NEXT SCHEDULED UPDATE 5 November 2009 No. unique visitors = 52,816 up to 27 March 2005 (2062 days) + Scroll Down OR Jump to Latest Entry.
April 2006 "Pain Relief Drugs from the Sea"
Analgesic Component of Venom (ACV1) from Cone Snails :
For a one-page description of Cone Shells and their Conotoxins click here For a video simulation of cone shell envenomaton click here
Bruce Livett's more recent publications (1998-2009)
Site Map of Cone Shells and Conotoxins HomePage
3. SEARCH this site using FreeFind
Visit the Please mail me
suggestions at the following address: b.livett@unimelb.edu.au
Great Classical Music, Jazz & Opera at Global Music Network;
31 October, 2009
Evolution of Conus in the Cape Verde Islands
Regina Lopes da Cunha (2009) Tempo and mode of evolution of the genus Conus in the Cape Verde Islands. 12th Annual Molluscan Forum, Thursday 12th November 2009, The Dorothea Bate Room, Natural History Museum, London, UK
20 October, 2009
Novel I(2)-superfamily conotoxins from Conus spurius, Gulf of Mexico
Zamora-Bustillos R, Aguilar MB, Falcón A. (2009) Identification, by molecular cloning, of a novel type of I(2)-superfamily conotoxin precursor and two novel I(2)-conotoxins from the worm-hunter snail Conus spurius from the Gulf of Méxic. Peptides. 2009 Oct 14. [Epub ahead of print]
Laboratorio de Neurofarmacología Marina, Departamento de Neurobiología Celular y Molecular, Instituto de Neurobiología, Universidad Nacional Autónoma de México, Campus Juriquilla, Querétaro 76230, México.
Abstract: cDNA was prepared from the venom duct of a single Conus spurius specimen collected near the coast of Campeche, México. From it, PCR products were generated aiming to clone I-conotoxin precursors. Thirty clones were sequenced and predicted to encode ten distinct precursors: seven of I(2)-conotoxins and three of I(2)-like-conotoxins. These precursors contain three different, mature toxins, sr11a, sr11b and sr11c, of which two are novel and one (sr11a) has been previously purified and characterized from the venom of this species. The precursors include a 26- (I(2)) or 23- residue signal peptide (I(2)-like), a 31-residue "pro" region (I(2)-like), and a 32-residue mature toxin region (I(2) and I(2)-like). In addition, all the precursors have a 13-residue "post" region which contains a gamma-carboxylation recognition sequence that directs the gamma-carboxylation of Glu-9 and Glu-10 of toxin sr11a and, possibly, Glu-13 of toxin sr11b and Glu-9 of toxin sr11c. This is the first time that a "post" region has been found in precursors of I-conotoxins that also contain a "pro" region. The "post" peptide is enzymatically processed to yield the amidated mature toxin sr11a, which implies that gamma-carboxylation occurs before amidation. Phylogenetic analysis at the whole precursor level indicates that the I(2)-like-conotoxins of C. spurius are more related to I(2)-conotoxins than to I(1)- and I(3)-conotoxins from other species, and that they might represent a new subgroup of the I(2)-superfamily. The three I-conotoxins from C. spurius have charge differences at seven to nine positions, suggesting that they might have different molecular target types or subtypes.
17 October, 2009
Human envenomation by Conus regius
Haddad, V. Jr., Coltro, M and Simone, LRL (2009) Report of a human accident caused by Conus regius (Gastropoda, Conidae). Revista da Sociedade Brasileira de Medicina Tropical 42: 446-448.[Article.pdf]
Botuscatu School of Medicine, Sao Paulo State Universiy, Botucatu, SP, Brazil; Vital Brasil Hospital, Butantan Institute, Sao Paulo, SP, Brazil; Femorale Shells, Sao Paulo, SP, Brazil. Address correspondence to Prof. Vidal Haddad Junior, Caixa Postal 557, 18618-000 Botucatu, SP, Brazil
Abstract: Conus regius is a venomous molluc in the Conidae family, which includes species responsible for severe or even fatal accidents affecting human beings. This is the first report of a clinical case involving this species. It consisted a puncture in the right hand of a diver who presented parashesia and movement diffikculty in the whole limb. The manifestations disappeared after around twelve hours, without sequelae.
Marine pharmacology - a REVIEW
Glaser KB, Mayer AMS (2009) A renaissance in marine pharmacology: from preclinical curiosity to clinical reality Biochem Pharmacol 2009,Sep,01;78(5):440-448
Cancer Research R47J-AP9, Abbott Laboratories, Abbott Park, IL 60064-6121, USA. keith.glaser@abbott.com
Abstract: Marine pharmacology, the pharmacology of marine natural products, has been for some time more associated with marine natural products chemistry rather than mainstay pharmacology. However, in recent years a renaissance has occurred in this area of research, and has seen the US Food & Drug Administration (FDA) approval in 2004 of Prialt (ziconotide, omega-conotoxin MVIIA) the synthetic equivalent of a conopeptide found in marine snails, used for the management of severe chronic pain. Furthermore Yondelis) (trabectedin, ET-743) an antitumor agent scovered in a marine colonial tunicate, and now produced synthetically, receiving Orphan Drug designation from the European Commission (EC) and FDA for soft tissue sarcomas and ovarian cancer and its registration in 2007 in the EU for the treatment of soft tissue sarcoma. The approval/marketing of so few marine natural products has come after many years of research primarily by the academic community and the sporadic involvement of major pharmaceutical companies. This commentary, through the opinions provided by several leaders in the marine natural products field, will examine the potential reasons and perceptions from both the academic and pharmaceutical communities regarding the development of marine natural products as viable therapeutic entities.
Some recent cone shell and conopeptide papers
More to come as now back on line and updating from 10 April 2008
MS identification of conotoxin Vc1.1 and conopeptides along venom duct of Conus victoriae
Townsend, A., Livett, BG, Bingham, J-P, Truong, H-T, Karas, JA, O’Donnell, P, Williamson, NA, Purcell, AW and Scanlon D (2009) Mass spectral identification of Vc1.1 and differential distribution of conopeptides in the venom duct of Conus victoriae. Effect of post-translational modifications and disulfide isomerisation on bioactivity. Int. J. Peptide Res and Therap. 15 (3): 195-203. [Abstract], [ Article.pdf]
Dept. Biochemistry & Molecular Biology and Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, VIC 3010, Australia; Metabolic Pharmaceuticals Ltd., Level 3, 509 St. Kilda Rd. Melbourne, VIC. 3004, Australia; Dept Molecular Biosciences and Bioengineering, University of Hawaii, Honolulu, HI 96822, USA
Abstract: Molluscs of the genus Conus (cone shells) are carnivorous, feeding on marine worms, small fish and other marine molluscs. They capture their prey by injecting venom containing hundreds of neurally active peptide components. These peptides are classed as conotoxins and consist of small disulfide-bonded peptides exhibiting a high degree of post-translational modifications (PTMs). The functional roles of these modifications remain largely unknown. Two of the most frequently observed modifications are gamma-carboxylation of glutamate and hydroxylation of proline (Buczek et al. Cell Mol Life Sci 62:3067, 2005). Vc1.1 is an alpha-conotoxin from Conus victoriae (Sandall et al. Biochemistry 42(22):6904–6911, 2003) and the only form of this peptide which has been detected in the venom is the gamma-glutamate and hydroxyproline (Vc1.1.P6O:E14Gla) version of the molecule (Jakubowski et al. Toxicon 47(6):688–699, 2006). In order to investigate the role of PTMs, we did mass spectral profiling of the venom duct of C. victoriae looking at changes in mass and the number of peptides detected. We synthesised a number of predicted Vc1.1-PTM peptides together with the three possible disulfide isoforms of Vc1.1 and assessed the possible functional role of the PTM conopeptides by measuring the in vitro activity at the cognate neuronal nicotinic acetylcholine receptors (nAChRs). In addition we looked for their presence Vc1.1 venom by mass spectrometry and by this approach we were able to detect unmodified Vc1.1 in C. victoriae venom for the first time.
Dicarba analogues of alpha-Conotoxin ImI
MacRaild CA, Illesinghe J, van Lierop BJ, Townsend AL, Chebib M, Livett BG, Robinson AJ, Norton RS (2009) Structure and activity of (2,8)-dicarba-(3,12)-cystino alpha-ImI, an alpha-conotoxin containing a nonreducible cystine analogue. J Med Chem. 52 : 755-762. [Abstract], [ Article.pdf], [ SUPPORTING INFO.pdf] The Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Parkville 3050, Victoria, Australia. Abstract: The alpha-conotoxins are potent and selective antagonists of nicotinic acetylcholine receptors (nAChR). Exploitation of these and other peptides in research and clinical settings has been hampered by the lability of the disulfide bridges that are essential for toxin structure and activity. One solution to this problem is replacement of cystine bridges with nonreducible dicarba linkages. We explore this approach by determining the solution structure and functional characteristics of a dicarba analogue of the alpha-conotoxin alpha-ImI, (2,8)-dicarba-(3,12)-cystino alpha-ImI. The structure of the dicarba analogue was similar to that of native alpha-ImI, with differences attributable to the different covalent geometry of the disulfide and dicarba bridges. Dicarba-alpha-ImI maintained inhibitory activity of nAChR comparable to that of native alpha-ImI in two in vitro assays. These findings confirm the potential of the dicarba linkage to improve stability while maintaining alpha-conotoxin function.
Dicarba analogues of alpha-Conotoxin RgIA
Robinson AJ, van Lierop BJ, Garland RD, Teoh E, Elaridi J, Illesinghe JP and Jackson WR (2009) Regioselective formation of interlocked dicarba bridges in naturally occurring cyclic peptide toxins using olefin metathesis. Chem. Commun. (Camb). 2009 Jul 28:(28)4293-4295.
School of Chemistry, Centre for Green Chemistry, Monash University, Clayton 3800, Victoria, Australia.
Abstract: Bis-dicarba analogues of native dicystine-containing-conotoxin Rg1A, an analgesic peptide isolated from cone snail venom, were constructed on resin using a regioselective metathesis–hydrogenation strategy.
Conus anemone vs Conus novaehollandiae
John K. Tucker (2009) Floraconus anemone: An Example of
Circular Overlap? The Cone Collector 11: 8-24
Great Rivers Field Station, Illinois Natural History Survey, USA
Abstract: This article in The Cone Collector Vol. 11 (2009) provides an extensive account of the naming and distribution of Conus anemone and variants around the coast of Australia
Conus anemone
Discrimination of alpha-conotoxins for subtypes of nicotinic receptors
Turner M, Eidemiller S, Martin B, Narver A, Marshall J, Zemp L, Cornell KA, McIntosh JM, McDougal OM. (2009) Structural basis for alpha-conotoxin potency and selectivity. Bioorg Med Chem. 2009 Aug 15;17(16):5894-5899
Department of Chemistry and Biochemistry, Boise State University, ID 83725-1520, USA.
Abstract: Parkinson's disease is a debilitating movement disorder characterized by altered levels of alpha(6)beta(2) * ( * indicates the possible presence of additional subunits) nicotinic acetylcholine receptors (nAChRs) localized on presynaptic striatal catecholaminergic neurons. alpha-Conotoxin MII (alpha-CTx MII) is a highly useful ligand to probe alpha(6)beta(2) nAChRs structure and function, but it does not discriminate among closely related alpha(6) * nAChR subtypes. Modification of the alpha-CTx MII primary sequence led to the identification of alpha-CTx MII[E11A], an analog with 500-5300-fold discrimination between alpha(6) * subtypes found in both human and non-human primates. alpha-CTx MII[E11A] binds most strongly (femtomolar dissociation constant) to the high affinity alpha(6) nAChR, a subtype that is selectively lost in Parkinson's disease. Here, we present the three-dimensional solution structure for alpha-CTx MII[E11A] as determined by two-dimensional (1)H NMR spectroscopy to 0.13+/-0.09A backbone and 0.45+/-0.08A heavy atom root-mean-square deviation from mean structure. Structural comparisons suggest that the increased hydrophobic area of alpha-CTx MII[E11A] relative to other members of the alpha-CTx family may be responsible for its exceptionally high affinity for alpha6alpha4beta2 * nAChR as well as discrimination between alpha(6)beta(2) and alpha(3)beta(2) containing nAChRs. This finding may enable the rational design of novel peptide analogs that demonstrate enhanced specificity for alpha(6) * nAChR subunit interfaces and provide a means to better understand nAChR structural determinants that modulate brain dopamine levels and the pathophysiology of Parkinson's disease.
