The voltage-gated sodium channel 1.7 (NaV1.7) has recently emerged as a promising analgesic target. Gain-of-function mutations in the SNC9A gene encoding the pore-forming α-subunit of NaV1.7 cause painful inherited neuropathies whereas loss-of-function mutations result in a congenital indifference to all forms of pain. Thus, selective blockers of NaV1.7 are likely to be powerful analgesics. However, NaV1.7 is only one of nine human NaV subtypes, and improper function of certain members of this ion channel family can cause debilitating or even lethal channelopathies. Thus, therapeutics designed to target NaV1.7 must have exquisite selectivity. Of particular concern for a NaV1.7-targeted analgesic would be off-target effects on NaV1.5, which is responsible for the rising phase of the cardiac action potential, or the muscle-specific subtype NaV1.4.
Modulation of NaV channels is a dominant pharmacology in spider venoms, and hence we decided to screen an extensive panel of >200 spider venoms for blockers of this channel. Using an in-house, high-throughput FLIPR-based screen, 36% of all spider venoms that we assayed were found to contain potent blockers of the human NaV1.7 channel. Using this assay, we purified a total of 41 peptidic blockers of human NaV1.7 from 25 “hit” venoms. Sequencing of these peptides revealed that they fall into three distinct structural classes, although they all contain three disulfide bonds.
One of these structural classes, which contains a large number of related toxins that nevertheless have diverse selectivities against the various NaV subtypes is of particular interest. Toxins from this family inhibit NaV channel activation by binding to the voltage sensor of channel domain II. A novel approach for rapidly mapping the pharmacophore of these toxins circumvents the need to produce and purify mutant toxins. Careful structural and functional characterization of this family of toxins is providing detailed information on the residues responsible for the interaction of these toxins not just with the desired therapeutic target (NaV1.7) but also critical off-target subtypes such as NaV1.5. It is anticipated that development of detailed structure-function relationships for this class of toxins will enable us to engineer highly specific blockers of the human NaV1.7 channel that will be therapeutically useful for the treatment of chronic pain.
Structural, functional, and in vivo analgesic data will be presented for one novel peptide with more than 100-fold selectivity for NaV1.7 over the critical off-target subtypes NaV1.4 and NaV1.5.