title: Precise and combinatorial PEGylation generates a low-immunogenic and stable form of human growth hormone.
abstract: In this study, we aimed to develop a safe and stable form of human growth hormone (hGH) and to refine PEGylation methods for therapeutic proteins via genetic code expansion. Through this precise approach, a series of polyethylene glycol (PEG) moieties and sites were combined in various ways. Additionally, the effects of combinatorial PEGylation on the biological, pharmacological, and immunogenic properties of hGH in vitro and vivo were analyzed. Our results showed that combinatorial PEGylation at Y35, G131, and K145 significantly reduced immunogenicity and improved pharmacokinetic (PK) profiles compared with mono-PEGylation, while retaining biological activity. Upon re-examination of the pharmacodynamics in hypophysectomized rats, multi-PEGylated hGH was found to be much more stable than mono-PEGylated hGH. Thus, this method for combinatorial, precise PEGylation may facilitate the development of next-generation, long-acting hGH with low immunogenicity.
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Ling
authors.initials: L
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Jingxian
authors.initials: J
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Yiming
authors.initials: Y
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China; Center for Translational Medicine, Peking Union Medical College Hosptial, Beijing 100730, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Xiaochun
authors.initials: X
authors.lastname: Cai
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Ziwei
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: LongLong
authors.initials: L
authors.lastname: Si
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Huan
authors.initials: H
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: YongXiang
authors.initials: Y
authors.lastname: Zheng
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: ChuanLing
authors.initials: C
authors.lastname: Zhang
authors.affiliation: National Institutes for Food and Drug Control, Beijing 100050, China.
authors.forename: Chenggang
authors.initials: C
authors.lastname: Liang
authors.affiliation: National Institutes for Food and Drug Control, Beijing 100050, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Li
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Lihe
authors.initials: L
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China. Electronic address: deminzhou@bjmu.edu.cn.
authors.forename: Demin
authors.initials: D
authors.lastname: Zhou
mesh_terms: D000818:Animals; D000917:Antibody Formation; D002460:Cell Line; D057930:Click Chemistry; D005815:Genetic Code; D019382:Human Growth Hormone / Q000737:chemistry* / Q000235:genetics / Q000276:immunology* / Q000493:pharmacokinetics; D006801:Humans; D008958:Models, Molecular; D011092:Polyethylene Glycols / Q000737:chemistry* / Q000493:pharmacokinetics; D055550:Protein Stability; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000493:pharmacokinetics
chemical_list: D011994:Recombinant Proteins; D019382:Human Growth Hormone; D011092:Polyethylene Glycols
keywords: Copper-free click chemistry; Expanded genetic code; Multisite specific PEGylation; Recombinant human growth hormone