production of novel 13-residue acyclic conopeptide. This peptide, Mo1659, can be liberated from the carrier protein using CNBr cleavage and subsequent purification using RP-HPLC methods. The yield of isotopically enriched peptides is high, ranging from 3 to 4mg of purified peptide from a 500ml culture, indicating that this system can be widely used for peptide production. Biosynthetic Mo1659 is active on non-inactivating K(+) channel much like the natural Mo1659, despite the absence of C-terminal amidation. Heteronuclear NMR studies show that the peptide exists in a conformational equilibrium involving proline-10. To our knowledge this is the first report of the production of an isotopically (15)N/(13)C-enriched conopeptide.
4 October, 2005
Mechanism of action of Contulakin-G
Cognetix Inc. of Salt Lake City, Utah, have announced in a Business Wire dated 27 September 2005 that it has identified the novel mechanism of action for CGX-1160, its lead compound in clinical development for the treatment of chronic intractable pain.
About Cognetix. Cognetix, Inc. is a private drug discovery and development company focused on the commercialization of conopeptide-based CNS pharmaceuticals. Conopeptides are a highly specialized, but structurally and functionally diverse group of peptides produced by venomous Conus sea snails, or cone snails These snails use a complex venom "cocktail" composed of between 50 and 200 distinct and unique peptides. These peptides target a variety of therapeutically important ion channels and receptors with unprecedented potency and selectivity and represent a large and diverse pharmacopeia of bioactive molecules. This potent and subtype selective activity at many molecular targets is of primary interest in Cognetix' pharmaceutical drug development program. . The Company has a strong drug discovery program in the field of pain, powered by a platform consisting of the most extensive conopeptide libraries and conopeptide-related intellectual property position in the world. Cognetix believes it has the largest patent portfolio in the world on conopeptide sequences, uses, synthesis, and methods. This patent portfolio includes more than 80 issued or allowed U.S. and international patents and an additional 80 patents pending.
Further information: Contact Cognetix, Inc. Roger L. Flowerdew, 801-581-0400 rflowerdew@cognetix.com www.cognetix.com
21 September, 2005
Cone shells on display
20 September, 2005
Review on pharmaceutical potential of marine toxins
Watters, M.R. (2005) Tropical marine neurotoxins: venoms to drugs.
Semin Neurol. 25: 278-289. REVIEW
Abstract: Neurotoxic venoms are common among tropical marine creatures, which have specialized apparatuses for delivery of the venoms. These include jellyfish and anemones, venomous cone snails, venomous fish, stingrays, sea snakes, and venomous octopuses. Numerous toxic neuropeptides are found within these venoms, and some can discriminate between closely related intracellular targets, a characteristic that makes them useful to define cation channels and attractive for drug development. A synthetic derivative of an omega-conotoxin is now available, representing a new class of analgesics. In general, toxic marine venoms contain proteins that are heat labile, providing opportunity for therapeutic intervention following envenomation, while ingestible seafood toxins are thermostable toxins. Ingestible toxins found in the tropics include those associated with reef fish, pufferfish, and some shellfish, which serve as food-chain vectors for toxins produced by marine microorganisms.
Engineered Peptides
Clark, R.J., Fischer, H., Dempster, L., Daly, N.L., Rosengren, K.J., Nevin, S.T., Meunier,
F.A., Adams, D.J. and Craik, D.J. (2005) Engineering stable peptide toxins by means of backbone cyclization: Stabilization of the {alpha}-conotoxin MII.
Proc Natl Acad Sci U S A. 102: 13767-13772.
Abstract: Conotoxins (CTXs), with their exquisite specificity and potency, have recently created much excitement as drug leads. However, like most peptides, their beneficial activities may potentially be undermined by susceptibility to proteolysis in vivo. By cyclizing the alpha-CTX MII by using a range of linkers, we have engineered peptides that preserve their full activity but have greatly improved resistance to proteolytic degradation. The cyclic MII analogue containing a seven-residue linker joining the N and C termini was as active and selective as the native peptide for native and recombinant neuronal nicotinic acetylcholine receptor subtypes present in bovine chromaffin cells and expressed in Xenopus oocytes, respectively. Furthermore, its resistance to proteolysis against a specific protease and in human plasma was significantly improved. More generally, to our knowledge, this report is the first on the cyclization of disulfide-rich toxins. Cyclization strategies represent an approach for stabilizing bioactive peptides while keeping their full potencies and should boost applications of peptide-based drugs in human medicine.
