In vitro affinity maturation to improve the efficacy of a hypoxia-inducible factor 1α single-domain intrabody

Min Hu; Guangbo Kang; Xin Cheng; Jiewen Wang; Ruowei Li; Zixuan Bai; Dong Yang; He Huang

doi:10.1016/j.bbrc.2020.06.097

In vitro affinity maturation to improve the efficacy of a hypoxia-inducible factor 1α single-domain intrabody

Biochem Biophys Res Commun. 2020 Sep 3;529(4):936-942. doi: 10.1016/j.bbrc.2020.06.097. Epub 2020 Jul 30.

Authors

Min Hu¹, Guangbo Kang¹, Xin Cheng¹, Jiewen Wang¹, Ruowei Li¹, Zixuan Bai¹, Dong Yang², He Huang³

Affiliations

¹ Department of Biochemical Engineering, School of Chemical Engineering & Technology, Tianjin University, Tianjin, 300350, China; Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, 300072, China.
² Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, 300072, China; School of Environmental Science & Engineering, Tianjin University, Tianjin, 300072, China. Electronic address: dongyang@tju.edu.cn.
³ Department of Biochemical Engineering, School of Chemical Engineering & Technology, Tianjin University, Tianjin, 300350, China; Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), Tianjin University, Tianjin, 300072, China. Electronic address: huang@tju.edu.cn.

PMID: 32819602
DOI: 10.1016/j.bbrc.2020.06.097

Abstract

Affinity is an important property of therapeutic antibodies, so improving affinity is critical to the biological activity and clinical efficacy. An anti-HIF-1α nanobody, VHH212, was screened via a native ribosome display library with a 26.6 nM of K_D value was used as the parent. In this paper, a Venn-intersection of multi-algorithms screening (VIMAS) strategy for computer-aided binding affinity prediction was designed. Homology modeling and protein docking methods were used to substitute the need for a crystal structure. Finally, a mutant with a 17.5-fold enhancement in binding affinity (1.52 nM) was obtained by using the VIMAS strategy. Furthermore, the biological activity of mutants was verified at the cellular level. Targeting HIF-1α can sensitize PDAC (pancreatic ductal adenocarcinoma) tumors to gemcitabine, which is a potential co-treatment method for pancreatic cancer patients. Our results showed that the cytotoxicity of gemcitabine on pancreatic cancer cell lines increased with the enhanced-affinity of an intrabody under combined treatment.

Keywords: Affinity maturation; Combined treatment; Homology modeling; Hypoxia-inducible factor; Nanobody.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Algorithms*
Antibody Affinity
Antibody Specificity
Antimetabolites, Antineoplastic / pharmacology
Antineoplastic Agents, Immunological / chemistry
Antineoplastic Agents, Immunological / metabolism
Antineoplastic Agents, Immunological / pharmacology*
Binding Sites
Cell Line, Tumor
Cell Survival / drug effects
Cell Survival / genetics
Deoxycytidine / analogs & derivatives
Deoxycytidine / pharmacology
Gemcitabine
Humans
Hypoxia-Inducible Factor 1, alpha Subunit / antagonists & inhibitors*
Hypoxia-Inducible Factor 1, alpha Subunit / genetics
Hypoxia-Inducible Factor 1, alpha Subunit / immunology
Molecular Docking Simulation
Molecular Dynamics Simulation
Mutation
Pancreatic Ducts / immunology
Pancreatic Ducts / pathology
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Single-Domain Antibodies / chemistry
Single-Domain Antibodies / genetics
Single-Domain Antibodies / pharmacology*
Structural Homology, Protein
User-Computer Interface

Substances

Antimetabolites, Antineoplastic
Antineoplastic Agents, Immunological
HIF1A protein, human
Hypoxia-Inducible Factor 1, alpha Subunit
Single-Domain Antibodies
Deoxycytidine
Gemcitabine