N-terminus of Etanercept is Proteolytically Processed by Dipeptidyl Peptidase-4.
Pharm Res
; 39(10): 2541-2554, 2022 Oct.
Article
de En
| MEDLINE
| ID: mdl-35986123
ABSTRACT
PURPOSE:
Biologics are structurally heterogeneous and can undergo biotransformation in the body. Etanercept (ETN) is a fusion protein composed of a soluble tumor necrosis factor (TNF) receptor and the Fc portion of human immunoglobulin G1. The N-terminus of ETN has a putative sequence cleaved by dipeptidyl peptidase-4 (DPP-4). The purpose of this study was to investigate the biotransformation of ETN in humans and mice and evaluate its effects on functional properties.METHODS:
An analytical method using liquid chromatography-mass spectrometry (LC-MS/MS) was established. The N-terminal heterogeneity of ETN was assessed in the serum of patients with rheumatoid arthritis or mice receiving ETN. The in vitro N-terminal truncation was explored using recombinant DPP-4. The binding affinity to TNF-α or TNF-ß was investigated using an in-house enzyme-linked immunosorbent assay.RESULTS:
In the formulations, about 90% of ETN had an intact N-terminus, while the N-terminal truncated form was most abundant in the serum of the patients with rheumatoid arthritis and mice. Recombinant human DPP-4 cleaved two amino acids from the N-terminus of ETN in vitro. Sitagliptin, a DPP-4 inhibitor, inhibited N-terminal truncation both in vivo and in vitro. However, N-terminal truncation did not affect the binding ability to TNF-α or TNF-ß and the pharmacokinetics of ETN. ETN biosimilars exhibited similar characteristics to the reference product in vivo and in vitro.CONCLUSIONS:
ETN undergoes N-terminal truncation in the body, and DPP-4 cleaves exogenous ETN via N-terminal proteolysis. The application of an MS-based assay will detect novel biotransformation of therapeutic proteins.Mots clés
Texte intégral:
1
Collection:
01-internacional
Base de données:
MEDLINE
Sujet principal:
Polyarthrite rhumatoïde
/
Antirhumatismaux
/
Inhibiteurs de la dipeptidyl-peptidase IV
/
Produits pharmaceutiques biosimilaires
Limites:
Animals
/
Humans
Langue:
En
Journal:
Pharm Res
Année:
2022
Type de document:
Article
Pays d'affiliation:
Japon