Conotoxins as pain killers - REVIEW
Zhao CJ, Dai QY. (2009) [Recent advances in study of antinociceptive conotoxins]
[Article in Chinese] Yao Xue Xue Bao. 2009 Jun;44(6):561-565.
Institute of Biotechnology, Academy of Military Medical Sciences, Beijing 100071,
China.
Abstract: The Conus venom is secreted by the duct and theca of venom. Most of conotoxins
are composed of 10-40 amino acid residues with several disulfide bridges. They
can specifically target neurotransmitter receptors including nAChRs, calcium ion
channels, sodium ion channels and potassium ion channels, etc. Some conotoxins,
such as that target N-Ca2+ channels, nAChR alpha9alpha10 subtype, TTX-R Na+
channels or NMDA receptors, have potent antinociceptive activities, omega-MVIIA,
an Ca2+ channels blocker was approved by FDA in December, 2004 for marketing.
Because of lower molecular weight and high specificity, conotoxins are the
powerful pharmacology tools and potent analgesics without addiction. This review
briefly summarizes the research progress of antinociceptive conotoxins and
addresses on their targets and structure-activity relationships.
Omega toxins - REVIEW
Bingham JP, Mitsunaga E, Bergeron ZL(2009) Drugs from Slugs - Past, Present and Future Perspectives of omega-Conotoxin
Research. Chem Biol Interact. 2009 Oct 1. [Epub ahead of print]
Department of Molecular Biosciences and Bioengineering, University of Hawaii,
Honolulu, HI, 96822, USA.
Abstract: Peptides from the venom of carnivorous cone shells have provided six decades of
intense research, which has led to the discovery and development of novel
analgesic peptide therapeutics. Our understanding of this unique natural marine
resource is however somewhat limited. Given the past pharmacological record,
future investigations into the toxinology of these highly venomous tropical
marine snails will undoubtedly yield other highly selective ion channel
inhibitors and modulators. With over a thousand conotoxin-derived sequences
identified to date, those identified as ion channel inhibitors represent only a
small fraction of the total. Here we discuss our present understanding of
conotoxins, focusing on the omega-conotoxin peptide family, and illustrate how
such a seemingly simple snail has yielded a highly effective clinical drug.
Novel alpha conotoxin from Conus quercinus
Peng C, Chen W, Han Y, Sanders T, Chew G, Liu J, Hawrot E, Chi C, Wang C (2009).
Characterization of a novel alpha4/4-conotoxin, Qc1.2, from vermivorous Conus
quercinus.Acta Biochim Biophys Sin (Shanghai). 2009 Oct;41(10):858-864.
Institute of Protein Research, Tongji University, Shanghai, China.
Abstract: As part of continuing studies of the identification of gene organization and
cloning of novel alpha-conotoxins, the first alpha4/4-conotoxin identified in a
vermivorous Conus species, designated Qc1.2, was originally obtained by cDNA and
genomic DNA cloning from Conus quercinus collected in the South China Sea. The
predicted mature toxin of Qc1.2 contains 14 amino acid residues with two
disulfide bonds (I-III, II-IV connectivity) in a native globular configuration.
The mature peptide of Qc1.2 is supposed to contain an N-terminal
post-translationally processed pyroglutamate residue and a free carboxyl
C-terminus. This peptide was chemically synthesized and refolded for further
characterization of its functional properties. The synthetic Qc1.2 has two
interconvertible conformations in aqueous solution, which may be due to the
cis-trans isomerization of the two successive Pro residues in its first Cys loop.
Using the Xenopus oocyte heterologous expression system, Qc1.2 was shown to
selectively inhibit both rat neuronal alpha3beta2 and alpha3beta4 subtypes of
nicotinic acetylcholine receptors with low potency. A block of about 63% and 37%
of the ACh-evoked currents was observed, respectively, and the toxin dissociated
rapidly from the receptors. Compared with other characterized alpha-conotoxin
members, the unusual structural features in Qc1.2 that confer to its receptor
recognition profile are addressed.
Neuroprotective effects of SNX185
Shahlaie K, Lyeth B, Gurkoff GG, Muizelaar JP, Berman RF (2009) Neuroprotective Effects of Selective N-Type VGCC Blockade on Stretch
Injury-Induced Calcium Dynamics in Cortical Neurons. J Neurotrauma. 2009 Sep 22. [Epub ahead of print]
University of California, Davis, Neurological Surgery, 4860 Y Street, Suite 3740,
Sacramento, California, United States, 95817, 916-734-3071, 916-452-2580;
krshahlaie@ucdavis.edu.
Abstract: Acute elevation in intracellular calcium ([Ca2+]i) following traumatic brain
injury (TBI) can trigger cellular mechanisms leading to neuronal dysfunction and
death. The mechanisms underlying these processes are incompletely understood, but
calcium influx through N-type VGCCs appears to play a central role. The present
study examined the time course of [Ca2+]i flux, glutamate release, and loss of
cell viability following injury using an in vitro neuronal-glial cortical cell
culture model of TBI. The effects of N-channel blockade with SNX185 (e.g.,
omega-conotoxin TIVA) before or after injury were also examined. Neuronal injury
produced a transient elevation in [Ca2+]i, increased glutamate release, and
resulted in neuronal and glial death. SNX185 administered before or immediately
after cell injury reduced glutamate release and increased survival of neurons and
astrocytes, whereas delayed treatment did not improve cell survival but
significantly facilitated the return of [Ca2+]i to baseline levels. The new
findings that N-type VGCCs are critically involved in injury-induced glutamate
release and recovery of [Ca2+]i argue for continued investigation of this
treatment strategy for clinical management of TBI. In particular, SNX-185 may
represent an effective class of drugs that can significantly protect injured
neurons from secondary insults that commonly occur after TBI.
Role of the Ca(V)2.3subtype in nociception
Yang L, Stephens GJ(2009) Effects of neuropathy on high-voltage-activated Ca(2+) current in sensory
neurones.Cell Calcium. 2009 Aug 31. [Epub ahead of print]
School of Pharmacy, University of Reading, PO Box 228, Whiteknights, Reading RG6
6AJ, United Kingdom.
Abstract: Voltage-dependent Ca(2+) channels (VDCCs) have emerged as targets to treat
neuropathic pain; however, amongst VDCCs, the precise role of the Ca(V)2.3
subtype in nociception remains unproven. Here, we investigate the effects of
partial sciatic nerve ligation (PSNL) on Ca(2+) currents in small/medium diameter
dorsal root ganglia (DRG) neurones isolated from Ca(V)2.3(-/-) knock-out and
wild-type (WT) mice. DRG neurones from Ca(V)2.3(-/-) mice had significantly
reduced sensitivity to SNX-482 versus WT mice. DRGs from Ca(V)2.3(-/-) mice also
had increased sensitivity to the Ca(V)2.2 VDCC blocker omega-conotoxin. In WT
mice, PSNL caused a significant increase in omega-conotoxin-sensitivity and a
reduction in SNX-482-sensitivity. In Ca(V)2.3(-/-) mice, PSNL caused a
significant reduction in omega-conotoxin-sensitivity and an increase in
nifedipine sensitivity. PSNL-induced changes in Ca(2+) current were not
accompanied by effects on voltage-dependence of activation in either
Ca(V)2.3(-/-) or WT mice. These data suggest that Ca(V)2.3 subunits contribute,
but do not fully underlie, drug-resistant (R-type) Ca(2+) current in these cells.
In WT mice, PSNL caused adaptive changes in Ca(V)2.2- and Ca(V)2.3-mediated
Ca(2+) currents, supporting roles for these VDCCs in nociception during
neuropathy. In Ca(V)2.3(-/-) mice, PSNL-induced changes in Ca(V)1 and Ca(V)2.2
Ca(2+) current, consistent with alternative adaptive mechanisms occurring in the
absence of Ca(V)2.3 subunits.
Alpha-conotoxin ImII and its ribbon isomer (ImIIiso)both block human alpha7 and muscle nAChRs equivalently
Kasheverov IE, Zhmak MN, Fish A, Rucktooa P, Khruschov AY, Osipov AV, Ziganshin
RH, D'hoedt D, Bertrand D, Sixma TK, Smit AB, Tsetlin VI(2009) Interaction of alpha-conotoxin ImII and its analogs with nicotinic receptors and acetylcholine-binding proteins: additional binding sites on Torpedo receptor. J Neurochem. 2009 Aug 27. [Epub ahead of print]
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of
Sciences, Moscow, Russia.
Abstract: alpha-Conotoxins interact with nicotinic acetylcholine receptors
(nAChRs) and acetylcholine-binding proteins (AChBPs) at the sites for
agonists/competitive antagonists. alpha-Conotoxins blocking muscle-type or alpha7
nAChRs compete with alpha-bungarotoxin. However, alpha-conotoxin ImII, a close
homolog of the alpha7 nAChR-targeting alpha-conotoxin ImI, blocked alpha7 and
muscle nAChRs without displacing alpha-bungarotoxin (Ellison et al. 2003, 2004),
suggesting binding at a different site. We synthesized alpha-conotoxin ImII, its
ribbon isomer (ImIIiso), 'mutant' ImII(W10Y) and found similar potencies in
blocking human alpha7 and muscle nAChRs in Xenopus oocytes. Both isomers
displaced [(125)I]-alpha-bungarotoxin from human alpha7 nAChRs in the cell line
GH(4)C(1) (IC(50) 17 and 23 muM, respectively) and from Lymnaea stagnalis and
Aplysia californica AChBPs (IC(50) 2.0-9.0 muM). According to SPR measurements,
both isomers bound to immobilized AChBPs and competed with AChBP for immobilized
alpha-bungarotoxin (K(d) and IC(50) 2.5-8.2 muM). On Torpedo nAChR,
alpha-conotoxin [(125)I]-ImII(W10Y) revealed specific binding (K(d) 1.5-6.1 muM)
and could be displaced by alpha-conotoxin ImII, ImIIiso and ImII(W10Y) with
IC(50) 2.7, 2.2 and 3.1 muM, respectively. As alpha-cobratoxin and
alpha-conotoxin ImI displaced [(125)I]-ImII(W10Y) only at higher concentrations
(IC(50)>/= 90 muM), our results indicate that alpha-conotoxin ImII and its
congeners have an additional binding site on Torpedo nAChR distinct from the site
for agonists/competitive antagonists.
Safety and Efficacy of intrathecal Ziconotide (PRIALT)
Smith HS, Deer TR(2009) Safety and efficacy of intrathecal ziconotide in the management of severe chronic
pain. Ther Clin Risk Manag. 2009 Jun;5(3):521-34. Epub 2009 Jul 12.
Albany Medical College, Department of Anesthesiology, Albany, New York, USA.
Abstract: Ziconotide is a conopeptide intrathecal (IT) analgesic which is approved by the
US Food and Drug Administration (FDA) for the management of severe chronic pain.
It is a synthetic equivalent of a naturally occurring conopeptide found in the
venom of the fish-eating marine cone snail and provides analgesia via binding to
N-type voltage-sensitive calcium channels in the spinal cord. As ziconotide is a
peptide, it is expected to be completely degraded by endopeptidases and
exopeptidases (Phase I hydrolytic enzymes) widely located throughout the body,
and not by other Phase I biotransformation processes (including the cytochrome
P450 system) or by Phase II conjugation reactions. Thus, IT administration, low
plasma ziconotide concentrations, and metabolism by ubiquitous peptidases make
metabolic interactions of other drugs with ziconotide unlikely. Side effects of
ziconotide which tend to occur more commonly at higher doses may include: nausea,
vomiting, confusion, postural hypotension, abnormal gait, urinary retention,
nystagmus/amblyopia, drowsiness/somnolence (reduced level of consciousness),
dizziness or lightheadedness, weakness, visual problems (eg, double vision),
elevation of serum creatine kinase, or vestibular side effects. Initially, when
ziconotide was first administered to human subjects, titration schedules were
overly aggressive and led to an abundance of adverse effects. Subsequently,
clinicians have gained appreciation for ziconotide's relatively narrow
therapeutic window. With appropriate usage multiple studies have shown ziconotide
to be a safe and effective intrathecal analgesic alone or in combination with
other intrathecal analgesics.
M-Superfamily of Conotoxins - REVIEW
Jacob RB, McDougal OM(2009) The M-superfamily of conotoxins: a review.