Conotoxin ImI from Conus imperialis
Kang, T.S., Vivekanandan, S., Jois, S.D., Kini, R.M. (2005) Effect of C-Terminal Amidation on Folding and Disulfide-Pairing of alpha-Conotoxin ImI. Angew Chem Int Ed Engl. 44 : 6333-6637.
Abstract: No abstract
Chi-Conotoxin Xen2174, from Conus marmoreus
Nielsen, C.K., Lewis, R.J., Alewood, D., Drinkwater, R., Palant, E., Patterson, M., Yaksh,
T.L., McCumber, D., Smith, M.T. (2005) Anti-allodynic efficacy of the chi-conopeptide, Xen2174, in rats with neuropathic pain. Pain 118: 112-124.
Abstract: Xen2174 is a structural analogue of Mr1A, a chi-conopeptide recently isolated from the venom of the marine cone snail, Conus marmoreus. Although both chi-conopeptides are highly selective inhibitors of the norepinephrine transporter (NET), Xen2174 has superior chemical stability relative to Mr1A. It is well-known that tricyclic antidepressants (TCAs) are also potent NET inhibitors, but their poor selectivity relative to other monoamine transporters and various G-protein-coupled receptors, results in dose-limiting side-effects in vivo. As TCAs and the alpha(2)-adrenoceptor agonist, clonidine, have established efficacy for the relief of neuropathic pain, this study examined whether intrathecal (i.t.) Xen2174 alleviated mechanical allodynia in rats with either a chronic constriction injury of the sciatic nerve (CCI-rats) or an L5/L6 spinal-nerve injury. The anti-allodynic responses of i.t. Mr1A and i.t. morphine were also investigated in CCI-rats. Paw withdrawal thresholds were assessed using calibrated von Frey filaments. Bolus doses of i.t. Xen2174 produced dose-dependent relief of mechanical allodynia in CCI-rats and in spinal nerve-ligated rats. Dose-dependent anti-allodynic effects were also produced by i.t. bolus doses of Mr1A and morphine in CCI-rats, but a pronounced 'ceiling' effect was observed for i.t. morphine. The side-effect profiles were mild for both chi-conopeptides with an absence of sedation. Confirming the noradrenergic mechanism of action, i.t. co-administration of yohimbine (100nmol) with Xen2174 (10nmol) abolished Xen2174s anti-allodynic actions. Xen2174 appears to be a promising candidate for development as a novel therapeutic for i.t. administration to patients with persistent neuropathic pain.
26 July, 2005
Updates to Conus Biodiversity Website
Dr. Alan Kohn has announced some additions and updates to his Conus Biodiversity Website.
We have also added a new menu labeled “Information.” New pages here include:
In the near future we hope to add additional images of primary types of species described through 1900, and images of the shells of specimens whose genes and proteins have now been sequenced."
25 July, 2005
Conus victoriae venom component, alpha-conotoxin Vc1.1 (ACV1) enters Phase I clinical trials
Two articles about our cone shell work have appeared in the local press.
Pain-killer comes out of its shell
Another article "Molecular miners find pain relief drugs from the sea" [With an increasing age demographic in our society the need for more effective pain suppressing compounds is a priority. ACV1 may fill this unmet need] This appeared in Yztimes ("YZage" web bulletin)
4 July, 2005
alpha-conotoxin discriminates between alpha9alpha10 and alpha7 nicotinic receptors
McIntosh, J.M., Plazas, P.V., Watkins, M., Gomez-Casati, M.E,, Olivera, B.M. and Elgoyhen, A.B (2005). A novel alpha-conotoxin, PeIA, cloned from Conus pergrandis discriminates between rat alpha9alpha10 and alpha 7 nicotinic cholinergic receptors. J Biol Chem. 280: 30107-30112
Abstract: The alpha9 and alpha10 nicotinic cholinergic subunits assemble to form the receptor believed to mediate synaptic transmission between efferent olivocochlear fibers and hair cells of the cochlea, one of the few examples of postsynaptic function for a non-muscle nicotinic acetylcholine receptor (nAChR). However, it has been suggested that the expression profile of alpha9 and alpha10 overlaps with that of alpha7 in the cochlea and in sites like dorsal root ganglion neurons, peripheral blood lymphocytes, developing thymocytes and skin. We now report the cloning, total synthesis and characterization of a novel toxin alpha-conotoxin PeIA that discriminates between alpha9alpha10 and alpha7 nAChRs. This is the first toxin to be identified from Conus pergrandis, a species found in deep waters of the Western Pacific. alpha-Conotoxin PeIA displayed a 260-fold higher selectivity for alpha-bungarotoxin-sensitive alpha9alpha10 nAChRs compared to alpha-bungarotoxin-sensitive alpha7 receptors. The IC(50) of the toxin was 6.9 +/- 0.5 nM and 4.4 +/- 0.5 nM, for recombinant alpha9alpha10 and wild-type hair cell nAChRs, respectively. alpha-Conotoxin PeIA bears high resemblance to alpha-conotoxins MII and GIC isolated from Conus magus and Conus geographus, respectively. However, neither alpha-conotoxin MII nor alpha-conotoxin GIC at concentrations of 10 muM, blocked acetylcholine responses elicited in Xenopus oocytes injected with the alpha9 and alpha10 subunits. Among neuronal non-alpha-bungarotoxin sensitive receptors, alpha-conotoxin PeIA was also active at alpha3beta2 receptors and chimeric alpha6/alpha3beta2beta3 receptors. alpha-Conotoxin PeIA represents a novel probe to differentiate responses mediated either through alpha9alpha10 or alpha7 nAChRs in those tissues where both receptors are expressed.
