Engineering a High-Affinity Anti-Methoxy Poly(ethylene glycol) (mPEG) Antibody for Sensitive Immunosensing of mPEGylated Therapeutics and mPEG Molecules

Chiao Yu Hsiao, Jun Lun Meng, Jung Zhe Hong, Xuan Huong Ly, Meng Hsuan Lin, Chin Yuan Chang, Minh Tram T. Nguyen, Tian Lu Cheng, Wen Wei Lin, Pierre Alain Burnouf, Talal Salem Al-Qaisi, En Shuo Liu, Yu-Cheng Su*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Sensitive quantification of methoxy poly(ethylene glycol) (mPEG)-conjugated therapeutics for pharmacokinetic determination is critical for mPEGylated drug development. However, sensitive measurement of low-molecular-weight (lmw) mPEG compounds remains challenging due to epitope competition between backbone-specific anti-PEG antibodies. Here, we engineered a high-affinity methoxy-specific anti-mPEG antibody for sensitive quantification of free mPEG molecules and mPEGylated therapeutics. The affinity-enhanced h15-2Y antibody variant shows a 10.3-fold increase in mPEG-binding activity compared to parental h15-2b. h15-2Y-based sandwich ELISA can effectively quantify lmw mPEG5Kand high-molecular-weight (hmw) mPEG20Kat concentrations as low as 3.4 and 5.1 ng mL-1, respectively. Moreover, lmw mPEG compounds (560, 750, 1000, and 2000 Da) can be efficiently quantified via h15-2Y-based competitive ELISA with detection limits at nanomolar levels. This study provides a promising approach for application in the quantitative analysis of the various sizes of mPEG molecules to accelerate the timeline of mPEG-conjugated drug development.

Original languageEnglish
Pages (from-to)2180-2188
Number of pages9
JournalBioconjugate Chemistry
Volume33
Issue number11
DOIs
StatePublished - 16 Nov 2022

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