Cell Mol Life Sci. 2009 Aug 25. [Epub ahead of print]
Department of Chemistry and Biochemistry, Boise State University, 1910 University
Drive, Boise, ID, 83725-1520, USA.
Abstract: The focus of this review is the M-superfamily of Conus venom peptides. Disulfide
rich peptides belonging to the M-superfamily have three loop regions and the
cysteine arrangement: CC-C-C-CC, where the dashes represent loops one, two, and
three, respectively. Characterization of M-superfamily peptides has demonstrated
that diversity in cystine connectivity occurs between different branches of
peptides even though the cysteine pattern remains consistent. This superfamily is
subdivided into five branches, M-1 through M-5, based on the number of residues
in the third loop region, between the fourth and fifth cysteine residues.
M-superfamily peptides appear to be ubiquitous in Conus venom. They are largely
unexplained in indigenous biological function, and they represent an active area
of research within the scientific community.
Nicotinic receptors that mediate alpha-CtxMII-resistant
dopamine release
Grady SR, Salminen O, McIntosh JM, Marks MJ, Collins AC(2009) Mouse Striatal Dopamine Nerve Terminals Express alpha4alpha5beta2 and Two Stoichiometric Forms of alpha4beta2*-Nicotinic Acetylcholine Receptors. J Mol Neurosci. 2009 Aug 20. [Epub ahead of print]
Institute for Behavioral Genetics, University of Colorado, Boulder, CO, USA.
Abstract: Wild-type and alpha5 null mutant mice were used to identify nicotinic cholinergic
receptors (nAChRs) that mediate alpha-conotoxin MII (alpha-CtxMII)-resistant
dopamine (DA) release from striatal synaptosomes. Concentration-effect curves for
ACh-stimulated release (20 s) were monophasic when wild-type synaptosomes were
assayed but biphasic with synaptosomes from the alpha5 null mutant. Deleting the
alpha5 gene also resulted in decreased maximal ACh-stimulated
alpha-CtxMII-resistant DA release. When a shorter perfusion time (5 s) was used,
biphasic curves were detected in both wild-type and alpha5 null mutants,
indicative of high- and low-sensitivity (HS and LS) activity. In addition,
DHbetaE-sensitive (HS) and DHbetaE-resistant (LS) components were found in both
genotypes. These results indicate that alpha-CtxMII-resistant DA release is
mediated by alpha4alpha5beta2, (alpha4)(2)(beta2)(3) (HS), and
(alpha4)(3)(beta2)(2) (LS) nAChRs.
Omega-conotoxin GVIA mimetics
Andersson A, Baell JB, Duggan PJ, Graham JE, Lewis RJ, Lumsden NG, Tranberg CE,
Tuck KL, Yang A(2009) Omega-conotoxin GVIA mimetics based on an anthranilamide core: effect of
variation in ammonium side chain lengths and incorporation of fluorine. Bioorg Med Chem. 2009 Sep 15;17(18):6659-70. Epub 2009 Jul 30.
Institute for Molecular Bioscience, The University of Queensland, St. Lucia, QLD
4072, Australia.
Abstract: A number of omega-conotoxin GVIA mimetics based on an anthranilamide core were
prepared and tested for their affinity for rat brain Ca(v)2.2 channels. Features
such as the presence of hydroxyl and fluoro substituents on the tyrosine side
chain mimic, the length of the chains on the lysine/arginine side chain mimics
and the use of diguanidino and diamino substituents rather than
mono-guanidine/mono-amine substitution were examined. The diguanidinylated
compounds proved to be the most active and deletion of the hydroxyl substituent
had a limited influence on activity. The SAR associated with variation in the
lysine/arginine side chain mimics was not strong. The introduction of a fluoro
substituent into the tyrosine mimic produced the most active compound prepared in
this study (2g), with an EC(50) at rat brain Ca(v)2.2 channels of 6 microM.
Rapid detection of peptide toxins in serum
Fang J, Dong F, Wang N, He K, Liu B, Wu S, Li A, Zhang X(2009) Rapid detection of conotoxin SO(3) in serum using Cu-chelated magnetic beads coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
J Anal Toxicol. 2009 Jun;33(5):272-277.
National Center of Biomedical Analysis, Beijing 100039, China.
Abstract: A novel method based on Cu-chelated magnetic beads (Cu-Magbeads) and
matrix-assisted laser desorption/ionization time-of-flight mass spectrometry
(MALDI-TOF-MS) was developed for the rapid detection of peptide toxins in serum.
The peptides in the serum were efficiently adsorbed by the Cu-Magbeads, eluted
with methanol solution, and assayed by MALDI-TOF-MS. Specific peptides were
identified according to their characteristic mass-to-charge ratio values.
Conotoxin SO(3), a synthesized peptide, was used as a model to evaluate the
method. Conotoxin SO(3) was detected in human serum, as well as bovine and murine
serum, with a detection sensitivity in the low femtomole range. The assay was
performed within 40 min, without the need for a specific antibody or an expensive
reagent. It shows potential for future use in clinical and emergency rescue
practice because of its simplicity, high speed, and high sensitivity.
Protective effects of omega-conotoxin GVIA in optic neuritis
Gadjanski I, Boretius S, Williams SK, Lingor P, Knöferle J, Sättler MB, Fairless
R, Hochmeister S, Sühs KW, Michaelis T, Frahm J, Storch MK, Bähr M, Diem R(2009) Role of N-type voltage-dependent calcium channels in autoimmune optic neuritis. Ann Neurol. 66(1): 81-93
Department of Neurology, Georg-August University, Göttingen, Germany.
Abstract: OBJECTIVE: The aim of this study was to investigate the role of voltage-dependent
calcium channels (VDCCs) in axon degeneration during autoimmune optic neuritis.
METHODS: Calcium ion (Ca(2+)) influx into the optic nerve (ON) through VDCCs was
investigated in a rat model of optic neuritis using manganese-enhanced magnetic
resonance imaging and in vivo calcium imaging. After having identified the most
relevant channel subtype (N-type VDCCs), we correlated immunohistochemistry of
channel expression with ON histopathology. In the confirmatory part of this work,
we performed a treatment study using omega-conotoxin GVIA, an N-type specific
blocker. RESULTS: We observed that pathological Ca(2+) influx into ONs during
optic neuritis is mediated via N-type VDCCs. By analyzing the expression of VDCCs
in the inflamed ONs, we detected an upregulation of alpha(1B), the pore-forming
subunit of N-type VDCCs, in demyelinated axons. However, high expression levels
were also found on macrophages/activated microglia, and lower levels were
detected on astrocytes. The relevance of N-type VDCCs for inflammation-induced
axonal degeneration and the severity of optic neuritis was corroborated by
treatment with omega-conotoxin GVIA. This blocker led to decreased axon and
myelin degeneration in the ONs together with a reduced number of
macrophages/activated microglia. These protective effects were confirmed by
analyzing the spinal cords of the same animals. INTERPRETATION: We conclude that
N-type VDCCs play an important role in inflammation-induced axon degeneration via
two mechanisms: First, they directly mediate toxic Ca(2+) influx into the axons;
and second, they contribute to macrophage/microglia function, thereby promoting
secondary axonal damage. Ann Neurol 2009;66:81-93.
Flourescent alpha-conotoxin
Hone AJ, Whiteaker P, Christensen S, Xiao Y, Meyer EL, McIntosh JM(2009) A novel fluorescent alpha-conotoxin for the study of alpha7 nicotinic acetylcholine receptors. J Neurochem. 11: 80-89.
Interdepartmental Program in Neuroscience, University of Utah, Salt Lake City,
Utah, USA.
Abstract: Homomeric alpha7 nicotinic acetylcholine receptors are a well-established,
pharmacologically distinct subtype. The more recently identified alpha9 subunit
can also form functional homopentamers as well as alpha9alpha10 heteropentamers.
Current fluorescent probes for alpha7 nicotinic ACh receptors are derived from
alpha-bungarotoxin (alpha-BgTx). However, alpha-BgTx also binds to alpha9* and
alpha1* receptors which are coexpressed with alpha7 in multiple tissues. We used
an analog of alpha-conotoxin ArIB to develop a highly selective fluorescent probe
for alpha7 receptors. This fluorescent alpha-conotoxin, Cy3-ArIB[V11L;V16A],
blocked ACh-evoked alpha7 currents in Xenopus laevis oocytes with an IC(50) value
of 2.0 nM. Observed rates of blockade were minute-scale with recovery from
blockade even slower. Unlike FITC-conjugated alpha-BgTx, Cy3-ArIB[V11L;V16A] did
not block alpha9alpha10 or alpha1beta1deltaepsilon receptors. In competition
binding assays, Cy3-ArIB[V11L;V16A] potently displaced [(125)I]-alpha-BgTx
binding to mouse hippocampal membranes with a K(i) value of 21 nM. Application of
Cy3-ArIB[V11L;V16A] resulted in specific punctate labeling of KXalpha7R1 cells
but not KXalpha3beta2R4, KXalpha3beta4R2, or KXalpha4beta2R2 cells. This labeling
could be abolished by pre-treatment with alpha-cobratoxin. Thus,
Cy3-ArIB[V11L;V16A] is a novel and selective fluorescent probe for alpha7
receptors.
Selective alpha6 antagonist, alpha-conotoxin MII[H9A;L15A]
Jackson KJ, McIntosh JM, Brunzell DH, Sanjakdar SS, Damaj MI(2009) The role of {alpha}6-containing nicotinic acetylcholine receptors in nicotine reward and withdrawal. J Pharmacol Exp Ther. 331: 547-554
Virginia Commonwealth University.
Abstract: The alpha6 nicotinic acetylcholine receptor (nAChR) subunit is involved in
nicotine-stimulated dopamine release in the striatum. It is expressed in brain
regions and co-expressed with nAChR subtypes implicated in nicotine dependence
behaviors; hence, this subunit may play a role in nicotine dependence. Using the
alpha6-selective antagonist, alpha-conotoxin H9A; L15A (MII[H9A;L15A]), we
determined the role of alpha6* nAChRs in the pharmacological and behavioral
effects of nicotine. We measured effects of pre-treatment with MII[H9A;L15A] on
analgesia, locomotion, and body temperature following a single injection of
nicotine. Effects of MII[H9A;L15A] on nicotine reward were measured using the
conditioned place preference (CPP) paradigm. We further measured physical
(somatic signs, hyperalgesia) and affective (anxiety-related behavior,
conditioned place aversion (CPA)) nicotine withdrawal behaviors following
extended nicotine exposure. Results showed that MII[H9A;L15A] did not block acute
nicotine effects on the behaviors measured. Conversely, MII[H9A:l15A] blocked the
expression of nicotine CPP, as well as withdrawal-associated CPA and
anxiety-related behavior in the elevated plus maze, but not withdrawal-induced
somatic signs or hyperalgesia. These results suggest a role for the alpha6 nAChR
subunit in nicotine reward and affective nicotine withdrawal, but not acute
nicotine-induced or physical withdrawal behaviors.
Conotoxins as drug leads - REVIEW
Halai R, Craik DJ.(2009) Conotoxins: natural product drug leads. Nat Prod Rep. 26:526-536
The University of Queensland, Institute for Molecular Bioscience, Brisbane,
Queensland 4072, Australia.
Abstract: Venomous marine cone snails harbour a variety of small disulfide-rich peptides
called conotoxins, which target a broad range of ion channels, membrane
receptors, and transporters. More than 700 species of Conus are thought to exist,
each expressing a wide array of different peptides. Within this large repertoire
of toxins, individual conotoxins are able to discriminate between different
subtypes and isoforms of ion channels, making them valuable pharmacological
probes or leads for drug design. This review gives a brief background to the
discovery of conotoxins and describes their sequences, biological activities, and
applications in drug design.
Conus ebreus - variations in genomic alleles at Okinawa, Guam and Hawaii
Duda TF Jr, Chang D, Lewis BD, Lee T Geographic variation in venom allelic composition and diets of the widespread
predatory marine gastropod Conus ebraeus PLoS One. 2009 4:e6245.