Are cone snail venoms of danger to fish ?
Mebs, D. and Kauferstein, S. (2005) Ichthyotoxicity caused by marine cone snail venoms? Toxicon. 46: 355-356
Zentrum der Rechtsmedizin, University of Frankfurt, Kennedyallee 104, D-69596 Frankfurt am Main, Germany.
Abstract: Ten venoms from marine cone snails were tested for ichthyotoxic effects on zebra fish (Brachydanio rerio) when added to the water. Only two venoms, from Conus capitaneus and Conus episcopatus, produced lethal effects at high concentrations (50-300mug/ml) within 20-90min. No sedative or hypnotic symptoms were observed. The experiments confirm that Conus venoms exert a quick and prompt activity only by parenteral injection into the prey as it is performed by the snail.
24 June, 2005
Human trials for pain relief drugs from sea
See these links for further details.
University of Melbourne: UniNews article
ASX announcement from Metabolic Pharmaceuticals
Please click on the hyperlink below:
http://www.metabolic.com.au/?lo2mt&4z6U&1dtY&mBg
Armishaw, C.J. and Alewood, P.F.(2005) Conotoxins as research tools and drug leads. Curr Protein Pept Sci. 6: 221-240.
Abstract: The complex mixture of biologically active peptides that constitute the venom of Conus species provides a rich source of ion channel neurotoxins. These peptides, commonly known as conotoxins, exhibit a high degree of selectivity and potency for different ion channels and their subtypes making them invaluable tools for unravelling the secrets of the nervous system. Furthermore, several conotoxin molecules have profound applications in drug discovery, with some examples currently undergoing clinical trials. Despite their relatively easy access by chemical synthesis, rapid access to libraries of conotoxin analogues for use in structure-activity relationship studies still poses a significant limitation. This is exacerbated in conotoxins containing multiple disulfide bonds, which often require synthetic strategies utilising several steps. This review will examine the structure and activity of some of the known classes of conotoxins and will highlight their potential as neuropharmacological tools and as drug leads. Some of the classical and morerecent approaches to the chemical synthesis of conotoxins, particularly with respect to the cont rolled formation of disulfide bonds will be discussed in detail. Finally, some examples of structure-activity relationship studies will be discussed, as well as some novel approaches for designing conotoxin analogues.
23 June, 2005
*Leipold, E., *Hansel, A., **Olivera, B.M., ***Terlau, H. and *Heinemann, S.H. (2005) Molecular interaction of delta-conotoxins with voltage-gated sodium channels. FEBS Letters, 579 : 3881-3884.
*Institute of Molecular Cell Biology, Research Unit Molecular and Cellular Biophysics, Friedrich Schiller University Jena, Drackendorfer Strasse 1, D-07747 Jena, Germany
Abstract: Various neurotoxic peptides modulate voltage-gated sodium (NaV) channels and thereby affect cellular excitability. delta-Conotoxins from predatory cone snails slow down inactivation of NaV channels, but their interaction site and mechanism of channel modulation are unknown. Here, we show that delta-conotoxin SVIE from Conus striatus interacts with a conserved hydrophobic triad (YFV) in the domain-4 voltage sensor of NaV channels. This site overlaps with that of the scorpion alpha-toxin Lqh-2, but not with the alpha-like toxin Lqh-3 site. delta-SVIE functionally competes with Lqh-2, but exhibits strong cooperativity with Lqh-3, presumably by synergistically trapping the voltage sensor in its “on” position.
