Mulcahy, J. V. | Search bioRxiv

bioRxiv2024

Structural basis for saxitoxin congener binding and neutralization by anuran saxiphilins

Dinoflagellates and cyanobacteria in harmful red tide algal blooms produce saxitoxin (STX) and [~]50 congeners that block voltage-gated sodium channel (NaV) function and disrupt bioelectrical signals1-4. Consuming seafood carrying these lethal toxins causes paralytic shellfish poisoning (PSP), a growing public health hazard due to climate change5-7 that motivates efforts to detect these toxins and counteract their noxious effects. Although structural studies of NaVs8,9 and anuran soluble STX binding proteins known as saxiphilins (Sxphs)10,11 revealed convergent binding modes for the bis-guanidinium STX core10,11, the structural basis for STX congener recognition is unknown. Here, we show that American bullfrog (Rana catesbeiana) RcSxph10,11 and High Himalaya frog (Nanorana parkeri) NpSxph10 use a pre-organized pocket to sequester STX congeners through a binding mode shared with STX. This lock and key recognition yields a tradeoff between a relatively rigid high-affinity toxin binding site in which bound waters are crucial and the ability of Sxphs to accommodate STX congener modifications. Importantly, functional studies show that Sxphs act as toxin sponges that reverse NaV block by multiple STX congeners and can detect these bis-guanidinium toxins in a radioligand receptor binding assay (RBA) for PSP toxin environmental testing12,13. Our findings establish how Sxphs sequester diverse neurotoxins and reveal structural factors underlying STX congener binding differences between Sxphs and NaVs that are rooted in the distinct toxin binding orientations on these two targets. These insights expand the molecular foundation required for understanding toxin sponge action and for guiding development of new means to monitor PSTs and mitigate their harmful effects.

biophysics↗

bioRxiv2019

Discovery of a Selective, State-Independent Inhibitor of NaV1.7 by Modification of Guanidinium Toxins

The voltage-gated sodium channel isoform NaV1.7 is highly expressed in small diameter dorsal root ganglion neurons and is obligatory for nociceptive signal transmission. Genetic gain-of-function and loss-of-function NaV1.7 mutations have been identified in select individuals, and are associated with episodic extreme pain disorders and insensitivity to pain, respectively. These findings implicate NaV1.7 as a key pharmacotherapeutic target for the treatment of pain. While several small molecules targeting NaV1.7 have been advanced to clinical development, no NaV1.7-selective compound has shown convincing efficacy in clinical pain applications. Here we describe the discovery and characterization of ST-2262, a NaV1.7 inhibitor that blocks the extracellular vestibule of the channel with an IC50 of 72 nM and greater than 200-fold selectivity over off-target sodium channel isoforms, NaV1.1-1.6 and NaV1.8. In contrast to other NaV1.7 inhibitors that preferentially inhibit the inactivated state of the channel, ST-2262 is equipotent against resting and inactivated protein conformers. In a non-human primate model, animals treated with ST-2262 exhibit markedly reduced sensitivity to noxious heat. These findings establish the extracellular vestibule of the sodium channel as a viable receptor site for selective ligand design and provide insight into the pharmacology of state-independent inhibition of NaV1.7. Significance StatementPain is among the most common reasons for seeking medical care, yet many frequently prescribed drugs, particularly the opioids, cause problematic side effects and carry a risk of addiction. Voltage-gated sodium ion channels (NaVs) have emerged as promising targets for the development of non-opioid pain medicines. NaVs are involved in the propagation of electrical signals along neurons throughout the body. Humans born without a functional copy of one sodium channel subtype, NaV1.7, are unable to experience most types of pain. In the present work, we disclose the discovery and characterization of a selective inhibitor of NaV1.7 that reduces sensitivity to a painful thermal stimulus in non-human primates. Findings from this work may help guide the development of novel, non-addictive drug candidates as alternatives to opioids.

neuroscience↗