Glycosylation decreases aggregation and immunogenicity of adalimumab fab secreted from Pichia pastoris.

Abstract:

:Glycoengineering of therapeutic proteins has been applied to improve the clinical efficacy of several therapeutics. Here, we examined the effect of glycosylation on the properties of the Fab of the therapeutic antibody, adalimumab. An N-glycosylation site was introduced at position 178 of the H-chain constant region of adalimumab Fab through site-directed mutagenesis (H: L178N Fab), and the H: L178N Fab was produced in Pichia pastoris. Expressed mutant Fab contained long and short glycan chains (L-glyco Fab and S-glyco Fab, respectively). Under the condition of aggregation of Fab upon pH shift-induced stress, both of L-glyco Fab and S-glyco Fab were less prone to aggregation, with L-glyco Fab suppressing aggregation more effectively than the S-glyco Fab. Moreover, the comparison of the antigenicity of glycosylated and wild-type Fabs in mice revealed that glycosylation resulted in the suppression of antigenicity. Analysis of the pharmacokinetic behavior of the Fab, L-glyco Fab, and S-glyco Fab indicated that the half-lives of glycosylated Fabs in the rats were shorter than that of wild-type Fab, with L-glyco Fab having a shorter half-life than S-glyco Fab. Thus, we demonstrated that the glycan chain influences Fab aggregation and immunogenicity, and glycosylation reduces the elimination half-life in vivo.

journal_name

J Biochem

journal_title

Journal of biochemistry

authors

Nakamura H,Kiyoshi M,Anraku M,Hashii N,Oda-Ueda N,Ueda T,Ohkuri T

doi

10.1093/jb/mvaa116

subject

Has Abstract

pub_date

2020-10-27 00:00:00

eissn

0021-924X

issn

1756-2651

pii

5941469

pub_type

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