Journal of Chemical Ecology, Vol. 19, No. 10, 1993

Review

NEUROTOXIC A C Y L P O L Y A M I N E S FROM SPIDER VENOMS 1

KEVIN D. M c C O R M I C K and J E R R O L D M E I N W A L D * Cornell Institute for Research in Chemical Ecology (CIRCE) Department of Chemistry, Cornell University Ithaca, New York 14853-1301 (Received March 22, 1993; accepted May 5, 1993)

Key Words--Spiders, Arachnida, venoms, neuroactive components, acylpolyamines, synthesis. INTRODUCTION In their relations with humans, spiders have acquired considerable respect, often inspiring emotions ranging from mild discomfort to genuine terror. 1-3 One of the few literary instances in which spiders are treated realistically but affectionately is E.B. White's classic, "Charlotte's Web. ''4 It does not appear entirely unreasonable to attribute our tendency to avoid contact with spiders, at least in part, to the ability of some species to produce and deliver venoms that paralyze and/or kill their prey. 5 The capacity to intoxicate invertebrate and vertebrate prey is clearly an extremely important factor in the chemical ecology of this fascinating group of arachnids. It is also the reason for the recent wave of "arachnophilia" which has manifested itself in the chemical and pharmacological literature of the past decade, coinciding with the realization that spider venoms contain many neuroactive components, of relatively low ( < 1000 ainu) molecular weight, which may have important uses. Our own entry into this field * To whom correspondence should be addressed. The format of citations and references is nonstandard by agreement with the authors.

2411 0098-0331/93/1000-2411$07.00/0 9 1993PlenumPublishingCorporation

2412

McCORMICKAND MEINWALD

has necessitated a critical examination of the relevant chemical literature. 6 It is our purpose here to present an overview of recent research on spider-derived neurotoxins, designating all those species that have been studied chemically, describing briefly the types of neurochemical activity associated with the various toxins, listing the chemical structures that have been determined to date, and outlining the syntheses that have been accomplished. Venomous animals from a wide variety of taxa have evolved powerful arsenals stocked with chemical weapons able to stun, paralyze, and kill other organisms. Many venom components function as neurotoxins, which often exhibit remarkable specificities and binding affinities for particular neural receptors or ion channels. For example, toxins from snakes, scorpions, and predatory snails have been found to block specifically certain ligand-dependent 7'8 and voltage-dependent ion channels. 9-~2 Consequently, these compounds are proving to be useful tools for neurochemical research. There are additional reasons why neurotoxins are important. Many neural diseases and disorders are based on perturbations in the functioning of neural receptors and ion channels. As an example, nerve cell death in ischemia ~3' ~4 (associated with strokes and heart attacks) and epilepsy 15"16 is often caused by an uncontrolled firing of the N-methyl-o-aspartate (NMDA) receptors, which allows toxic levels of calcium to enter the cells. Knowledge of the structures of venomous compounds that block these responses and of their mechanism(s) of action can be expected to aid in the development of drugs with which to treat these neurological problems. Spiders are now receiving considerable attention in this context. Spiders are completely carnivorous, and many depend on their venom for survival. They are abundant throughout the world; more than 30,000 species from over 70 different families had been described by the early 1980s. ~7 Until recently, however, spiders have been largely neglected by chemists because of the small size of most arachnids, coupled with the difficulty in collecting their venom. Black widow spiders (Latrodectus mectans), for example, are only 6-11 mm long, and contain about 0.2 mg of venom per individual. 18 Nevertheless, it is now possible to collect spider venom nondestructively, 19'2~ and new methods are available to isolate, bioassay, and characterize venom components in very small amounts. These advances have facilitated the study of many new species.