22 June, 2005
Brown, M.A., Begley, G.S., Czerwiec, E., Stenberg, L.M., Jacobs, M., Kalume, D.E., Roepstorff, P., Stenflo, J., Furie, B.C. and Furie, B. (2005) Precursors of novel Gla-containing conotoxins contain a carboxy-terminal recognition site that directs gamma-carboxylation. Biochemistry 44: 9150-9159.
Marine Biological Laboratory, Woods Hole, Massachusetts 02543, Center for Hemostasis and Thrombosis Research, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts 02215, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, DK-5230 Odense M, Denmark, and Department of Clinical Chemistry, Lund University, University Hospital, Malmo S-205 02, Sweden.
Abstract: Vitamin K-dependent gamma-glutamyl carboxylase catalyzes the conversion of glutamyl residues to gamma-carboxyglutamate. Its substrates include vertebrate proteins involved in blood coagulation, bone mineralization, and signal transduction and invertebrate ion channel blockers known as conotoxins. Substrate recognition involves a recognition element, the gamma-carboxylation recognition site, typically located within a cleavable propeptide preceding the targeted glutamyl residues. We have purified two novel gamma-carboxyglutamate-containing conotoxins, Gla-TxX and Gla-TxXI, from the venom of Conus textile. Their cDNA-deduced precursors have a signal peptide but no apparent propeptide. Instead, they contain a C-terminal extension that directs gamma-carboxylation but is not found on the mature conotoxin. A synthetic 13-residue "postpeptide" from the Gla-TxXI precursor reduced the K(m) for the reaction of the Conus gamma-carboxylase with peptide substrates, including FLEEL and conantokin-G, by up to 440-fold, regardless of whether it was positioned at the N- or C-terminal end of the mature toxin. Comparison of the postpeptides to propeptides from other conotoxins suggested some common elements, and amino acid substitutions of these residues perturbed gamma-carboxylation of the Gla-TxXI peptide. The demonstration of a functional and transferable C-terminal postpeptide in these conotoxins indicates the presence of the gamma-carboxylation recognition site within the postpeptide and defines a novel precursor structure for vitamin K-dependent polypeptides. It also provides the first formal evidence to prove that gamma-carboxylation occurs as a post-translational rather than a cotranslational process.
22 June, 2005
Tom Avril has written a nice piece for the Philadelphia Inquirer about the use of Ziconotide for pain relief. The article is entitled "From depths of the ocean, bright hope for medicines".
21 June, 2005
Monika Bayrhuber and colleagues, from Max-Plank institute, Gottingen, Germany, describe a new 60-residue neurotoxin from the venom of Conus striatus that retains 2 out of the 3 disufide links typically found in these small, basic protein module which targets voltage-gated potassium channels.
Bayrhuber, M., Vijayan, V., Ferber, M., Graf, R., Korukottu, J., Imperial, J., Garrett, J.E., Olivera, B.M., Terlau, H., Zweckstetter, M. and Becker, S. (2005) Conkunitzin-S1 Is the first member of a new Kunitz-type neurotoxin family: Structural and functional characterization. J. Biol. Chem. 280: 23766-23770
Abstract: Conkunitzin-S1 (Conk-S1) is a 60-residue neurotoxin from the venom of the cone snail Conus striatus that interacts with voltage-gated potassium channels. Conk-S1 shares sequence homology with Kunitz-type proteins but contains only two out of the three highly conserved cysteine bridges, which are typically found in these small, basic protein modules. In this study the three-dimensional structure of Conk-S1 has been solved by multidimensional NMR spectroscopy. The solution structure of recombinant Conk-S1 shows that a Kunitz fold is present, even though one of the highly conserved disulfide cross-links is missing. Introduction of a third, homologous disulfide bond into Conk-S1 results in a functional toxin with similar affinity for Shaker potassium channels. The affinity of Conk-S1 can be enhanced by a pore mutation within the Shaker channel pore indicating an interaction of Conk-S1 with the vestibule of potassium channels.
21 June, 2005
Don Barclay, is presently tackling some of his "yet-to-be-identified" cones from Guam. His mystery page can be viewed at: http://www.manandmollusc.net/Mystery_shell_pages/donB-mystery.html. If you can help with identification please send your ideas to Don Barclay.