Department of Ecology and Evolutionary Biology and Museum of Zoology, University
of Michigan, Ann Arbor, MI, USA. tfduda@umich.edu
Abstract: BACKGROUND: Members of the predatory gastropod genus Conus use a venom comprised
of a cocktail of peptide neurotoxins, termed conotoxins or conopeptides, to
paralyze prey and conotoxin gene family members diversify via strong positive
selection. Because Conus venoms are used primarily to subdue prey, the evolution
of venoms is likely affected by predator-prey interactions. METHODOLOGY/PRINCIPAL
FINDINGS: To identify the selective forces that drive the differentiation of
venoms within species of Conus, we examined the distribution of alleles of a
polymorphic O-superfamily conotoxin locus of Conus ebraeus at Okinawa, Guam and
Hawaii. Previous analyses of mitochondrial cytochrome oxidase I gene sequences
suggest that populations of C. ebraeus, a worm-eating Conus, are not structured
genetically in the western and central Pacific. Nonetheless, because the sample
size from Guam was relatively low, we obtained additional data from this location
and reexamined patterns of genetic variation at the mitochondrial gene at
Okinawa, Guam and Hawaii. We also utilized a DNA-based approach to identify prey
items of individuals of C. ebraeus from Guam and compared this information to
published data on diets at Okinawa and Hawaii. Our results show that conotoxin
allelic frequencies differ significantly among all three locations, with
strongest differentiation at Hawaii. We also confirm previous inferences that C.
ebraeus exhibits no genetic differentiation between Okinawa, Guam and Hawaii at
the mitochondrial locus. Finally, DNA-based analyses show that eunicid
polychaetes comprise the majority of the prey items of C. ebraeus at Guam; while
this results compares well with observed diet of this species at Okinawa, C.
ebraeus preys predominantly on nereid polychaetes at Hawaii.
CONCLUSIONS/SIGNIFICANCE: These results imply that strong selection pressures
affect conotoxin allelic frequencies. Based on the dietary information, the
selection may derive from geographic variation in dietary specialization and
local coevolutionary arms races between Conus and their prey.
Peptide interactions with the nicotinic receptor
Kasheverov IE, Utkin YN, Tsetlin VI (2009)Naturally occurring and synthetic peptides acting on nicotinic acetylcholine
receptors. Curr Pharm Des. 15:2430-2452.
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of
Sciences, Moscow, Russian Federation.
Abstract: Nicotinic acetylcholine receptors (nAChRs) are pentameric membrane-bound proteins
belonging to the large family of ligand-gated ion channels. nAChRs possess
various binding sites which interact with compounds of different chemical nature,
including peptides. Historically first peptides found to act on nAChR were
synthetic fragments of snake alpha-neurotoxins, competitive receptor antagonists.
Later it was shown that fragments of glycoprotein from rabies virus, having
homology to alpha-neurotoxins, and polypeptide neurotoxins waglerins from the
venom of Wagler's pit viper Trimeresurus (Tropidolaemus) wagleri bind in a
similar way, waglerins being efficient blockers of muscle-type nAChRs.
Neuropeptide substance P appears to interact with the channel moiety of nAChR.
beta-Amyloid, a peptide forming senile plaques in Alzheimer's disease, also can
bind to nAChR, although the mode of binding is still unclear. However, the most
well-studied peptides interacting with the ligand-binding sites of nAChRs are
so-called alpha-conotoxins, peptide neurotoxins from marine snails of Conus
genus. First alpha-conotoxins were discovered in the late 1970s, and now it is a
rapidly growing family due to isolation of peptides from multiple Conus species,
as well as to cloning, and chemical synthesis of new analogues. Because of their
unique selectivity towards distinct nAChR subtypes, alpha-conotoxins became
valuable tools in nAChR research. Recent X-ray structures of alpha-conotoxin
complexes with acetylcholine-binding protein, a model of nAChR ligand-binding
domains, revealed the details of the nAChR ligand-binding sites and provided the
basis for design of novel ligands.
Novel I-superfamily conotoxins from Conus in the South China Sea
Liu Z, Xu N, Hu J, Zhao C, Yu Z, Dai Q (2009) Identification of novel I-superfamily conopeptides from several clades of Conus
species found in the South China Sea. Peptides 30:1782-1787
Beijing Institute of Biotechnology, Beijing 100071, People's Republic of China.
Novel M-superfamily conotoxin from Conus litteratus
Wang L, Liu J, Pi C, Zeng X, Zhou M, Jiang X, Chen S, Ren Z, Xu A.
(2009) Identification of a novel M-superfamily conotoxin with the ability to enhance
tetrodotoxin sensitive sodium currents. Arch Toxicol. 83:925-932
State Key Laboratory of Biocontrol, Guangdong Province Key Laboratory of
Pharmaceutical Functional Genes, Department of Biochemistry, College of Life
Sciences, Sun Yat-sen (Zhongshan) University, 135 Xingangxi Road, 510275
Guangzhou, People's Republic of China.
Abstract: In this work, a novel M-superfamily conotoxin, designated lt3a, was purified from
the crude venom of Conus litteratus. Combined with peptide sequencing, MALDI-TOF
mass spectrometry and cDNA cloning techniques, the amino acid sequence of lt3a
was supposed to be DgammaCCgamma OQWCDGACDCCS, where O is hydroxyproline and
gamma is carboxyglutamate. The Cys framework of lt3a (-CC-C-C-CC-) is similar to
that of psi-, mu-, kappaM-conotoxins, which are representatives of M-conotoxins.
Peptide lt3a is categorized into M1 branch based on the number of residues in the
last Cys loop. Whole cell patch-clamp study on adult rat dorsal root ganglion
neurons indicated that lt3a could enhance tetrodotoxin-sensitive sodium currents.
This is a previously unknown function of M-superfamily conotoxins.
Use of selective conotoxins to block nicotinic receptors
Anderson DJ, Malysz J, Grønlien JH, El Kouhen R, Håkerud M, Wetterstrand C,
Briggs CA, Gopalakrishnan M.(2009) Stimulation of dopamine release by nicotinic acetylcholine receptor ligands in
rat brain slices correlates with the profile of high, but not low, sensitivity
alpha4beta2 subunit combination. Biochem Pharmacol.78:844-851.
Abstract: Neuroscience Research, Global Pharmaceutical Research and Development, Abbott
Laboratories, Abbott Park, IL 60064, USA. david.j.anderson@abbott.com
alpha4beta2 neuronal nicotinic receptors (nAChRs) can exist in high and low
sensitivity states possibly due to distinct stoichiometries during subunit
assembly: (alpha4)(2)(beta2)(3) pentamer (high sensitivity, HS) and
(alpha4)(3)(beta2)(2) pentamer (low sensitivity, LS). To determine if there is a
linkage between HS or LS states and receptor-mediated responses in brain, we
profiled several clinically studied alpha4beta2* nAChR agonists for the
displacement of radioligand binding to alpha4beta2 [(3)H]-cytisine sites in rat
brain membranes, effects on stimulation of [(3)H]-dopamine release from slices of
rat prefrontal cortex and striatum, and activation of HS and LS human alpha4beta2
nAChRs expressed in Xenopus laevis oocytes. Binding affinities (pK(i)) and
potency (pEC(50)) values for [(3)H]-dopamine release closely correlated with a
rank order: varenicline>(-)-nicotine>AZD3480>dianicline congruent with ABT-089.
Further, a good correlation was observed between [(3)H]-dopamine release and HS
alpha4beta2 pEC(50) values, but not between [(3)H]-dopamine release and LS
alpha4beta2. The relative efficacies of the agonists ranged from full to partial
agonists. Varenicline behaved as a partial agonist in stimulating [(3)H]-dopamine
release and activating both HS and LS alpha4beta2 nAChRs expressed in oocytes.
Conversely, ABT-089, AZD3480 and dianicline exhibited little efficacy at LS
alpha4beta2 (<5%), were more effective at HS alpha4beta2 nAChRs, and in
stimulating cortical and striatal [(3)H]-dopamine release >or=30%. In the
presence of alpha-conotoxin MII to block alpha6beta2* nAChRs, the alpha4beta2*
alpha-conotoxin-insensitive [(3)H]-dopamine release stimulated by these ligands
correlates well with their interactions at HS, but not LS. In summary, this study
provides support for HS alpha4beta2* nAChR involvement in neurotransmitter
release.
Complexity of conotoxins revealed by MS
Davis J, Jones A, Lewis RJ. (2009) Remarkable inter- and intra-species complexity of conotoxins revealed by LC/MS. Peptides 30:1222-1227.
The University of Queensland, Institute for Molecular Biosciences, Brisbane
Q4072, Australia.
Abstract: Cone snails have evolved an assortment of venom peptides as an evolutionary
strategy for rapid prey immobilization and defence. Earlier studies estimated
approximately 100 conopeptides per species. In this study we optimized liquid
chromatography and electrospray ionization mass spectrometry for the detection of
conopeptides in crude venom to characterize conopeptides present in the venom of
individual specimens of Conus textile, C. imperialis and C. marmoreus. Using this
approach, we have expanded the predicted number of venom peptides 10-fold to an
estimate of 1000-1900 conopeptides per species. Our investigation has also
revealed a surprisingly high level of intra-species variation that distinguishes
cone snails from other venomous species including spiders and scorpions. Given
this inherent diversity and variability, more sensitive bioassays and sequencing
techniques will be required to fully explore conotoxin bioactivity.
Complications with Ziconotide
[No Authors listed]. Ziconotide: new drug. Limited analgesic efficacy, too many adverse effects. Prescrire Int. 17:179-182.
[No authors listed]
Abstract: (1) When oral morphine does not relieve severe pain and when there is no specific
treatment for the underlying cause, the first option is to try subcutaneous or
intravenous administration. If this standard treatment fails or is poorly
tolerated, intrathecal injection is usually preferred as the direct route to the
central nervous system. However, one-quarter to one-half of patients still do not
achieve adequate pain relief, and adverse effects are relatively frequent; (2)
Ziconotide is not an opiate and is not related to the usual classes of drugs that
interfere with nervous transmission in the posterior horn of the spinal cord.
Marketing authorization has been granted for "severe, chronic pain in patients
who require intrathecal analgesia". The Summary of Product Characteristics (SPC)
recommends continuous infusion via an intrathecal catheter connected to a pump;
(3) Clinical evaluation of ziconotide does not include any trials versus morphine
in patients with nociceptive pain, or any trials versus tricyclic or
antiepileptic drugs in patients with neurogenic pain; (4) In a trial in 220
patients in whom systemic morphine had failed, the mean pain score on a 100-mm
visual analogue scale was 69.8 mm after three weeks on ziconotide, compared to
75.8 mm with placebo. This difference, although statistically significant, is
clinically irrelevant. The proportion of "responders" (reduction of at least 30%
in the initial pain score) was respectively 16.1% and 12.0% (no statistically
significant difference); (5) The two other placebo-controlled trials included 112
patients with pain linked to cancer or HIV infection, and 257 patients with
non-cancer pain. After a titration phase lasting 5 to 6 days, a combined analysis
of the two trials showed that the mean pain score was 48.8 mm with ziconotide and
68.4 mm with placebo (statistically significant difference). However, many
patients did not complete the titration phase. Efficacy also appeared to differ
according to the type of pain; ziconotide was more effective on cancer pain than
on neurogenic pain; (6) The main adverse effects of ziconotide in clinical trials
were cerebellovestibular disorders such as ataxia, dizziness, and gait disorders,
as well as confusion, hallucinations (increased in cases of overdose), nausea,
vomiting, postural hypotension, and urine retention. About 40% of patients had an
elevation in muscle creatine kinase activity, through an unknown mechanism; (7)
Intrathecal administration carries a risk of infection (especially meningitis).
Some patients might experience a paradoxical increase in pain with ziconotide;
(8) In practice, the efficacy of ziconotide in relieving neurogenic pain remains
to be established. In cancer pain, the available evidence showing that ziconotide
is effective after opiate failure is too weak in view of the potential risks. It
is better to re-examine and, if possible, correct the reasons for opiate
treatment failure rather than prescribe ziconotide.
Peptidergic toxins as tools for nicotinic receptors
Tsetlin V, Utkin Y, Kasheverov I. (2009) Polypeptide and peptide toxins, magnifying lenses for binding sites in nicotinic
acetylcholine receptors. Biochem Pharmacol. 78:720-731
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of
Sciences, Moscow 117997, Russian Federation. vits@mx.ibch.ru
Abstract: At present the cryo-electron microscopy structure at 4A resolution is known for
the Torpedo marmorata nicotinic acetylcholine receptor (nAChR), and
high-resolution X-ray structures have been recently determined for bacterial
ligand-gated ion channels which have the same type of spatial organization.