NEUROTOXINS FROM SPIDERS Research into the detailed chemistry of spider venoms began over 30 years ago when Fischer and Bohn analyzed the venoms of some very large South American bird spiders. 2~ At that time, full characterization of the venom constituents was not possible. However, the venoms were shown to contain both

SPIDERVENUMS

2413

proteins and polyamines. Since this initial study, venom components from over 85 spider species representing 15 different families have been investigated (Table 1), although fewer than a dozen of these species have been studied in depth. The spider venoms are composed chiefly of proteins and polypeptides, but they also contain low-molecular-weight ( < 1000 amu) organic compounds and inorganic salts. 22 Most recent research has been devoted to neurotoxic substances, although some attention has also been directed toward venom-derived enzymes. The venom of the aggressive brown recluse spider (Loxosceles reclusa), for example, contains a powerful sphingomyelinase capable of extensive tissue damageY Categorizing the different types of spider neurotoxins on the basis of their biological activity is difficult. The nervous system is complex, and determining how individual toxins act requires a variety of bioassays to probe these complexities. For example, black widow spider venom contains two large (130 and 150 kDa) neurotoxic proteins. One of these, a-latrotoxin, is very toxic to vertebrate animals but not to insects. 24 The other, a-latroinsectotoxin, is active against insects but not vertebratesY Interestingly, both neurotoxins function by a similar mechanism. They bind to recognition sites along the presynaptic membrane, and then form transmembrane Ca 2+ ion channels stimulating a massive release of neurotransmitter. 26'z7 The different target selectivities seem to be due to recognition of different binding sites along the presynaptic membrane. Other than that, both neurotoxins function identically and have a high degree of sequence homology. 24'27 Many of the earlier bioassays determined only whether a compound was neurotoxic to some animal species. Few mechanistic studies were done, and consequently the mode of action of many neurotoxins remained unknown. However, as knowledge of the nervous system has improved, so have the bioassays. The precise mechanisms by which some neurotoxins function are now known, and spider toxins are revealing new information about neural receptors and ion channels. For example, FTX, an intriguing venom component of as yet undetermined structure or homogeneity isolated from Agelenopsis aperta, was found to block an unknown type of calcium current in Purkinje cells; this study led to the discovery of the P-type calcium channel. 166"171,174

COMPILATION OF STRUCTURES Most neurotoxins identified so far are proteins or polypeptides, of which four different classes are known: (1) high-molecular-weight proteins that create divalent cation channels which cause massive transmitter release from presynaptic nerve terminals, (2) small polypeptides that function similarly to the first class, (3) polypeptide blockers of presynaptic calcium channels that inhibit signal

2414

McCoRMICK AND MEINWALD

TABLE 1. SPIDER TAXONOMY, INDICATING SPECIES WHOSE VENOMS HAVE BEEN AT LEAST PARTIALLY CHARACTERIZED Araneae Orthognatha Mesothelae (atypical tarantulas) Liphistiidae Anthrodiaetidae Mecicobothriidae Atypidae Opisthothelae (typical tarantulas) Themphosidae Acanthoscurria atrox 21 Acanthoscurria cratus 28 Acanthoscurria emelia 29 Acanthoscurria sp. (Arizona) a9 31 Acanthoscurria sp. (Honduras) 29 Aphonopelma chalcodes 3z Brachypelma smithii 33 Dugesiella hentzi29"3~ 33 36 Eurypelma californicum 37"38 Eurypelma emilia 33 Eurypelma vellutinum 21 Grammostola moUicoma 2t Grammostola actaeon 2~ Grammostola pulchripes 21 Harpactirella sp. 32 Lasiodora klugii 21 Pamphobeteus platyomma 39 Pamphobeteus roseus 21"a~ Pamphobeteus soracabae 39 Pamphobeteus tetracanthus 2~ Phormictopus cancerises 41 Pterinochilus sp. 42'4~

Paratropididae Pycnothelidae Barychelidae Migidae Dipluridae (funnel-web spiders) Atrax infensus 44 h t r a x robustus 45-54 Atrax versutus 55

Ctenizidae (trap-door spiders) Aganippe berlandi 44 Aptostichus schlingeri 56 Hebestatis theveniti 32 Phoneutria nigriventer 57-66 Phoneutria fero 67' 68