20 June, 2005
Al-Sabi, Ahmed (2004): Structural and functional studies of kM-conotoxin RIIIK interaction with Shaker-related potassium channels from trout fish (tSha 1). Dissertation: Bremen, November 2004.
19 June, 2005
An informative web page on Conus purpurascens and its potassium channel directed venom component (k-conotoxin PVIIA) and its sodium channel toxin (delta-conotoxin PVIA), is available here, at a site hosted by Prof. W. Stuhmer Director, the Max-Planck-Institut für experimentelle Medizin Abteilung Molekulare Biologie neuronaler Signale, Gottingen, Germany, in conjunction with the laboratory of Prof. Baldomero Olivera in Salt Lake City, Utah, USA.
16 June, 2005
Celie, P.H., Kasheverov, I.E., Mordvintsev, D.Y., Hogg, R.C., van Nierop, P., van Elk, R., van Rossum-Fikkert, S.E., Zhmak, M.N., Bertrand, D., Tsetlin, V., Sixma, T.K. and Smit, A.B. (2005) Crystal structure of nicotinic acetylcholine receptor homolog AChBP in complex with an alpha-conotoxin PnIA variant. Nat Struct Mol Biol, 12: 582-588
Abstract: Conotoxins (Ctx) form a large family of peptide toxins from cone snail venoms that act on a broad spectrum of ion channels and receptors. The subgroup alpha-Ctx specifically and selectively binds to subtypes of nicotinic acetylcholine receptors (nAChRs), which are targets for treatment of several neurological disorders. Here we present the structure at a resolution of 2.4 A of alpha-Ctx PnIA (A10L D14K), a potent blocker of the alpha(7)-nAChR, bound with high affinity to acetylcholine binding protein (AChBP), the prototype for the ligand-binding domains of the nAChR superfamily. alpha-Ctx is buried deep within the ligand-binding site and interacts with residues on both faces of adjacent subunits. The toxin itself does not change conformation, but displaces the C loop of AChBP and induces a rigid-body subunit movement. Knowledge of these contacts could facilitate the rational design of drug leads using the Ctx framework and may lead to compounds with increased receptor subtype selectivity.
15 June, 2005
John Roach for Pulse of the Planet reports in National Geographic News June 14, 2005, about "Toxic Snail Venoms Yielding New Painkillers, Drugs"
Studying cone snail venom, researchers have derived other new treatments for pain, epilepsy, and incontinence. The drugs are in clinical and preclinical trials. Scientists say more experimental drugs are in development at research labs around the world. "They could have a cure to prostate cancer, a cure to AIDS. I have no idea," said Jon-Paul Bingham, a biochemist at Clarkson University in Potsdam, New York. Bingham is a leading researcher on cone snails. He said there are more than 500 cone snail species, each able to produce more than a hundred unique toxins. Each toxin is a potential new drug. Scientists have studied less than one percent of them.
Bingham worries that "it's getting harder and harder to [collect] these snails." Cone snails are found primarily in coral reefs in warm, tropical waters. But as Eric Chivian, the founder and director of the Center for Health and the Global Environment at the Harvard Medical School in Cambridge, Massachusetts, noted, "We are destroying coral reefs." According to Chivian, some 26 percent of the world's reefs are damaged beyond repair, and another 30 to 50 percent are severely degraded. The proven potential of drugs derived from cone snail venom is the best example of the cost associated with the loss of the world's coral reefs, Chivian said. "I'm totally convinced cone snails as a group may have more potential for new medicines than any other genus in nature," Chivian said. (A genus is a group of closely related species.) "The number of different toxins they have developed over some 30 to 50 million years of evolution is unparalleled in nature."
In nature, cone snails use their venom to immobilize prey, such as fish, mollusks, and worms. Cone snails harpoon their prey with pointed tongues that are shaped like hypodermic needles. The snails then pump their prey's flesh full of toxins. "The poor animal, even if it gets away, [it] breaks off the harpoon. The likelihood of it being fatal is 100 percent, basically," Bingham said. "For any [human] who has been stung by a cone shell, the chance of mortality is 90 percent." The potency and complexity of the venom fascinates scientists. Over millions of years cone snails have evolved toxins that target specific species in specific environments. No one toxin is exactly like another. In addition, any cone snail can alter the compounds in its venom at will. Scientists are studying how each cone snail toxin affects its victim. The results are improving our understanding of how cells, such as those that make up the nervous system, interact and communicate. The research is also leading to the development of new drugs. "We are learning what nature has put in these animals and applying the strategy to other organisms," Bingham said. Consider, for example, the painkiller ziconotide, the cone-snail-venom-derived drug that was approved by the FDA in December.