Together all these structures provide the basis for better understanding
functioning of muscle-type and neuronal nAChRs, as well as of other Cys-loop
receptors: 5HT3-, glycine-, GABA-A and some other. Detailed information about the
ligand-binding sites in nAChRs, necessary both for understanding the receptor
functioning and for rational drug design, became available when the X-ray
structures were solved for the acetylcholine-binding proteins (AChBP), excellent
models for the ligand-binding domains of all Cys-loop receptors. Of special value
in this respect are the X-ray structures of AChBP complexes with agonists and
antagonists. Among the latter are the complexes with polypeptide and peptide
antagonists, that is with protein neurotoxins from snake venoms and peptide
neurotoxins (alpha-conotoxins) from poisonous marine snails of Conus genus. The
role of a bridge between the AChBP and nAChRs is played by the X-ray structure of
the ligand-binding domain of alpha1 subunit of nAChR in the complex with
alpha-bungarotoxin. The purpose of this review is to show the role of well-known
and new polypeptide and peptide neurotoxins, from the earlier days of nAChRs
research until present time, in identification of different nAChR subtypes and
mapping their binding sites.
Intrathecal Ziconotide
Kapural L, Lokey K, Leong MS, Fiekowsky S, Stanton-Hicks M, Sapienza-Crawford AJ,
Webster LR.(2009) Intrathecal ziconotide for complex regional pain syndrome: seven case reports. Pain Pract 9:296-303
The Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA. KAPURAL@ccf.org
Abstract: Ziconotide is a nonopioid analgesic currently indicated as monotherapy, but
frequently used in combination with opioids, for the management of severe chronic
pain in patients for whom intrathecal (IT) therapy is warranted and who are
intolerant of, or whose pain is, refractory to other treatments. There is a
paucity of information regarding ziconotide use in patients with complex regional
pain syndrome (CRPS). Seven cases in which IT ziconotide was used in patients
with CRPS were analyzed. All patients (4 male, 3 female; age range, 14 to 52
years) had experienced inadequate pain relief with multiple conventional and
interventional treatments. Three patients received ziconotide monotherapy
exclusively; 4 patients received ziconotide monotherapy initially, then
combination IT therapy. The mean ziconotide dose was 5.2 mcg/d (range, 0.5 to 13
mcg/d) at initiation and 24.7 mcg/d (range, 0.06 to 146 mcg/d) at the last
available assessment. The mean duration of ziconotide therapy was 3.1 years
(range, 26 days to 8 years). At ziconotide initiation, the mean visual analog
scale (VAS) score was 89.3 mm (range, 75 to 100 mm); VAS scores decreased by a
mean of 47.5% (range, 5% to 100%) at last assessment. Of the 5 patients who
experienced substantial improvement in pain, edema, skin abnormalities, and/or
mobility with ziconotide therapy, 2 have discontinued ziconotide and are pain
free. Another patient experienced marked reversal of both edema and advanced skin
trophic changes. Adverse events included urinary retention, depression, anxiety,
and hallucinations. Adverse events generally resolved spontaneously, with
treatment, or with ziconotide discontinuation/dose reduction. Although further
studies are required, ziconotide holds promise as an effective treatment for
CRPS.
alpha6-nAChRs
Yang KC, Jin GZ, Wu J.(2009) Mysterious alpha6-containing nAChRs: function, pharmacology, and pathophysiology. Acta Pharmacol Sin. 30:740-751
Divisions of Neurology, Barrow Neurological Institute, St Joseph's Hospital and
Medical Center, Phoenix, AZ 85013, USA.
Abstract: Neuronal nicotinic acetylcholine receptors (nAChRs) are the superfamily of
ligand-gated ion channels and widely expressed throughout the central and
peripheral nervous systems. nAChRs play crucial roles in modulating a wide range
of higher cognitive functions by mediating presynaptic, postsynaptic, and
extrasynaptic signaling. Thus far, nine alpha (alpha2-alpha10) and three beta
(beta2, beta3, and beta4) subunits have been identified in the CNS, and these
subunits assemble to form a diversity of functional nAChRs. Although alpha4beta2-
and alpha7-nAChRs are the two major functional nAChR types in the CNS,
alpha6*-nAChRs are abundantly expressed in the midbrain dopaminergic (DAergic)
system, including mesocorticolimbic and nigrostriatal pathways, and particularly
present in presynaptic nerve terminals. Recently, functional and pharmacological
profiles of alpha6*-nAChRs have been assessed with the use of alpha6 subunit
blockers such as alpha-conotoxin MII and PIA, and also by using alpha6 subunit
knockout mice. By modulating DA release in the nucleus accumbens (NAc) and
modulating GABA release onto DAergic neurons in the ventral tegmental area (VTA),
alpha6*-nAChRs may play important roles in the mediation of nicotine reward and
addiction. Furthermore, alpha6*-nAChRs in the nigrostriatal DAergic system may be
promising targets for selective preventative treatment of Parkinson's disease
(PD). Thus, alpha6*-nAChRs may hold promise for future clinical treatment of
human disorders, such as nicotine addiction and PD. In this review, we mainly
focus on the recent advances in the understanding of alpha6*-nAChR function,
pharmacology and pathophysiology.
Intrathecal Ziconotide - a REVIEW
Rauck RL, Wallace MS, Burton AW, Kapural L, North JM.(2009) Intrathecal ziconotide for neuropathic pain: a review. Pain Pract.9:327-337
Wake Forest University Health Sciences, Winston-Salem, North Carolina 27103, USA. rrauck@ccrpain.com
Abstract: Neuropathic pain is a considerable burden that affects activities of daily living. The management of neuropathic pain can be challenging because of multiple etiologies and complex manifestations. Ziconotide is a nonopioid intrathecal (IT) analgesic option for patients with neuropathic pain refractory to conventional treatments. The objective of this article is to review the published literature on ziconotide for the treatment of neuropathic pain. Relevant publications were identified through searches of all years of 6 databases, which included PubMed, EMBASE, and CINAHL. Search terms used were ziconotide, SNX-111, MVIIA, Prialt, and neuropathic pain. Publications were included if ziconotide was intrathecally administered (either alone or in combination with other IT agents) to treat neuropathic pain of any etiology and if pain assessment was an outcome measure. Data extracted included study design, IT drug doses, pain outcome measures, and adverse events (AEs). Twenty-eight articles met the inclusion criteria: 5 were preclinical studies and 23 were clinical studies. In the preclinical studies, ziconotide demonstrated antiallodynic effects on neuropathic pain. Data from double-blind, placebo-controlled (DBPC) trials indicated that patients with neuropathic pain reported a mean percent improvement in pain score with ziconotide monotherapy that ranged from 15.7% to 31.6%. A low starting dose and slow titration of ziconotide resulted in an improved safety profile in the aforementioned trials. Common AEs associated with ziconotide include nausea and/or vomiting, dizziness, confusion, urinary retention, and somnolence. Evidence from DBPC trials, open-label studies, case series, and case studies suggests that ziconotide, as either monotherapy or in combination with other IT drugs, is a potential therapeutic option for patients with refractory neuropathic pain. Additional studies are needed to establish the long-term efficacy and safety of ziconotide for neuropathic pain.
Intrathecal Ziconotide in Europe
Kress HG, Simpson KH, Marchettini P, Ver Donck A, Varrassi G. (2009) Intrathecal therapy: what has changed with the introduction of ziconotide. Pain Pract. 9:338-347
Medical University of Vienna, A-1090 Vienna, Austria. hans-georg.kress@meduniwien.ac.at
Abstract: Administering drugs into the intrathecal space is becoming more popular in the treatment of patients with intractable pain or intolerable side effects of systemic analgesic treatments. Although morphine and ziconotide are the only intrathecal analgesics currently approved by regulatory authorities in the U.S. (Food and Drug Administration) and Europe (national-level approval by individual countries for morphine and European Agency for the Evaluation of Medicinal Products approval for ziconotide), a wide variety of opioid and non-opioid drugs are being used in this way. There is no official guidance concerning the selection of these drugs or their use in combinations and a paucity of efficacy and safety data from randomized controlled trials. The polyanalgesic initiative aims to summarize the current knowledge and to facilitate rational choices of intrathecal drug and drug combinations for the management of chronic pain. The most recent polyanalgesic consensus recommendations were published in 2007. In this review, we shall examine these recommendations, which are tailored toward those practicing intrathecal analgesia in the U.S., and discuss how they should be implemented in Europe, where the healthcare systems and regulations of the medical authorities are different.
Cost effectiveness of Ziconotide treatment in the UK
Dewilde S, Veridian L, Maclaine GDH (2009) Cost-effectiveness of ziconotide in intrathecal pain management for severe chronic pain patients in the UK. Curr Med Res Opin 25:2007-2019
Services in Health Economics, 1000 Brussels, Belgium. sd@SHE-consulting.be
Abstract: OBJECTIVE: To examine the cost-effectiveness of using intrathecal ziconotide in the treatment of severe chronic pain compared to best supportive care for patients with intractable chronic pain in the United Kingdom. METHODS: Using a simulation model, the analysis evaluated the cost and health economic consequences of using ziconotide as a treatment for severe chronic pain. The modelled population and clinical data were based on a randomised controlled trial in which the main outcome was reduction in pain as measured by the visual analogue scale of pain intensity (VASPI). Resource use data were elicited using a modified Delphi panel and costed using published sources. Utility values were derived from a separate research study. The main outcome measure was the cost per quality-adjusted life-year (QALY). Extensive scenario analysis was conducted to evaluate parameter uncertainty. RESULTS: Overall, findings were robust to most assumptions. The cost-effectiveness of ziconotide compared to best supportive care (BSC) was pound 27,443 per QALY (95% CI pound 18,304-38,504). Scenarios were investigated in which discount rates, the time horizon, the threshold for qualifying as a responder, pump-related assumptions, utilities, ziconotide drug dose, and the patient discontinuation rate with ziconotide were varied. The most sensitive parameter was the dosage of ziconotide: using the lower and upper bounds of the average ziconotide dosage observed in the long-term open-label study changed the incremental cost-effectiveness ratio (ICER) to pound 15,500 [pound 8206-25,405] and pound 44,700 [pound 30,541-62, 670]. CONCLUSIONS: Ziconotide may offer an economically feasible alternative solution for patients for whom current treatment is inappropriate or ineffective. The main study limitation is that some model inputs, mainly related to resource use, are based on assumptions or expert interviews.
***************************************************************************************************
More to come as now back on line and updating from 10 June 2008
***************************************************************************************************
10 June 2008
alpha-Conotoxin Lp1.1 from Conus leopardus
Peng C, Han Y, Sanders T, Chew G, Liu J, Hawrot E, Chi C, Wang C. (2008)
alpha4/7-conotoxin Lp1.1 is a novel antagonist of neuronal nicotinic
acetylcholine receptors. Peptides. 29:1700-1707.
Institute of Protein Research, Tongji University, Shanghai 200092, China.
Abstract: Cone snails comprise approximately 700 species of venomous molluscs which have evolved the ability to generate multiple toxins with varied and exquisite selectivity. alpha-Conotoxin is a powerful tool for defining the composition and function of nicotinic acetylcholine receptors which play a crucial role in excitatory neurotransmission and are important targets for drugs and insecticides. An alpha4/7 conotoxin, Lp1.1, originally identified by cDNA and genomic DNA cloning from iConus leopardus, was found devoid of the highly conserved Pro residue in the first intercysteine loop. To further study this toxin, alpha-Lp1.1 was chemically synthesized and refolded into its globular disulfide isomer. The synthetic Lp1.1 induced seizure and paralysis on freshwater goldfish and selectively reversibly inhibited ACh-evoked currents in Xenopus oocytes expressing rat alpha3beta2 and alpha6alpha3beta2 nAChRs. Comparing the distinct primary structure with other functionally related alpha-conotoxins could indicate structural features in Lp1.1 that may be associated with its unique receptor recognition profile.
Conantokin-P and -E: Novel conopeptides from Conus purpurascens and Conus ermineus
Gowd KH, Twede V, Watkins M, Krishnan KS, Teichert RW, Bulaj G, Olivera BM.(2008)
Conantokin-P, an unusual conantokin with a long disulfide loop. Toxicon 52: 203-213.
Department of Biology, University of Utah, Salt Lake City, UT 84112, USA.