Actinopodidae

Labidognatha Hypochiloidea Gradungulidae Hypochilidae Neocribetlatae Filistatidae Oecobiidae Eresidae Dinopidae Uloboridae Dictynidae Amaurobiidae Amphinectidae Neolanidae Psechridae Stiphiidae Tengellidae Zoropsidae Acanthoctenidae Ecribellatae Haplogynae (primitive hunters and weavers) Sicariidae Scytodidae Loxoscelidae Loxosceles laela 69"7~ Loxosceles gaucho 71 Loxosceles reclusa-brown

recluse23,72 95 Loxosceles refescens 96"97 Loxosceles reufipes 96

Diguetidae Plectreuridae Plectreurys tristes 4t'98 lot

Caponiidae Oonopidae Tetrablemmidae Pacullidae Ochyroceratidae Leptonetidae Telemidae Textracellidae Dysderidae Segestridae Segestrie florentina i02,103

TABLE l .

Entelogynae Trionycha (higher web weavers) Pholicidae Symphytognathidae Amaurobius insignis 44

Theridiidae (comb-footed) spiders Achaearanea tepidariorum 1~ to5 Episinus ?106 Latrodeetus hasseltt~4 Latrodectus hesperus ~~ Latrodectus mectans--black widowIS,20.24-27,68,106,108 135 Latrodectus pallidus 1~ 186 Latrodectus dahli '36 Steatoda bipunctata 1~ Steatoda paykulliana ~~ 126,~37,138 Steatoda triangulosa 1~ Theridion impressum 1~ Theridion varians ~~

Nicodamidae Nesticidae Hadrotarsidae Linyphiidae Micryphantidae Theridiosomatidae Araneidae (typical orb weavers) Agalenatea redii 139 Araneus diadematus 139 Araneus gemma 41"140-142 Araneus Argiope Argiope Argiope

tartaricus ~48 auranta 4~' ~4z,~45 bruennichi 97 lobata 139"t43,146-149

Argiope florida 14~ 142 Argiope trifasciata~4~ ~a2 Mangora acalypha 139 Neosocona adianta ~39 Neosocona arabesca 4~' ~45,~50 Neoscona cruciferoides 139 geoscona nautica 97 Nephila clavats 97" 151-159 Nephila edulis 44 Nephila maculata '52" 154,155 Nuctenea folium139 Zygiella caspica 139

Tetragnathidae Agelenidae (funnel-web weavers) Agelena opulenta '6~ Agelenopsis aperta 16H78 Hololena curta 1~ 179-181 Tegenaria domestica ~82

CONTINUED

Argronetidae Sesidae Hahniidae Hersiliidae Urocteidae Mimetidae Archaeidae Mecysmaucheniidae Zodariidae Palpimanidae Stenochilidae Pisauridae (nursery-web spiders) Dolomedes okefinokensis 6"188 Dolomedes sulfureus 97

Lycossidae Lycosa erythrognatha 57"67 Lycosa godeffroyi 44" 184 Lycosa labrea 41 Lycosa singoriensis 185 187 Lycosa tarentula 188 Sosippus californicus 189

Oxyopidae Peucetia viridens 4 ~"145 Senoculidae Toxopidae Dionycha (two clawed hunting spiders) Ammoxenidae Platoridae Gnaphosidae Lampona cylindrate 44

Prodidomidae Homalonychidae Cithaeronidae Clubionidae Anyphaenidae Amaurobioidae Zoridae Ctenidae Sparassidae Isopeda immanis 44

Thomisidae Philodroimidae Aphantochilidae Salticidae Lyssomanidae Unknown family Chiranthium japonicum 97 Eriophora transmarina 44 Namea salantri 44 Scodra griseipes 19~ 191

