Generation of a Novel High-Affinity Antibody Binding to PCSK9 Catalytic Domain with Slow Dissociation Rate by CDR-Grafting, Alanine Scanning and Saturated Site-Directed Mutagenesis for Favorably Treating Hypercholesterolemia

Overview

Journal Biomedicines

Specialty Biomedical Engineering

Date 2021 Dec 24

PMID 34944600

Citations 1

Authors

Zhengli Bai

Menglong Xu

Ying Mei

Tuo Hu

Panpan Zhang

Manman Chen

Wenxiu Lv

Chenchen Lu

Shuhua Tan

Affiliations

Soon will be listed here.

Abstract

Inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) has become an attractive therapeutic strategy for lowering low-density lipoprotein cholesterol (LDL-C). In this study, a novel high affinity humanized IgG1 mAb (named h5E12-L230G) targeting the catalytic domain of human PCSK9 (hPCSK9) was generated by using CDR-grafting, alanine-scanning mutagenesis, and saturated site-directed mutagenesis. The heavy-chain constant region of h5E12-L230G was modified to eliminate the cytotoxic effector functions and mitigate the heterogeneity. The biolayer interferometry (BLI) binding assay and molecular docking study revealed that h5E12-L230G binds to the catalytic domain of hPCSK9 with nanomolar affinity ( = 1.72 nM) and an extremely slow dissociation rate (, 4.84 × 10 s), which interprets its quite low binding energy (-54.97 kcal/mol) with hPCSK9. Additionally, h5E12-L230G elevated the levels of LDLR and enhanced the LDL-C uptake in HepG2 cells, as well as reducing the serum LDL-C and total cholesterol (TC) levels in hyperlipidemic mouse model with high potency comparable to the positive control alirocumab. Our data indicate that h5E12-L230G is a high-affinity anti-PCSK9 antibody candidate with an extremely slow dissociation rate for favorably treating hypercholesterolemia and relevant cardiovascular diseases.

Citing Articles

Comparison of "framework Shuffling" and "CDR Grafting" in humanization of a PD-1 murine antibody.

Wang Y, Chen Y, Xu H, Rana G, Tan X, He M Front Immunol. 2024; 15:1395854.

PMID: 39076979 PMC: 11284016. DOI: 10.3389/fimmu.2024.1395854.

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