Abstract: The conantokins are a family of Conus venom peptides (17-27AA) that are N-methyl-d-aspartate (NMDA) receptor antagonists. Conantokins lack disulfide bridges (six out of seven previously characterized peptides are linear), but contain multiple residues of gamma-carboxyglutamate. These post-translationally modified amino acids confer the largely helical structure of conantokins by coordinating divalent metal ions. Here, we report that a group of fish-hunting cone snails, Conus purpurascens and Conus ermineus, express a distinctive branch of the conantokin family in their venom ducts. Two novel conantokins, conantokin-P (Con-P) and conantokin-E (Con-E) are 24AA long and contain five gamma-carboxyglutamate residues. These two peptides are characterized by a long disulfide loop (12 amino acids including two Gla residues between the Cys residues). The oxidative folding studies of Con-P revealed that the formation of the disulfide bond proceeded significantly faster in the presence of Ca(++) ions. Circular dichroism suggested that Con-P is less helical than other previously characterized conantokins. Con-P blocks NMDA receptors containing NR2B subunit with submicromolar potency. Furthermore, the subtype-selectivity for different NR2 subunits differs from that of the previously characterized conantokins. Our results suggest that different branches of the phylogenetic tree of cone snails have evolved distinct groups of conantokins, each with its own unique biochemical features.
Novel conotoxin family from Conus distans
Chen P, Garrett JE, Watkins M, Olivera BM.(2008)
Purification and characterization of a novel excitatory peptide from Conus
distans venom that defines a novel gene superfamily of conotoxins.
Toxicon. 52: 139-145
Department of Biology, University of Utah, 257 South, 1400 East, Salt Lake City, UT 84112, USA. pc15@utah.edu
Abstract: An excitatory peptide, di16a, with 49 amino acids and 10 cysteine residues was purified and characterized from the venom of Conus distans. Five AA residues were modified: one gamma-carboxyglutamate (Gla), and four hydroxyproline (Hyp) residues. A cDNA clone encoding the precursor for the peptide was characterized; the peptide has a novel cysteine framework and a distinctive signal sequence that differs from any other conotoxin superfamily. The peptide was chemically synthesized and folded, and synthetic and native materials were shown to co-elute. Injection of the synthetic peptide causes a hyperexcitable phenotype in mice greater than 3 weeks of age at lower doses, and lethargy at higher doses. The peptide defines both a previously uncharacterized gene superfamily of conopeptides, and a new Cys pattern with three vicinal Cys residues.
(abstracts to follow)
Two novel conotoxins from Conus caracteristicus
Yuan DD, Liu L, Shao XX, Peng C, Chi CW, Guo ZY. (2009) New conotoxins define the novel I3-superfamily. Peptides. 30:861-865..
Institute of Protein Research, College of Life Sciences and Technology, Tongji University, Shanghai, PR China.
Abstract: We purified two novel conotoxins, designated as ca11a and ca11b, from the venom of Conus caracteristicus. Based on the amino acid sequence of mature ca11a, we cloned its full-length cDNA. Based on the signal peptide of ca11a, several ca11a-like conotoxins were cloned from C. caracteristicus and C. pulicarius. These novel conotoxins have an I-superfamily cysteine pattern but with a novel signal peptide sequence, suggesting they belong to a new branch of I-superfamily, designated as I(3)-superfamily. Additionally, two O-superfamily conotoxins were also cloned based on the signal peptide of ca11a, suggesting a possible evolutionary relationship between O- and I-superfamilies.
Novel conotoxin family from Conus caracteristicus
Yuan DD, Liu L, Shao XX, Peng C, Chi CW, Guo ZY.(2008) Isolation and cloning of a conotoxin with a novel cysteine pattern from Conus
caracteristicus. Peptides. 29:1521-1525.
Institute of Protein Research, College of Life Sciences and Technology, Tongji University, 1239 Siping Road, Shanghai 200092, PR China.
Abstract: A new conotoxin, ca16a, containing 8 cysteine residues was purified, sequenced, and cloned from a worm-hunting snail, Conus caracteristicus. This conotoxin is an extremely hydrophilic peptide comprising 34 residues, with 4 acidic and 4 basic residues. It is rich in polar Gly, Ser, and Thr residues and includes a hydroxylated Pro residue. The cysteine arrangement pattern of ca16a (-C-C-CC-C-CC-C-, designated as framework #16) is distinct from that of other known conotoxins. Furthermore, the signal peptide sequence of this conotoxin does not share any homology with those of other conotoxins. Leu residues account for almost 50% of its 20-residue signal peptide. The unique cysteine framework and signal peptide sequence of ca16a suggest that it belongs to a new conotoxin superfamily.
alphaA(S)-conotoxin analog, alphaA-OIVA[K15N], a selective high-affinity inhibitor of fetal muscle nAChRs
Teichert RW, Garcia CC, Potian JG, Schmidt JJ, Witzemann V, Olivera BM, McArdle JJ.(2008) Peptide-toxin tools for probing the expression and function of fetal and adult subtypes of the nicotinic acetylcholine receptor. Ann N Y Acad Sci. 1132:61-70.
Department of Biology, University of Utah, Salt Lake City, Utah, USA. russ_teichert@yahoo.com
Abstract: Although the neuromuscular nicotinic acetylcholine receptor (nAChR) is one of the most intensively studied ion channels in the nervous system, the differential roles of fetal and adult subtypes of the nAChR under normal and pathological conditions are still incompletely defined. Until recently, no pharmacological tools distinguished between fetal and adult subtypes. Waglerin toxins (from snake venom) and alphaA(S)-conotoxins (from cone-snail venom) have provided such tools. Because these peptides were characterized by different research groups using different methods, we have: 1) more extensively tested their subtype selectivity, and 2) begun to explore how these peptides may be used in concert to elucidate expression patterns and functions of fetal and adult nAChRs. In heterologous expression systems and native tissues, Waglerin-1 and an alphaA(S)-conotoxin analog, alphaA-OIVA[K15N], are high-affinity, highly selective inhibitors of the adult and fetal muscle nAChRs, respectively. We have used the peptides and their fluorescent derivatives to explore the expression and function of the fetal and adult nAChR subtypes. While fluorescent derivatives of these peptides indicated a gradual transition from fetal to adult muscle nAChRs in mice during the first 2 weeks postnatal, we unexpectedly observed a steeper transition in functional expression in the mouse diaphragm muscle using electrophysiology. As a toolkit of pharmacological agents with complementary specificity, alphaA-OIVA[K15N] and Waglerin-1 should have further utility in determining the roles of fetal and adult nAChR subtypes in development, in mature tissues, and under pathological conditions.
A novel conotoxin lt6c from Conus litteratus blocks Na channels
Wang L, Pi C, Liu J, Chen S, Peng C, Sun D, Zhou M, Xiang H, Ren Z, Xu A.(2008) Identification and characterization of a novel O-superfamily conotoxin from Conus litteratus. J Pept Sci. 14:1077-1083.
State Key Laboratory of Biocontrol, Guangdong Province Key Laboratory of Pharmaceutical Functional Genes, Department of Biochemistry, College of Life Sciences, Sun Yat-sen (Zhongshan) University, Guangzhou, 510275, People's Republic of China.
Abstract: A novel conotoxin named lt6c, an O-superfamily conotoxin, was identified from the cDNA library of venom duct of Conus litteratus. The full-length cDNA contains an open reading frame encoding a predicted 22-residue signal peptide, a 22-residue proregion and a mature peptide of 28 amino acids. The signal peptide sequence of lt6c is highly conserved in O-superfamily conotoxins and the mature peptide consists of six cysteines arranged in the pattern of C-C-CC-C-C that is defined the O-superfamily of conotoxins. The mature peptide fused with thioredoxin, 6-His tag, and a Factor Xa cleavage site was successfully expressed in Escherichia coli. About 12 mg lt6c was purified from 1L culture. Under whole-cell patch-clamp mode, lt6c inhibited sodium currents on adult rat dorsal root ganglion neurons. Therefore, lt6c is a novel O-superfamily conotoxin that is able to block sodium channels.
mu-conotoxins SIIIA and SIIIB from Conus striatus blocks Na channels
Schroeder CI, Ekberg J, Nielsen KJ, Adams D, Loughnan ML, Thomas L, Adams DJ, Alewood PF, Lewis RJ.(2008) Neuronally selective mu -conotoxins from Conus striatus utlise an alpha-helical motif to target mammalian sodium channels. J Biol Chem. 283:21621-21628.
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland 4072, Australia.
Abstract: Mu-conotoxins are small peptide inhibitors of muscle and neuronal tetrodotoxin (TTX)-sensitive voltage-gated sodium channels (VGSCs). Here we report the isolation of mu-conotoxins SIIIA and SIIIB by (125)I-TIIIA-guided fractionation of milked Conus striatus venom. SIIIA and SIIIB potently displaced (125)I-TIIIA from native rat brain Na(v)1.2 (IC(50) values 10 and 5 nm, respectively) and muscle Na(v)1.4 (IC(50) values 60 and 3 nm, respectively) VGSCs, and both inhibited current through Xenopus oocyte-expressed Na(v)1.2 and Na(v)1.4. An alanine scan of SIIIA-(2-20), a pyroglutamate-truncated analogue with enhanced neuronal activity, revealed residues important for affinity and selectivity. Alanine replacement of the solvent-exposed Trp-12, Arg-14, His-16, Arg-18 resulted in large reductions in SIIIA-(2-20) affinity, with His-16 replacement affecting structure. In contrast, [D15A]SIIIA-(2-20) had significantly enhanced neuronal affinity (IC(50) 0.65 nm), while the double mutant [D15A/H16R]SIIIA-(2-20) showed greatest Na(v)1.2 versus 1.4 selectivity (136-fold). (1)H NMR studies revealed that SIIIA adopted a single conformation in solution comprising a series of turns and an alpha-helical motif across residues 11-16 that is not found in larger mu-conotoxins. The structure of SIIIA provides a new structural template for the development of neuronally selective inhibitors of TTX-sensitive VGSCs based on the smaller mu-conotoxin pharmacophore.
Ziconotide - UPDATE & REVIEW
Williams JA, Day M, Heavner JE.(2008) Ziconotide: an update and review. Expert Opin Pharmacother. 9:1575-1583.
Texas Tech University Health Sciences Center, 3601 Fourth Street, MS 818, Lubbock, TX 79430, USA.
Abstract: BACKGROUND: Ziconotide is the only N-type calcium channel blocker approved by the US FDA for the treatment of chronic pain. The approved indication is for the management of severe chronic pain in patients for whom intrathecal therapy is warranted and who are intolerant of or refractory to other treatments such as systemic analgesics, adjunctive therapies or intrathecal morphine. OBJECTIVE: The purpose of this article was to review the available safety, efficacy and dosing information for ziconotide. METHODS: The sources searched for literature from 1980 to January 2008 included Pub Med, MEDLINE and PREMEDLINE using the words ziconotide, conotoxins and pain. RESULTS/CONCLUSION: Ziconotide is administered intrathecally by infusion pump to block nociceptive signal transmission in the spinal cord. It is a synthetic neuroactive peptide equivalent to the omega conotoxin MVIIA, a constituent of the venom of the fish-hunting marine snail Conus magus. It is highly potent, has a steep dose-response curve, a slow onset of action and a narrow margin of safety and responses to dose adjustments are slow. Patients receiving ziconotide should be under the care of physicians experienced in the management of intrathecal infusion therapy for pain control and should have convenient access to medical facilities.
9: Imperial JS, Chen P, Sporning A, Terlau H, Daly NL, Craik DJ, Alewood PF,
Olivera BM.(2008)
Tyrosine-rich conopeptides affect voltage-gated K+ channels.
J Biol Chem. 2008 May 27. [Epub ahead of print]
10: Duda TF Jr, Remigio EA.(2008)
Variation and evolution of toxin gene expression patterns of six closely related
venomous marine snails.
Mol Ecol. 2008 Jun;17(12):3018-32. Epub 2008 May 16.
11: Fiedler B, Zhang MM, Buczek O, Azam L, Bulaj G, Norton RS, Olivera BM,
Yoshikami D.(2008)
Specificity, affinity and efficacy of iota-conotoxin RXIA, an agonist of
voltage-gated sodium channels Na(V)1.2, 1.6 and 1.7.
Biochem Pharmacol. 2008 Jun 15;75(12):2334-44. Epub 2008 Apr 6.
12: Doura MB, Gold AB, Keller AB, Perry DC.(2008)
Adult and periadolescent rats differ in expression of nicotinic cholinergic
receptor subtypes and in the response of these subtypes to chronic nicotine
exposure.
Brain Res. 2008 Jun 18;1215:40-52. Epub 2008 Apr 4.