2416

McCORMICKAND MEINWALD

transmission, and (4) polypeptide activators of presynaptic sodium channels that stimulate repetitive nerve firings. Examples of these classes are given in Table 2. Spider venoms also contain many compounds other than proteins and polypeptides that are neuroactive. Some of these compounds are well known from other sources as neural transmitters (serotonin, 2~ octopamine, 51 5-hydroxytryptamine, 68"114 5-methoxytryptamine, 51 histamine, 61"6s tyramine, 51 y-aminobutyric acid, 20~21"34'47'51'68 aspartic acid, 2~ and glutamic acid2~ Only one new, nonproteinaceous class of neuroactive compounds has been discovered in spider venoms, the acylpolyamines. These all contain an aromatic acyl end group and a polyamine chain. They were first discovered in the early to mid1980s from the spiders Nephila clavata 152 and Argiope lobata. 146 These neurotoxins function by blocking glutamate-sensitive calcium channels that are important in many neural functions including pain, 19z motor control, 193 and memory. 194 The glutamate-sensitive calcium channels have also been implicated in several neurodegenerative disorders such as amyotrophic lateral sclerosis 195 and Alzheimer's, and Huntington's diseases. 196 The desire to understand how these ion channels function has stimulated much research on spider venoms as well as on acylpolyamines from other sources. Many new acylpolyamines have now been identified in spider venoms, and quite remarkably, some closely related acylpolyamines have also been identified in certain wasp venoms. ~98'199 Currently, the neuronal activity reported for all but two of these acylpolyamines is glutamate-receptor blocking. Since glutamate is the active chemical messenger in insect neuromuscular junctions, this mode of action is hardly surprising (a recent review of the activities of these glutamate receptor affecting toxins has been published by P.N.R. Usherwood and I.S. Blagbrough)fl~176 The exceptions are CNS 2103 (as its trifluroacetic acid salt), which blocks L- and R-type voltage-sensitive calcium channels, 183 and FTX, which blocks P-type voltage-sensitive calcium channels. Additionally, one group of acylpolyamines, the nephilatoxins, has also been shown to degranulate mast cells, 2~ although the biological significance of this observation is not clear. The acylpolyamines can be subdivided into two groups: the amino acidcontaining acylpolyamines (Figure 1) and the non-amino acid-containing acytpolyamines (Figure 2). Both types function similarly as glutamate receptor blockers, and both incorporate the same family of aromatic end groups (Figure 3). The amino acid-containing acylpolyamines were the first to be discovered. To date, 34 of these compounds have been fully characterized, all from one spider family: the Araneidae (orb weavers) (Table 3). All of these possess one or two of the following basic amino acids between the acyl end group and the polyamine chain: asparagine, ornithine, and c0-N-methyllysine. In addition, some of these acylpolyamines possess up to three amino acids at the polyamine terminus, with arginine usually occupying the terminal position.

TABLE 2. EXAMPLES OF FOUR CLASSES OF POLYPEPTIDE NEUROTOXINS IDENTIFIED IN SPIDER VENOMS

Type I. Large proteins that bind to presynaptic membranes and form transmembrane, divalent cation channels, producing an influx of calcium ions that stimulates massive neurotransmitter release.

Lactrodectus mectans e~-Latrotoxin

130 kDa Forms channels in vertebrates but not 1170 amino acid sequence invertebrates 24,26,116-118 e~-Latroinsectotoxin 120 kDa Forms channels in invertebrates but not partial amino acid sequence vertebratesY.27, ~35 Steatodapaykulliana 110-130 kDa Forms ion channels, toxic to insectsJ 37 Type II. Low molecular weight pore forming polypeptides which cause massive transmitter release from nerve terminals. Latrodectus mectans, 5 kDa Forms ion channels through L. pallidus, Steatoda phospholipid membranes 1~ ~26,~34,197

bipunctata, S. paykulliana, S. triangulosa, Theridion impressum Sosippus californicus Sositoxin I

2.5 kDa Forms ion channels through 23 amino acid sequence phospholipid membranes ~89 Type III. Peptides that suppress neurotransmitter release from presynaptic stores. These function by blocking presynaptic ca2+ channels.

Agelenopsis aperta ~-Aga-IA w-Aga-IB w-Aga-llIA w-Aga-IIA w-Aga-IVA

7.5 kDa (66 aa) 66 amino acid sequence 7.5 kDa partial sequence 8.5 kDa 11 kDa partial sequence 5.2 kDa 48 amino acid sequence

Blocks L- and N-type Ca z+ channels. ~65.~68.~73

16 kDa two subunits 4 kDa 36-38 amino acid sequence

Blocks voltage dependent Ca 2+ channels vertebrate inactive t~ Irreversible presynaptic neuromuscular blockage in insects ~8~

