Spinning natural venoms into drug discovery: The journey of a new potent peptide blocker of the human Cav1.2 channel subtype from Poecilotheria subfusca spider

Abstract

Animal venoms have been explored as rich and valuable sources of new peptide modulators that target a large variety of ion channel subtypes. Over the last two decades, natural peptides have been identified with the ability to modulate several targets at a time. Post-discovery studies are used to then refine the identity of the best target. We employed a similar strategy to identify a novel and highly potent modulator of the human Cav1.2 channel; a high throughput screening of a large library of 200 animal venoms was initiated through automated whole-cell patch‐clamp experiments using engineered cells overexpressing human voltage-gated sodium NaV1.7 or NaV1.5 channel subtypes. Out of several positive venom fractions, one bioactive peptide, poecitoxin‐1a (PecTx‐1a) from the Poecilotheria subfusca spider venom, was purified for its higher potency to block NaV1.7 over NaV1.5 and identified based on mass spectrometry sequencing. The peptide is 35 amino acids long and belongs to the inhibitor cystine knot (ICK) structural family. Next, the synthetic peptide was generated and found identical to the native one, which allowed further characterization on a large set of voltage-gated calcium, voltage-gated potassium and inward-rectifier potassium (hKir) channel subtypes overexpressed in recombinant cells. This ion channel selectivity profiling uncovered hCaV1.2 as the best target of PecTx‐1a (IC50 of 24 nM) making this peptide the best-known inhibitor of this cardiac channel isoform to date. Further characterization of the peptide on the action potential and calcium currents of human induced pluripotent stem cell-derived cardiomyocytes indicates the pharmacological value of this peptide in modulating cardiac excitability. In conclusion, PecTx‐1a is the first high affinity ICK spider toxin that targets the L-type hCaV1.2 channel with high affinity and should therefore represent a valuable tool to study the CaV1.2 subtype physiological and pharmacological functions in healthy and pathological models.