13: Wang Y, Shao X, Li M, Wang S, Chi C, Wang C.(2008)
mr1e, a conotoxin from Conus marmoreus with a novel disulfide pattern.
Acta Biochim Biophys Sin (Shanghai). 2008 May;40(5):391-6.
14: Dwoskin LP, Wooters TE, Sumithran SP, Siripurapu KB, Joyce BM, Lockman PR,
Manda VK, Ayers JT, Zhang Z, Deaciuc AG, McIntosh JM, Crooks PA, Bardo MT.(2008)
N,N'-Alkane-diyl-bis-3-picoliniums as Nicotinic Receptor Antagonists: Inhibition
of Nicotine-induced Dopamine Release and Hyperactivity.
J Pharmacol Exp Ther. 2008 May 6. [Epub ahead of print]
15: Arredondo J, Chernyavsky AI, Jolkovsky DL, Pinkerton KE, Grando SA.(2008)
Receptor-mediated tobacco toxicity: acceleration of sequential expression of
alpha5 and alpha7 nicotinic receptor subunits in oral keratinocytes exposed to
cigarette smoke.
FASEB J. 2008 May;22(5):1356-68.
16: Papke RL, Dwoskin LP, Crooks PA, Zheng G, Zhang Z, McIntosh JM, Stokes C.(2008)
Extending the analysis of nicotinic receptor antagonists with the study of alpha6
nicotinic receptor subunit chimeras.
Neuropharmacology. 2008 Jun;54(8):1189-200. Epub 2008 Mar 28.
6 April 2008
Conotoxins and receptors
Olivera, BM (2008) Using Conus venom peptides to understand nervous systems and discover drugs. FASEB J. 22:252.1 [Meeting Abstract]
Miller MT, Hansen,SB, McIntosh JM, Olivera BM and Taylor P (2006) Structural insights into competitive and non-competitive nicotinic antagonists. FASEB J. 20:A244 [Meeting Abstract]
25 March 2008
alpha conotoxin Vc1.1 receives US patent
US Patent NumberUS Patent Number 7,348,400 B2 “a-Conotoxin Peptides with Analgesic Properties”, Bruce Livett, Zeinab Khalil, Ken Gayler, John Down, David Sandall, David Keays. Date of Patent : March 25, 2008
27 February, 2008
US patent win for unlisted Xenome
Nick Evans BioTechnologyNews.Net Tuesday, 25 March 2008
Abstract: Xen2174 is derived from the venom of the cone snail.
The ruling sees Xenome granted the patent for the class of drugs, originally assigned to the company by the University of Queensland, and means the company’s lead pain compound, Xen2174, is cleared for development without the risk of patent infringement claims in the US.
“We recruited 37 patients into that study and we’re in the process of analysing data, and those results will be available in about the middle of the year,” he said.
Under the US patent system, “interference” is declared if two patent applications are filed which set forth claims covering the same subject matter. Under those circumstances the patent office can require that each of the parties appear to determine who was the earliest to discover the claimed invention.
Xen2174 is a synthetic drug modelled on a peptide isolated from the venom of a cone shell found on Australia's Great Barrier Reef.
Xen2174 is currently being tested in a Phase 1/2 clinical trial targeting chronic intractable pain in cancer patients via single dose intrathecal injection. This study again focuses on safety under this type of administration as well as pharmacokinetics and signs of efficacy at various doses.
18 February, 2008
Disulfide-rich peptide toxins from Gemmula species
Heralde FM 3rd, Imperial J, Bandyopadhyay PK, Olivera BM, Concepcion GP,
Santos AD (2007) A rapidly diverging superfamily of peptide toxins in venomous Gemmula species.Toxicon. 2007 Dec 28; [Epub ahead of print] doi:10.1016/j.toxicon.2007.12.022
Abstract: The gem turrids (genus Gemmula Weinkauff, 1875) are venomous snails in the family Turridae. A gene superfamily of disulfide-rich peptides expressed in Gemmula venom ducts was characterized. Gemmula speciosa (Reeve, 1843) venom duct cDNA clones revealed two different conotoxin-like prepropeptide precursors, with identical signal sequences, a largely conserved pro region, and a cysteine-rich C-terminal mature peptide region. The conserved signal sequence was used to successfully amplify homologous genes from three other Gemmula species; all had the same pattern of Cys residues in the predicted mature venom peptide. Although the signal sequence and propeptide regions were highly conserved, the mature toxin regions diverged greatly in sequence, except that the Cys residues were conserved. We designate this as the Pg-gene superfamily (Pg-superfamily) of Gemmula venom peptides. Purification of two members of the family directly from G. speciosa venom was achieved; amino acid sequence analysis revealed that these peptides are highly posttranslationally modified. With at least 10-fold as many species of turrids as cone snails, identification of rapidly diversifying gene superfamilies such as the Pg-superfamily of Gemmula is essential before the facile and systematic discovery and characterization of peptide toxins from turrid venoms can be achieved.
15 February, 2008
Venom peptides from Conus ventricosus
Romeo C, Di Francesco L, Oliverio M, Palazzo P, Massilia GR, Ascenzi P,
Polticelli F and Schininà ME (2008) Conus ventricosus venom peptides profiling by HPLC-MS: A new insight in the intraspecific variation. J Sep Sci. 31:488-498 [Epub ahead of print]
Abstract: Conus is a genus of predatory marine gastropods that poison the prey with a complex mixture of compounds active on muscle and nerve cells. An individual cone snail's venom contains a mixture of pharmacological agents, mostly short,
structurally constrained peptides. This study is focused on the composition of the venom employed by Conus ventricosus Gmelin, 1791, a worm-hunting cone snail living in the Mediterranean Sea. For this purpose, LC coupled to MS techniques has been successfully used to establish qualitative and quantitative differences in conopeptides from minute amounts of venom ducts. We were able to prove variability in the venom conopeptide complement, possibly related to different trophic habits of the species in the Mediterranean Sea. Moreover, the information-rich MS techniques enabled us to identify two novel C. ventricosus peptides, here named Conotoxin-Vn and alpha-Conotoxin-Vn. On the basis of the structural data collected so far, we suggest that Conotoxin-Vn is a conopeptide belonging to the omega-family that recognizes calcium channels through a specific pharmacophore. Similarly, molecular modeling data suggest that alpha-Conotoxin-Vn should represent a competitive antagonist of neuronal nicotinic acetylcholine receptors (nAChRs).
7 February, 2008
Biogeographical patterns in Cape Verde Conus
Cunha RL*†, Tenorio MJ‡, Afonso C†, Castilho R† and Zardoya R*. (2008) Replaying the tape: recurring biogeographical patterns in Cape Verde Conus after 12 million years. Molecular Ecology 17: 885–901
Abstract: Isolated oceanic islands are excellent natural laboratories to test the relative role of historical contingency and determinism in evolutionary diversification. Endemics of the marine venomous snail Conus in the Cape Verde archipelago were originated from at least two independent colonizations of ‘small’ and ‘large’ shelled species separated by 12 million years. In this study, we have reconstructed phylogenetic relationships within large-shelled Conus (C. ateralbus, C. pseudonivifer, C. trochulus, and C. venulatus) based on mitochondrial cox1 and nad4 haplotype sequences. The reconstructed molecular phylogeny revealed three well-supported and relatively divergent clades (A, B, and C) that do not correspond to current species classification based on shell colour and banding patterns. Clade A grouped specimens assigned either to C. pseudonivifer or C. trochulus, clade B is composed of specimens assigned to C. venulatus, and clade C comprises specimens assigned either to C. venulatus or C. ateralbus. Geometric morphometric analyses found significant differences between the radular teeth shape of C. pseudonivifer/C. trochulus and C. venulatus/C. ateralbus. In clades A and B, northwestern Boavista and Maio specimens cluster together to the exclusion of eastern Boavista samples. In Sal, populations form a monophyletic island assemblage(clade C). The large-shelled Conus have remarkably replicated biogeographical patterns of diversification of small-shelled Conus. Similar selective forces (i.e. nonplanktonic lecithotrophy with limited larval dispersal and allopatric diversification) together with repeated instances of low sea level stands during glacial maxima that allowed connection between islands, have overcome the effect of historical contingency, and explain the observed recurring biogeographical patterns.
6 February, 2008
Structure and function of α-conotoxin RgIA and analogues
Ellison M1, Feng Z-P2, Park AJ1, Zhang X2, Olivera BM1, McIntosh JM1 and Norton RS2 (2008) α-RgIA, a novel α-conotoxin that blocks the α9α10 nAChR: Structure and identification of key receptor binding residues. Journal of Molecular Biology, In Press, Accepted Manuscript, Available online 4 February 2008
Abstract: α-Conotoxins are small disulfide-constrained peptides from cone snails which act as antagonists at specific subtypes of nicotinic acetylcholine receptors (nAChRs). The 13-residue peptide α-RgIA is a member of the α-4,3 family of α-conotoxins and selectively blocks the α9α10 nAChR subtype, in contrast to another well characterized member of this family, α-ImI, which is a potent inhibitor of the α7 and α3α2 nAChR subtypes. In this study, we have altered side chains in both the 4-residue and 3-residue loops of α-RgIA, and have modified its C-terminus. The effects of these changes on activity against α9α10 and α7 nAChRs were measured, the solution structures of α-RgIA and its Y10W, D5E and P6V analogues were determined from NMR data, and resonance assignments made for α-RgIA[R9A]. The structures for α-RgIA and its three analogues were well-defined except at the chain termini. Comparison of these structures with reported structures of α-ImI reveals a common two-loop backbone architecture within the α-4,3 family, but with variations in side chain solvent accessibility and orientation. Asp5, Pro6 and Arg7 in loop 1 are critical for blockade of both the α9α10 and α7 subtypes. In loop 2, α-RgIA[Y10W] had activity near that of wild-type α-RgIA, with high potency for α9α10 and low potency for α7, and had a similar structure to wild-type. By contrast, Arg9, in loop 2, is critical for specific binding to the α9α10 subtype, probably because it is larger and more solvent accessible than Ala9 in α-ImI. Our findings contribute to a better understanding of the molecular basis for antagonism of the α9α10 nAChR subtype, which is a target for the development of analgesics for treatment of chronic neuropathic pain.
Conotoxin presentations at Society for Neuroscience meeting November 2007
At the Society for Neuroscience meeting in SanDiego, CA, November 2007 there were a number of presentations on contoxins. A selection of these Abstracts is available here.
4 February, 2008
Structure of alpha-conotoxin RgIA: Comparison with ImI and Vc1.1
Clark RJ1, Daly NL1, Halai R1, Nevin ST2, Adams DJ2, Craik DJ1 (2008). The three-dimensional structure of the analgesic α-conotoxin, RgIA. FEBS Letters (DOI: 10.1016/j.febslet.2008.01.027):Received 10 January 2008; accepted 17 January 2008. published online 31 January 2008
Abstract: The α-conotoxin RgIA is a selective antagonist of the α9α10 nicotinic acetylcholine receptor and has been shown to be a potent analgesic and reduces nerve injury associated inflammation. RgIA was chemically synthesized and found to fold into two disulfide isomers, globular and ribbon. The native globular isomer inhibited ACh-evoked currents reversibly in oocytes expressing rat α9α10 nAChRs but the ribbon isomer was inactive. We determined the three-dimensional structure of RgIA using NMR methods to assist in elucidating the molecular role of RgIA in analgesia and inflammation.
31 January, 2008
From Cone Snail to Drugs: Olivera's UP Centenial Address
Ziconotide (omega-conotoxin MVIIA): non-clinical safety
Skov MJ, Beck JC, de Kater AW, Shopp GM. (2007) Nonclinical safety of ziconotide: an intrathecal analgesic of a new pharmaceutical class. Int J Toxicol. 26: 411-421.