Blocks L- and N-type Ca 2+ channels, t65. L68 Blocks N- and L-type Ca 2+ channels.~74 Blocks N-type Ca 2+ channels, inhibits c~-CgTx binding. 168 Blocks P-type Ca 2+ channels. 177

Hololena curta HoTX CT-I, II, and III

Plectreurys tristes blocks Ca 2§ channels vertebrate inactive99. ~0 Type IV. Polypeptides that cause repetitive firing in presynaptic neurons. This action is caused by activation of presynaptic voltage-sensitive sodium channels. a-PLTX I-III

7 kDa

Agelenopsis aperta tz-Aga I-VI

4.2 kDa 36-38 amino acid sequences

External sodium required for activation. Excitation is abolished with the addition of tetrodotoxin. ~64,167

8 kDa 77 amino acid sequence 6-6.5kDa partial amino acid sequence

Excitation is abolished with the addition of tetrodotoxin. 63,65,66

Phoneu~ia nignventer Phxl PhTx2

Excitation is abolished with the addition of tetrodotoxin. 66

2418

McCORMICK AND MEINWALD

NSTX-3 OH

! !

M

H

0

0

H

I

ii

II

I

II

~

S

i

H

~

I

H

H

H

~ O

I

II

il

NH

JSTX- i

JSTX-2

.o/~.;~

o

i

H

o

L

o

.I 2

JSTX-3 OH

0

H

O

"~~

"

H "~eos~

o

H

H

H

JSTX-4 OH

o

H

Ho~

o

H

~

H

~cos~ Nit2

NPTX-1 H

I

II

D

II

O

-~ "

O

I

H

I

H

H

I

H

i H NFFX-2

FIo. 1. Chemical structures of all the known amino acid-containing acylpolyamines identified from spider venoms.

SPIDER VENUMS

2419

NF~-3

NPTX-4 .

o

,'I

,m_~. .

Fil

H

l

H

NPTX-5

7

~/~i

o

r o

~

1

L.

.

J~

H

NPTX-6

p I

II

II

II

r

o

"

/

o

1

I /

I

~

NPTX-7 a

o

I

fl

.I"

/

.

F.

.

q

I

|1

I

/

NP'rX-8 H

0

0

H

I l

m

H

U

I ~

I

I

i

H

FIG. 1. Continued

I

2420

McCORMICK

AND MEINWALD

o

~.

bIPTX-9

~

~

i~

NPTX-IO

~..~..~

o

~-~o~

.

.

6 NPTX-II H

O

O

I

NPTX-12

[ H

Clavamine oH

o

.

o

H

a

~

H

N HO

Argiopine or Argiotoxin-636 OH

H

NH

Argiopinin I

~

H

0

I

.:." ~ ~oor~

6

O

II

uC

H I .

""\/

Ot

'

It

~

I

-~ Nrq

I II o

It

I II Na

Argiopinin II 14

H

?" I

E U

II -~

~

O

~ --"

I coNrb

II I

CI'I 3

II

,

,

=

H

FIG. 1. Continued

w II

U

SPIDER VENUMS

2421

Argiopinin III or Argiotoxin-659

I H

Argiopinin IV

Argiopinin V

NHC~

Pseudoargiopinin I

I

H

Pseudoargiopinin II

I

H

Pseudoargiopinin III H

O

Argiotoxin-480 OH

H

O

FIG. 1. Continued

2422

McCORMICK AND MEINWALD

Argiotoxin*650

Argiotoxin-494

Argiotoxin-622

Ho

h~

Argiotoxin-503 OH

~

O

~

o

~,

H

O

(~I~

I

H

Argiotoxin-673

o.

1

tt

Argioto• OH

I

I

I

H

Argiomxin-645

I

N

FIG. 1. Continued

y

.~

AG 452 or AGEL 452 HO

H

~ 1

OH

H

I

H

I

I

AG 489, AGEL 489 or HO 489

H

AG 505, AGEL 505 or HO 505

It It

AG 504 or AGEL 505a OH

H

I

H

I

~

H

H

1

I

H

AG488 or AGEL489a

H

AGEL468 or HO468 OH

OH

O

H

AGEL 448 OH

H

N

OH

~

N

H

~

'

~

N

~

~

NI~

H H

HO 4"]3 H

H

H

H

I

i

i

i tt

It

FIG. 2. Chemical structures of all known non-amino acid-containing acylpolyamines from spider venoms.

