WPK5, a Novel Kunitz-Type Peptide from the LeechInhibiting Factor XIa, and Its Loop-Replaced Mutant to Improve Potency.
Research article published in Biomedicines (2021)
Abstract
Kunitz-type proteins or peptides have been found in many blood-sucking animals, but the identity of them in leeches remained elusive. In the present study, five Kunitz-type peptides named WPK1-WPK5 were identified from the leech Whitmania pigra. Recombinant WPK1-WPK5 were expressed in Pichia pastoris GS115, and their inhibitory activity against Factor XIa (FXIa) was tested. WPK5 showed inhibitory activity against FXIa with an IC50 value of 978.20 nM. To improve its potency, the loop replacement strategy was used. The loop 1 (TGPCRSNLER) and loop 2 (QYGGC) in WPK5 were replaced by loop 1 (TGPCRAMISR) and loop 2 (FYGGC) in PN2KPI, respectively, and the resulting peptide named WPK5-Mut showed an IC50 value of 8.34 nM to FXIa, which is about 100-fold the potency of FXIa compared to that of WPK5. WPK5-Mut was further evaluated for its extensive bioactivity in vitro and in vivo. It dose-dependently prolonged APTT on both murine plasma and human plasma, and potently inhibited FeCl3-induced carotid artery thrombosis in mice at a dose of 1.5 mg/kg. Additionally, WPK5-Mut did not show significant bleeding risk at a dose of 6 mg/kg. Together, these results showed that WPK5-Mut is a promising candidate for the development of an antithrombotic drug.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Summary
Kunitz-type proteins or peptides have been found in many blood-sucking animals, but the identity of them in leeches remained elusive. In the present study, five Kunitz-type peptides named WPK1-WPK5 were identified from the leech.
Why This Matters for Hirudotherapy
This study identified five Kunitz-type peptides (WPK1-WPK5) from the leech Whitmania pigra, expressed them recombinantly, and found WPK5 inhibited coagulation Factor XIa (IC50 978.20 nM); an engineered loop-replaced variant, WPK5-Mut, was about 100-fold more potent (IC50 8.34 nM), prolonged clotting times in murine and human plasma, inhibited FeCl3-induced carotid thrombosis in mice at 1.5 mg/kg, and showed no significant bleeding at 6 mg/kg. This is among the most directly relevant entries to ASH's mission: it is concrete evidence that the medicinal-leech secretome yields novel, druggable anticoagulant peptides, and that protein engineering can sharpen a natural leech molecule into an antithrombotic drug candidate. The work is preclinical (in vitro assays and rodent models with an engineered mutant), not a clinical trial, so it supports the leech-derived drug-discovery rationale but says nothing about clinical hirudotherapy or human efficacy and safety.
Citation
WPK5, a Novel Kunitz-Type Peptide from the LeechInhibiting Factor XIa, and Its Loop-Replaced Mutant to Improve Potency.
Zheng YZ et al. · Biomedicines, 2021
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