Abstract: Ziconotide, a potent, selective, reversible blocker of neuronal N-type voltage-sensitive calcium channels, is approved in the United States for the
management of severe chronic pain in patients for whom intrathecal therapy is warranted, and who are intolerant or refractory to other treatment, such as systemic analgesics, adjunctive therapies, or intrathecal morphine. In the European Union, ziconotide is indicated for the treatment of severe chronic pain in patients who require intrathecal analgesia. Nonclinical investigations of ziconotide included a comprehensive characterization of its toxicology, incorporating acute and subchronic toxicity studies in rats, dogs, and monkeys; reproductive toxicity assessments in rats and rabbits; and mutagenic, carcinogenic evaluations performed in vivo and in vitro. Additional investigations assessed the potential for cardiotoxicity (rats) and immunogenicity (mice, rats, and guinea pigs), and the presence or absence of intraspinal granuloma formation and local cell proliferation and apoptosis (dogs). The resulting nonclinical toxicology profile was predictive of human adverse events reported in clinical trials and consistent with ziconotide's pharmacological activity. Frequently observed nonclinical behavioral effects included tremoring, shaking, ataxia, and hyperreactivity. Occurrences were generally transient and reversible upon cessation of treatment, and intolerable effects occurred at doses more than 45 times the maximum recommended clinical dose. Ziconotide was not associated with target organ toxicity, teratogenicity, or treatment-related gross or histopathological changes; it displayed no mutagenic or carcinogenic potential and no propensity to induce local cell proliferation or apoptosis. Although guinea pigs developed systemic anaphylaxis, antibodies to ziconotide were not detected in mice, rats, or guinea pigs, indicating low immunogenic potential. No evidence of granuloma formation was observed with intrathecal ziconotide treatment. In summary, the results from these nonclinical safety assessments revealed no significant toxicological risk to humans treated with ziconotide as recommended.
29 January, 2008
Conotoxin transcripts in Conus leopardus
Remigio, EA and Duda TF Jr (2008) Evolution of ecological specialization and venom of a predatory marine gastropod Molecular Ecology, OnlineEarly ArticlesPublished article online:
23-Jan-2008 doi: 10.1111/j.1365-294X.2007.03627.x
Abstract: Understanding the evolution of ecological specialization is important for making inferences about the origins of biodiversity. Members of the predatory, marine gastropod genus Conus exhibit a variety of diets and the ability to capture prey is linked to a venom comprised of peptide neurotoxins, termed conotoxins. We identified conotoxin transcripts from Conus leopardus, a species of Conus that uniquely preys exclusively on hemichordates, and compared its venom duct transcriptome to that of four other Conus species to determine whether a shift to a specialized diet is associated with changes in the venom composition of this species. We also examined the secondary structure of predicted amino acid sequences of conotoxin transcripts of C. leopardus to identify substitutions that may be linked to specialization on hemichordates. We identified seven distinct conotoxin sequences from C. leopardus that appear to represent transcripts of seven distinct loci. Expression levels and the diversity of conotoxins expressed by C. leopardus are considerably less than those of other Conus. Moreover, gene products of two transcripts exhibited unique secondary structures that have not been previously observed from other Conus. These results suggest that transition to a specialist diet is associated with reduction in the number of components expressed in venoms of Conus and that diverse venoms of Conus are maintained in species with a broad dietary width.
25 January, 2008
Conotoxins from Conus austini
Zugasti-Cruz A, Aguilar MB, Falcón A, Olivera BM, Heimer de la Cotera EP (2008). Two new 4-Cys conotoxins (framework 14) of the vermivorous snail Conus austini from the Gulf of Mexico with activity in the central nervous system of mice. Peptides. 2007 Dec 5; [Epub ahead of print]
Abstract: As part of continuing studies of the venom components present in Conus austini (syn.: Conus cancellatus), a vermivorous cone snail collected in the western Gulf of Mexico, Mexico, two major peptides, as14a and as14b, were purified and characterized. Their amino acid sequences were determined by automatic Edman sequencing after reduction and alkylation. Their molecular masses, established by matrix-assisted laser desorption ionization time-of-flight mass spectrometry, confirmed the chemical analyses and indicated that as14a and as14b have free C-termini. Each peptide contains 4-Cys residues arranged in a pattern (C-C-C-C, framework 14). The primary structure of as14a is GGVGRCIYNCMNSGGGLNFIQCKTMCY (experimental monoisotopic mass 2883.92Da; calculated monoisotopic mass 2884.20Da), whereas that of as14b is RWDVDQCIYYCLNGVVGYSYTECQTMCT (experimental monoisotopic mass 3308.63Da; calculated monoisotopic mass 3308.34Da). Both purified peptides elicited scratching and grooming activity in mice, and as14b also caused body and rear limb extension and tail curling immediately upon injection. The high sequence similarity of peptide as14a with peptide vil14a from the vermivorous C. villepinii suggests that the former might block K(+) channels.
3-D structure of conotoxin tx3a from Conus textile
McDougal OM, Turner MW, Ormond AJ, Poulter CD (2008). Three-Dimensional Structure of Conotoxin tx3a: An m-1 Branch Peptide of the M-Superfamily. Biochemistry. 2008 Jan 19; [Epub ahead of print]
Abstract: The M-superfamily, one of eight major conotoxin superfamilies found in the venom of the cone snail, contains a Cys framework with disulfide-linked loops labeled 1, 2, and 3 (-CC (1) C (2) C (3) CC-). M-Superfamily conotoxins can be divided into the m-1, -2, -3, and -4 branches, based upon the number of residues located in the third Cys loop between the fourth and fifth Cys residues. Here we provide a three-dimensional solution structure for the m-1 conotoxin tx3a found in the venom of Conus textile. The 15-amino acid peptide, CCSWDVCDHPSCTCC, has disulfide bonds between Cys (1) and Cys (14), Cys (2) and Cys (12), and Cys (7) and Cys (15) typical of the C1-C5, C2-C4, and C3-C6 connectivity pattern seen in m-1 branch peptides. The tertiary structure of tx3a was determined by two-dimensional (1)H NMR in combination with the combined assignment and dynamics algorithm for nuclear magnetic resonance (NMR) applications CYANA program. Input for structure calculations consisted of 62 inter- and intraproton, five phi angle, and four hydrogen bond constraints. The root-mean-square deviation values for the 20 final structures are 0.32 +/- 0.07 and 0.84 +/- 0.11 A for the backbone and heavy atoms, respectively. Surprisingly, the structure of tx3a has a "triple-turn" motif seen in the m-2 branch conotoxin mr3a, which is absent in mr3e, the only other member of the m-1 branch of the M-superfamily whose structure is known. Interestingly, injection of tx3a into mice elicits an excitatory response similar to that of the m-2 branch peptide mr3a, even though the conotoxins have different disulfide connectivity patterns.
20 January, 2008
Conorfamide-Sr2 from Conus spurius
Aguilar MB, Luna-Ramírez KS, Echeverría D, Falcón A, Olivera BM, Heimer de la
Cotera EP, Maillo M. (2008) Conorfamide-Sr2, a gamma-carboxyglutamate-containing FMRFamide-related peptide from the venom of Conus spurius with activity in mice and mollusks.
Peptides. 2007 Dec 5; [Epub ahead of print]
Abstract: A novel peptide, conorfamide-Sr2 (CNF-Sr2), was purified from the venom extract of Conus spurius, collected in the Caribbean Sea off the Yucatan Peninsula. Its primary structure was determined by automated Edman degradation and amino acid analysis, and confirmed by electrospray ionization mass spectrometry. Conorfamide-Sr2 contains 12 amino acids and no Cys residues, and it is only the second FMRFamide-related peptide isolated from a venom. Its primary structure PMgammaDPLgammaIIRI-nh(2), (gamma, gamma-carboxyglutamate; -nh(2), amidated C-terminus; calculated monoisotopic mass, 1468.72Da; experimental monoisotopic mass, 1468.70Da) shows two features that are unusual among FMRFamide-related peptides (FaRPs, also known as RFamide peptides), namely the novel presence of gamma-carboxyglutamate, and a rather uncommon C-terminal residue, Ile. CNF-Sr2 exhibits paralytic activity in the limpet Patella opea and causes hyperactivity in the freshwater snail Pomacea paludosa and in the mouse. The sequence similarities of CNF-Sr2 with FaRPs from marine and freshwater mollusks and mice might explain its biological effects in these organisms. It also resembles FaRPs from polychaetes (the prey of C. spurius), which suggests a natural biological role. Based on these similarities, CNF-Sr2 might interact with receptors of these three distinct types of FaRPs, G-protein-coupled receptors, Na(+) channels activated by FMRFamide (FaNaCs), and acid-sensing ion channels (ASICs). The biological activities of CNF-Sr2 in mollusks and mice make it a potential tool to study molecular targets in these and other organisms.
17 January, 2008
Do hydroxyprolines determine the folding and activity of conotoxins ?
Abstract: Hydroxylation of proline residue occurs in specific peptides and proteins derived from plants and animals, but the functional role of this modification has been characterized primarily in collagen. Marine cone snails produce disulfide-rich peptides that have undergone a plethora of posttranslational modifications, including proline hydroxylation. Although Conus snails extensively utilize proline hydroxylation, the consequences of this modification remain largely unexplored. In this work, we investigated the function of 4-hydroxyproline (Hyp) in conotoxins from three distinct gene families: mu-, omega-, and alpha-conotoxins. Analogues of mu-GIIIA, omega-MVIIC, alpha-GI, and alpha-ImI were synthesized with either Pro or Hyp, and their in vitro oxidative folding and biological activity were characterized. For GIIIA, which naturally contains three Hyp residues, the modifications improved the ability to block NaV1.4 sodium channels but did not affect folding. In contrast, the presence of Hyp in MVIIC had a significant impact on the oxidative folding but not on the biological activity. The folding yields for the MVIIC[Pro7Hyp] analogue were approximately 2-fold higher than for MVIIC under a variety of optimized oxidation conditions. For alpha-conotoxins ImI and GI, the hydroxylation of the conserved Pro residue improved their folding but impaired their activities against target receptors. Since prolyl-4-hydroxylase and protein disulfide isomerase coexist as a heterotetramer in the ER, we discuss the effects of Hyp on the folding of conotoxins in the context of cis-trans isomerization of Pro and Hyp. Taken together, our data suggest that proline hydroxylation is important for both in vitro oxidative folding and the bioactivity of conotoxins.
[The following hydroxyproline-containing conotoxins are included: mu-conotoxins GIIA, GIIIB, GIIIC, PIIIA; kM-conotoxin RIIK; psi-conotoxin PIIIE; alpha-conotoxin (4/7) EI; alpha-conotoxin (4/3) Reg1b; delta-conotoxin PVIA, SVIE; alphaA-conotoxins PIVA, EIVA; omega-conotoxin GVIA; chi-conotoxin MrIA, MrIB; and contryphans R, and Tx]
Two Cone Shell Articles
New Species of Cone Shells
One new species: Conus beatrix Tenorio, Poppe & Tagaro, 2007, collected on Aliguay Is., P.I. by Manuel J. Tenorio and Paul Kersten, and one new subspecies Conus recluzianus simanoki Tenorio, Poppe & Tagaro, 2007 collected at the Burma / Thailand border by Philippe Quisquandon, were published in Visaya, vol. 2, No. 2 (November, 2007).
The Cone Collector Volume 5 January 2008
Edited by António Monteiro, with layout by André Poremski.
5 January, 2008
Folding of conotoxins
Bulaj G, Olivera BM. (2008) Folding of conotoxins: formation of the native disulfide bridges during chemical synthesis and biosynthesis of Conus peptides. Antioxid Redox Signal. 10:141-56.
Abstract: Conopeptides from >700 species of predatory marine Conus snails provide an impressive molecular diversity of cysteine-rich peptides. Most of the estimated 50,000-100,000 distinct conopeptides range in size from 10 to 50 amino acid residues, often with multiple posttranslational modifications. The great majority contain from two to four disulfide bridges. As the biosynthetic and chemical production of this impressive repertoire of disulfide-rich peptides has been investigated, particularly the formation of native disulfide bridges, differences between in vivo and in vitro oxidative folding have become increasingly evident. In this article, we provide an overview of the molecular diversity of conotoxins with an emphasis on the cysteine patterns and disulfide frameworks. The conotoxin folding studies reviewed include regioselective and direct oxidation strategies, recombinant expression, optimization of folding methods, mechanisms of in vitro folding, and preliminary data on the biosynthesis of conotoxins in venom ducts. Despite these studies, how the cone snails efficiently produce properly folded conotoxins remains unanswered. As chemists continue to master oxidative folding techniques, insights gleaned from how conotoxins are folded in vivo will likely lead to the development of the new folding methods, as well as shed some light on fundamental mechanisms relevant to the protein folding problem.
1 January, 2008
Welcome to 2008
Continued in What's new in 2007
[HOME] |
On January 16, 2008, Prof. Baldomero M. Olivera PhD received an honorary degree and presented this Centenial address to assembled staff and graduates at the University of the Philippines: 