HO 452 OH

OH

O

H

HO 448 (same as Age1448?) OH

H

I

H

H

+ oxygen

H

HO 395 tt

I

H

H

H

H

HO 416a

H H

HO 416b OH

HO 359 H

I H

H

I H

Het 389 N~Sr~

li

Het 403 O

I

H

FIG. 2. Continued

SPIDER VENUMS

2425

Ape600

H

Ape728 H

H

H

CNS2103 H

H

H

1t

FI6. 2. Continued

OH H

OH

~

C~

~

H ~

C~

C02H

OH

I

I

H

H

OH CO2H

H

O

~

C02H

L L H H Fie. 3. The sevenacidscorrespondingto the acylend groupsidentifiedin spidervenom acylpolyamines.

JSTX-1 JSTX-2 JSTX-3 JSTX-4 NPTX-I NPTX-2 NPTX-3 NPTX-4 NPTX-5 NPTX-6 NPTX-7 NPTX-8 NPTX-9 NPTX- 10 NPTX- 11 NPTX- 12

NSTX-3

Compound

152, 155

Nephila maculata

158 152, 152, 152. 157 157 157 157 157 157 156, 156, 156, 157 157 157 157 157 157

158 153 158

Nephila clavata

Argiope lobata

Argiope trifasciata

Argiope florida

Argiope auranta

TABLE 3. AMINO ACID-CONTAINING ACYLPOLYAMINES FROM SPIDERS

Araneus gemma

207 208 207 208

203,206

202, 203, 204,205

Synthesis

~7

z

9

O',

b.~

~Structures are only partially characterized.

Clavamine Argiopine, Argiotoxin-636 Argiopinin I Argiopinin II Argiopinin III, Argiotoxin 659 Argiopinin 1V Argiopinin V Pseudoargiopinin I Pseudoargiopinin II Pseudoargiopinin III Argiotoxin-480 Argiotoxin-650 Argiotoxin-494 Argiotoxin-622 ~ Argiotoxin-673 Argiotoxin-503 Argiotoxin-517 Argiotoxin-645 a

159

142 142 142 142 142 142 142 142

142

147 147 147 147 147 147 147 147

140, 142

146, 147

142 142 142 142 142 142 142 142

142

142

145

145

145

140, 142

211

145, 211

209 145,210, 211, 212

4~ t~

2428

McCORMICK AND MEINWALD

The lengths of the polyamine chains show considerable variation. The smallest chain is only seven atoms long (pseudoargiopinin III), whereas the longest extends for 43 atoms (NPTX-6). These chains consist of NH, NCH3, or +N(CH3) 2 groups separated by segments of three to six carbon atoms. The carbon segments are linear chains of methylene groups, with the exception of some three and all of the six carbon segments, which appear as/3-alanine and lysine units, respectively. Interestingly, all of the lysine's terminal nitrogens are either mono- or dimethylated, but few of the other amino groups are substituted. Finally, the first segment within the polyamine chain of most known amino acid containing acylpolyamines is a five carbon unit. The non-amino acid-containing acylpolyamines (Figure 2), although discovered later, appear to be more widely distributed among spider species. Currently, 17 different compounds have been fully characterized from five different families: the Theraphosidae, Dipluridae, Ctenizidae, Agelenidae, and Pisauridae (Table 4). These compounds consist solely of an acyl end group attached to a polyamine chain. The carbon segments consist entirely of trimethylene, tetramethylene, and pentamethylene units. Some of the nitrogen atoms separating the first and second carbon segments occur as hydroxylamines, and a few terminal nitrogens are methylated. Unlike the amino acid-containing acylpolyamines, the first polyamine segment of the non-amino acid-containing compounds is usually a three or occasionally a four or five carbon unit.

