ProTx-II, a selective inhibitor of NaV1.7 sodium channels, blocks action potential propagation in nociceptors.
Voltage-gated sodium (Na(V)1) channels play a critical role in modulating the excitability of sensory neurons, and human genetic evidence points to Na(V)1.7 as an essential contributor to pain signaling. Human loss-of-function mutations in SCN9A, the gene encoding Na(V)1.7, cause channelopathy-associated indifference to pain (CIP), whereas gain-of-function mutations are associated with two inherited painful neuropathies. Although the human genetic data make Na(V)1.7 an attractive target for the development of analgesics, pharmacological proof-of-concept in experimental pain models requires Na(V)1.7-selective channel blockers. Here, we show that the tarantula venom peptide ProTx-II selectively interacts with Na(V)1.7 channels, inhibiting Na(V)1.7 with an IC(50) value of 0.3 nM, compared with IC(50) values of 30 to 150 nM for other heterologously expressed Na(V)1 subtypes. This subtype selectivity was abolished by a point mutation in DIIS3. It is interesting that application of ProTx-II to desheathed cutaneous nerves completely blocked the C-fiber compound action potential at concentrations that had little effect on Abeta-fiber conduction. ProTx-II application had little effect on action potential propagation of the intact nerve, which may explain why ProTx-II was not efficacious in rodent models of acute and inflammatory pain. Mono-iodo-ProTx-II ((125)I-ProTx-II) binds with high affinity (K(d) = 0.3 nM) to recombinant hNa(V)1.7 channels. Binding of (125)I-ProTx-II is insensitive to the presence of other well characterized Na(V)1 channel modulators, suggesting that ProTx-II binds to a novel site, which may be more conducive to conferring subtype selectivity than the site occupied by traditional local anesthetics and anticonvulsants. Thus, the (125)I-ProTx-II binding assay, described here, offers a new tool in the search for novel Na(V)1.7-selective blockers.
Schmalhofer
William A
WA
Department of Ion Channels, Merck Research Laboratories, Rahway, NJ 07065-0900, USA.
Calhoun
Jeffrey
J
Burrows
Rachel
R
Bailey
Timothy
T
Kohler
Martin G
MG
Weinglass
Adam B
AB
Kaczorowski
Gregory J
GJ
Garcia
Maria L
ML
Koltzenburg
Martin
M
Priest
Birgit T
BT
D000200:Action Potentials / Q000187:drug effects*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D006801:Humans; D015640:Ion Channel Gating; D008297:Male; D023421:Models, Animal; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009619:Nociceptors / Q000187:drug effects*; D011869:Radioligand Assay; D051381:Rats; D017207:Rats, Sprague-Dawley; D017386:Sequence Homology, Amino Acid; D026941:Sodium Channel Blockers / Q000494:pharmacology*; D015222:Sodium Channels / Q000737:chemistry / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D013111:Spider Venoms / Q000494:pharmacology*
D017931:DNA Primers; C470626:ProTx-II peptide; D026941:Sodium Channel Blockers; D015222:Sodium Channels; D013111:Spider Venoms