High Proteolytic Resistance of Spider-Derived Inhibitor Cystine Knots.

Kyoko Kikuchi,Mika Sugiura,Tadashi Kimura

doi:10.1155/2015/537508

Abstract

Proteolytic stability in gastrointestinal tract and blood plasma is the major obstacle for oral peptide drug development. Inhibitor cystine knots (ICKs) are linear cystine knot peptides which have multifunctional properties and could become promising drug scaffolds. ProTx-I, ProTx-II, GTx1-15, and GsMTx-4 were spider-derived ICKs and incubated with pepsin, trypsin, chymotrypsin, and elastase in physiological conditions to find that all tested peptides were resistant to pepsin, and ProTx-II, GsMTx-4, and GTx1-15 showed resistance to all tested proteases. Also, no ProTx-II degradation was observed in rat blood plasma for 24 hours in vitro and ProTx-II concentration in circulation decreased to half in 40 min, indicating absolute stability in plasma and fast clearance from the system. So far, linear peptides are generally thought to be unsuitable in vivo, but all tested ICKs were not degraded by pepsin and stomach could be selected for the alternative site of drug absorption for fast onset of the drug action. Since spider ICKs are selective inhibitors of various ion channels which are related to the pathology of many diseases, engineered ICKs will make a novel class of peptide medicines which can treat variety of bothering symptoms.

Highlights

Proteolytic degradation in gastrointestinal (GI) tract and blood plasma is a major barrier for peroral peptide drug development [1,2,3,4]
More than 60 FDA-approved peptide medicines are on the market, approximately 140 peptide drugs are currently in clinical trials, and more than 500 therapeutic peptides are in preclinical stage [5]. ω-Conotoxin MVIIA, known as ziconotide or Prialt, is the first FDA-approved peptide drug from marine cone snail Conus magus to treat severe chronic pain; ziconotide needs to be intrathecally administered because of poor in vivo stability [6]
HPRP and ProTx-I were almost completely digested within 1-hour incubation; other Inhibitor cystine knots (ICKs) peptides were degraded to approximately 80% of the original amount in 2 hours, but the degradation did not proceed beyond that point (Figure 1(b))

Summary

Introduction

Proteolytic degradation in gastrointestinal (GI) tract and blood plasma is a major barrier for peroral peptide drug development [1,2,3,4]. Despite high target selectivity, peptides are long thought to be unsuitable for oral delivery due to their susceptibility to degradation and low bioavailability. At this time, more than 60 FDA-approved peptide medicines are on the market, approximately 140 peptide drugs are currently in clinical trials, and more than 500 therapeutic peptides are in preclinical stage [5]. Several strategies to enhance proteolytic stability and bioavailability have been developed for oral administration of therapeutic peptides [8]. Modification of peptide termini, replacement of labile amino acid, and cyclization of a peptide are used to promote peptide stability in GI tract and plasma [7]; increasing molecular mass by PEGylation, coadministration of enzyme inhibitor and permeation enhancers, and encapsulation of peptides in biodegradable polymer microspheres

Methods

Results

Conclusion