SYNTHESIS OF SPIDER NEUROTOXINS

Largely because of the inevitably small supply of individual, biologically active components from spider venoms, synthesis is an extremely important tool for confirming postulated structures and for making useful amounts of material available for in vitro and in vivo experimentation. All of the successful synthetic work in this field is described in roughly a score o f recent papers, already referred to in the final columns of Tables 3 and 4. The syntheses combine many well known elements of polyamine and amino acid chemistry; they illustrate the importance of specific protecting groups, and the use of particular techniques for handling strongly basic, hydrophilic compounds, some of which are also sensitive to oxygen. To save space, each synthesis is presented simply as a chart with an indication of the reagents used to bring about each step. They are grouped in two sets: the first set (Schemes 1-14) describe polyamines containing amino acid moieties, and the second set (Schemes 15-19) describe those lacking amino acids. Within each group, the schemes are presented in alphabetical order with respect to the first author's names.

170

164, 171, 164, 171, 170

167, 176 167, 176

164, 167, 170, 171

167, 170, 171 164, 167, 170, 171

Agelenopsis aperta

181 181 181 18I 181 181 181

181

181

181

Hololena curta

aStmctures are only partially characterized.

AG 452 AGEL 452 AG 489 AGEL 489 HO 489 AG 505 AGEL 505 HO 505 AG 504 AGEL 505a AG 488 AGEL 489 AGEL 468 HO 468 AGEL 448 HO 473 HO 452 HO 448 HO 395 HO 416a HO 416b HO 359 Het 389 Her 403 Apc 600" Apc 728 a CNS 2103

Compound

32 32

Hebestatis Iheveniti

32

sp.

Harpactirella

32 32

Aphonopelma chalcodes

? 49

Atrax robustus

TABLE 4. NoN-AMINO ACID-CONTAINING ACYLPOLYAMINES FROM SPIDERS

Dolomedes okefinokensis

32

170

170

176

170

170

170

Synthesis

t.~ 4~ b~ ~D

2430

McCORMICK

B~"'--..-~.../--

~-.~I

~ - . / ~ I

Bn

2

AND

a

"

Bn

O

NH b,c

I

I

Bn

MEINWALD

[

Bn

_--

H

NHZ

o

ii

)

H O

Nit b,d(or e)f

I

1

H CONH2

Bn

OH

H

I

I

Bn

H

I

~

lb

H

O

0

Nil

l U

]

II

-

O O

t

I

H ~CONH2

t

H

I

H

g

H

NI-I2

I

H

~p~e OH

l

~ , ~ v

H

II

~ "Ir

fl

li

O

0

=-"

~

O

I

H CONH2

I

H

I

H

I

H

Nil

-

NH2

I

H

I

H

Argiotoxin 6 5 9

SCHEME. 1. Adams, 1987 (ref. 145). a. No~-Z-Nw-nitro-Arg, DCC, HOBt; b. TFA; c. BOC-Asp, DCC, HOBt; e. 2,4-dibenzyloxyphenyl acetyl chloride; f. 4-benzyloxyindole3-acetic acid, DCC, HOBt; g. H2, Pd-C, HOAc. CONCLUDING REMARKS With the structures of so many natural acylpolyamines known, and with synthetic samples of these and their analogs now becoming available in useful quantities, the stage has been set for an intensive study of the receptors at which these toxins act. How these receptors interact with polyamine neurotoxins must now become a major objective of future research. While the details of such research lie outside the scope of this review, they can be expected to illuminate what it is that the spiders, as applied neuropharmacologists, have been getting at. At the least, spider neurotoxins represent a powerful set of neurobiological research tools. Whether they ultimately provide a new generation of useful drugs and insecticides remains to be seen. From the viewpoint of the chemical ecologist, the chemical and biological studies that have been completed so far should certainly whet the appetite for a deeper understanding of how venoms influence the relationships between spiders and the rest of the world.

