The potencies of synthetic analogues of saxitoxin and the absolute stereoselectivity of decarbamoyl saxitoxin.
The potencies of synthetic saxitoxin (+/- STX) and six of its synthetic analogues, including the enantioselectively synthesized unnatural (-)enantiomer of decarbamoyl saxitoxin (dcSTX), were measured and compared to those of natural saxitoxin [(+)STX]. The analogues, all of which were racemic (+/-) mixtures except for dcSTX, varied in the substituents at the C6 position, the carbamoyl 'moeity', and the C12 position, the hydrated ketone. The ability of the toxins to inhibit the compound action potential (AP) and to displace radiolabeled natural saxitoxin (3H-STX) from nerve membranes at equilibrium were both used as potency assays. Biological activity of both (+)- and (-)dcSTX was analyzed by the kinetics of block of single Na+ channels reconstituted in planar lipid bilayer membranes, where it was demonstrated that only (+)dcSTX had biological activity. The potency of STX analogues fell markedly as the substituent at the C6 position became smaller; Ki values from the binding competition assay (at 4 degrees C) are: (+/-)6-methanolic-STX, 5 x 10(-10) M; (+/-)6-methyl-STX, 1 x 10(-6) M; (+/-)6-dihydro-STX, 3.5 x 10(-5) M. Replacement of the ketone at the C12 position by a methylene group was accomplished in two derivatives, although both also had substituents at the C6 position. The compound (+/-)6-methyl-12-deoxy-STX was about 0.03 as potent as (+/-)6-methyl-STX and only 10(-5) as potent as racemic (+/-)STX. In synthetic compounds where the benzyloxymethyl (-CH2OCH2C6H5) substituent occurred at the C6 position, the C12-methylene derivative still displayed some binding activity (Ki = 6 x 10(-4) M). However, when the same C6 derivatized compounds also contained a 6-membered heterocyclic group (-C3H8S2-) conjugated to carbon 12, the measured binding affinity was even further decreased (Ki = 2 x 10(-3) M). The findings show that substitutions on the carbon 6 position of STX have stronger effects on STX potency than previously believed, and that the toxin may form a hydrogen bond with the sodium channel at this site. Furthermore, the total removal of oxygen from the C12 position does not completely abolish the binding activity of the molecule.
Strichartz
G R
GR
Anesthesia Research Laboratories, Brigham and Women's Hospital, Boston, MA, USA.
Hall
S
S
Magnani
B
B
Hong
C Y
CY
Kishi
Y
Y
Debin
J A
JA
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001923:Brain Chemistry / Q000187:drug effects; D004594:Electrophysiology; D066298:In Vitro Techniques; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011817:Rabbits; D011894:Rana pipiens; D051381:Rats; D012530:Saxitoxin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000633:toxicity; D012584:Sciatic Nerve / Q000187:drug effects; D015222:Sodium Channels / Q000187:drug effects / Q000378:metabolism; D013237:Stereoisomerism; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism
D015222:Sodium Channels; D012530:Saxitoxin; C034903:decarbamylsaxitoxin