Programme Contents

Spider-venom peptides that target the human Na_v1.7 channel: potential analgesics for the treatment of chronic pain

The voltage-gated sodium channel 1.7 (Na_V1.7) has recently emerged as a promising analgesic target. Gain-of-function mutations in the SNC9A gene encoding the pore-forming α-subunit of Na_V1.7 cause painful inherited neuropathies whereas loss-of-function mutations result in a congenital indifference to all forms of pain. Thus, selective blockers of Na_V1.7 are likely to be powerful analgesics. However, Na_V1.7 is only one of nine human Na_V subtypes, and improper function of certain members of this ion channel family can cause debilitating or even lethal channelopathies. Thus, therapeutics designed to target Na_V1.7 must have exquisite selectivity. Of particular concern for a Na_V1.7-targeted analgesic would be off-target effects on Na_V1.5, which is responsible for the rising phase of the cardiac action potential, or the muscle-specific subtype Na_V1.4.

Modulation of Na_V 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 Na_V1.7 channel. Using this assay, we purified a total of 41 peptidic blockers of human Na_V1.7 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 Na_V subtypes is of particular interest. Toxins from this family inhibit Na_V 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 (Na_V1.7) but also critical off-target subtypes such as Na_V1.5. 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 Na_V1.7 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 Na_V1.7 over the critical off-target subtypes Na_V1.4 and Na_V1.5.