OBn

H

OBn

OBB

N

~.o.~j

BnO

BBO

H

OBn

O _

O-pNP

~..~,,~

o

o

f

Nit2

92%

O

BnO~.. ~

::

~

H

"~CON~

1

0

g,h,i

[

j,k

31%

46%

I

HO-

v

I

--OONIr JSTX-3

SCHEME. 2 . Hashimoto, 1987 fief. 206). a. NaBH4, MeOH; b. SOC12, Phil, A; c. NaCN, DMSO; d. KOH, EtOH, A; e. L-Asn-OpNP. TEA, DMF; f. H2N(CH2)sNH2, DMF; g. Z-CI, NaHCO3, Et20-H20; h. KOH, EtOH; i. p-nitrophenol, DCC, DMF; j. 1, HOBt, DMF; k. H2, Pd-C, HOAc.

~7~:"~I~,,~

NH2

a,b,c,a,b

gOO~sJ" " ~ , J ~

N

56%

0

l

~

c

~

/

~

/

~

/

~

~

~

W

I Bn

~

~

NPhth

] Bn

c,d 88%

NH

"~

~

[

[

[

":-

I

Bn

Bn

H

NHZ

Z

0

*" 79%

0

Nil

e,g(orh),Lj "

I H

I Bn

CONH2

OH

I

_/x.~v J

H

[I

HtY

I Bn

I n

" ~Z

I z

O

II o

O

"

I

:,,,CONH H2

I

I

n

NH

]

~

"

H

~2

I

H

Argiopke OH

H

It I1 ,~jx... j I

-I~

I

H

II

o

O

-

~

O

I

H

O3NH2

I

H

I

H

L

~

NH

-

~2

I

H

Argiotoxin-659

SCHEME. 3. Jasys, 1988 (ref. 211). a. NaBH4, PhCHO, MgSO4, MeOH; b. PhthN(CH2)3Br, KF-celite, CH3CN, A; c. H2NNH2, MeOH, ; gD; d. Z-Arg(Z2)-Su, CH2C12; e. TFA; f. BOC-Asn-O-pNP, TEA, CH2C12; g. 2,4-dibenzyloxyphenyl acetic acid, CH2C12; h. 4-benzyloxyindole-3-aceticacid, DCC, HOBt, CH2C12; i. H2, Pd(OH)2, HOAc; j. Amberlite CG-50, MeOH, HC1.

'tt3 ~

N

H

a,b,c ~,

I

I

N~

Z

!

d,e

I

N

~

N

~

N

H

I

T

! N

C~l3 OBz

NHZ

Z

I

H Nil

0

I

d,~d,g 56% 9

z

o H

ii

65%

I

~

!

H

I

43%

CH3 ~

CH3

CH3

II

H

I

I

!

NHZ

Z

I

h,i 88%

H

/

OH

I

~,.

H

0

I

II

v

J

[I

11

I

-"

o

-iN~

~

:~.

I

I

H

I

NI-I2

H

I

II ~

II

~

CONH2

I

H

CH3

11

o Argioto:~Jn-673

SCHEME. 4. Jasys, 1988 (ref. 211). a. PhthN(CH2)3Br, KF-celite, CH3CN, ; gD; b. H2NNH2, MeOH, A; c. Z-Arg(Z2)-Su, CH2C12; d. TFA; e. BOCHN(CH2)3Br, Na2CO3, CH3CN, A; f. BOC-Asn-O-pNP, TEA, CH2C12; g. 4-benzyloxyindole-3-acetic acid, DCC; HOBt, CH2C12; h. H2, Pd(OH)2, HOAc; i. Amberlite CG-50, MeOH, HC1. H

2

~

O

H

a,b,c 96%

d 69%

O

H

O

e

76%

N3

~t~...x~r ~1

O

O

H O

H

~[--~11~

f

89%

O

O g 51%

H ~

N

0

0

_

H

=

0

NIt2

NPTX-11

SCHEME. 5. Miyashita, 1992 (ref. 207). a. (BOC)20, aq. Na2CO3; b. MsC1, pyr, CH2C12; c. NAN3, DMF; d. TFA, CH2C12; BOC-Asn-OpNP, TEA, DMF; e. TFA, CH2C12; indole-3-acetyl-OpNP, TEA, DMF; f. H2, Pd-C, EtOH; BOC-Orn(Z)-OpNP, TEA, DMF; g. TFA, CH2C12; H2, Pd-C, EtOH.

SPIDER VENUMS

2433

H

O

O

I