Inhibition of ENaC Activity by Novel Peptide Trypsin‐Like Inhibitors Derived from Amphibian Skin Secretions
Authors: Carroll Emma L. , Douglas Lisa E.J. , Reihill James A. , Zhou Mei , Chen Tianbao , McGarvey Lorcan P. , Lundy Fionnuala T. , Hollywood Mark A. , Lockhart John C. , Martin S. Lorraine 
Published: June 2021

Impaired mucociliary clearance (MCC) constitutes the main hallmark of the chronic bronchitis phenotype in COPD. Whilst aberrant processes involved in mucus secretion and ciliary function are important drivers of abnormal MCC, dysregulation of airway hydration appears to be the principal determinant affecting this disorder (1). The hydration status of the airways is largely governed by interplay between the epithelial sodium channel (ENaC) and the cystic fibrosis transmembrane conductance regulator (CFTR). ENaC is activated by trypsin‐like (TL), channel activating proteases (CAPs) and work carried out in cystic fibrosis has shown that inhibition of these proteases increases airways surface liquid (ASL) volume and normalises MCC (2).The aim of this study was to characterise a number of natural TL peptide inhibitors, derived from amphibian skin secretions (frog peptides), based on their ability to inhibit CAPs and regulate ENaC function.Initial screening established an inhibitory profile for each frog peptide against known recombinant CAPs using relevant fluorogenic substrate‐based activity assays. The peptides were then added to fully differentiated human primary bronchial epithelial cells (HBEs) grown at air‐liquid interface (ALI) and ENaC activity determined by obtaining equivalent short circuit current (Ieq) readings using the Transepithelial Current Clamp system (TECC‐24; EP Devices).When examined against recombinant proteases (rTrypsin, rHAT, rMatriptase, rProstasin, rFurin), the frog peptides showed different inhibitory profiles with no single molecule demonstrating overall superiority. Of these, QUB‐2136 rapidly inhibited amiloride‐sensitive Ieq when applied apically to HBEs, to a greater degree than the other compounds investigated. Interestingly, we note that for several of these frog peptides, including QUB‐2136, inactivation of ENaC was so acute that the compounds behave more like direct channel blockers rather than acting indirectly through inhibition of the CAPs.We report for the first time novel biologically active natural peptides, derived from amphibian skin secretions, to be potent inhibitors of CAPs and rapid inactivators of ENaC‐mediated Na+ transport in airway cells. These compounds represent a novel strategy towards ENaC inhibition which has potential to rehydrate the airways, restore effective MCC and reduce the devastating pulmonary decline observed in CF, irrespective of CFTR mutation. Importantly, this strategy could be effective for other chronic airways disease patients such as COPD and non‐CF bronchiectasis.Support or Funding InformationBREATH (Border and REgions Airways Training Hub) is funded by the EU's INTERREG Va programme managed by the Special EU Programmes Body (SEUPB).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.