The painkiller (brand name: Prialt) is injected through a special pump into the fluid surrounding a patient's spinal cord. The drug blocks nerve channels that ordinarily transmit pain. Unlike opium-derived painkillers, such as morphine, ziconotide is said to pose a low risk of addiction or increased tolerance and has few side effects. "If you stop using the drug, the pain comes back. And no matter how long you've used [the drug], you don't have to increase the dosage to kill the pain," Bingham said. Given the potential of snail venom in the development of new drugs, Bingham said he is concerned that the rush to discover more promising toxins may contribute to the decline the world's cone snail species.
Cone Snail Conservation
Chivian, of Harvard Medical School, said the main threat to cone snails is the loss of coral reefs, which he attributes to bleaching (a type of slow death evident when multihued coral reefs turn a ghostly white) and disease outbreaks induced by global warming. Coastal development, pollution, and destructive fishing practices also negatively affect coral reefs. As for cone snails, their shells are collected and sold by the thousands at curio shops to tourists. Chivian said that additional collecting for biomedical research, if done irresponsibly, could further imperil the snails. But by far the greatest impact on the snails has been caused by the loss of coral reefs, Chivian said.
Bingham, the Clarkson University researcher, is taking no chances. He advocates responsible use of cone snails, including minimal collection for research. To that end, Bingham and his colleagues are among the few scientists in the world who "milk" cone snails for venom. Milking involves a tricky—and potentially deadly—process: enticing a captive cone snail to harpoon a condom-covered test tube opening. (Other scientists typically retrieve venom by dissecting the venom glands of dead snails.) Bingham notes that milking reduces the number of cone snails he needs to take from the wild for research. The process also allows him to study how and why the snails change the compounds in their venom over time. Each change in venom compounds yields a new toxin with unknown potential. Once scientists identify the compounds in any given cone snail toxin, researchers can readily make it in synthetic form, Bingham added, noting that this eliminates the need to extract more venom from the snails.
Chivian, meanwhile, said any measure to protect cone snails is appreciated. "We are losing species at a great rate without identifying them and without any knowledge of what value they may have for our health and our lives," he said.
15 June, 2005
14 June, 2005
The University of Melbourne posted a Press Release on Friday 10 June about progress by Metabolic Pharmaceuticals Limited, Melbourne, Australia, who are developing ACV1 (conotoxin Vc1.1 from Conus victoriae) for clinical use and announced that it is to the stage where it is ready to enter Phase I clinical trials.
Metabolic Pharmaceuticals Limited received an AusIndustry "Commercial Ready" Grant of AUD$449,902 to support a Phase 1 Clinical Trial of ACV1. This alpha-conotoxin has recently been shown to block ion transport through neuronal nicotinic receptors in excised segments of human sural nerve.
ACV1 (conotoxin Vc1.1) to enter clinical trials for diabetic neuropathy
Melbourne University researchers have found snails could be a pain cure for diabtes patients. Trials of a cone
shell peptide from the cone snail (Conus victoriae) have been successful in preventing pain in animal models of human neuropathic pain. University researcher Assoc. Prof. Bruce Livett and colleagues at the Bio21 Molecular Science & Biotechnology Institute and at the National Ag |
Lu, X.C., Williams, A.J., Wagstaff, J.D., Tortella, F.C. and Hartings, J.A. (2005). Effects of delayed intrathecal infusion of an NMDA receptor antagonist on ischemic injury and peri-infarct depolarizations. 
Aguilar, M.B., Lezama-Monfil, L., Maillo, M., Pedraza-Lara, H., Lopez-Vera, E., Heimer
de la Cotera, E.P. (2005) A biologically active hydrophobic T-1-conotoxin from the venom of Conus spurius. 
Aguilar, M.B., Lopez-Vera, E., Ortiz, E., Becerril, B., Possani, L.D., Olivera, B.M. and
Heimer de la Cotera, E.P. (2005) A novel conotoxin from Conus delessertii with posttranslationally modified lysine residues. 
Underhill, W. (2005) Nature's Drugs. 
Conus textile envenomates a mollusk
Dr. Livett with a selection of cone shells.
