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

  • Bioconjug Chem. 2022 Nov 16;33(11):2180-2188. doi: 10.1021/acs.bioconjchem.2c00416.
Chiao-Yu Hsiao  1 ,  Jun-Lun Meng  1 ,  Jung-Zhe Hong  1 ,  Xuan-Huong Ly  1 ,  Meng-Hsuan Lin  1 ,  Chin-Yuan Chang  1  2 ,  Minh-Tram T Nguyen  1 ,  Tian-Lu Cheng  2 ,  Wen-Wei Lin  3 ,  Pierre-Alain Burnouf  4 ,  Talal Salem Al-Qaisi  5 ,  En-Shuo Liu  6 ,  Yu-Cheng Su  1  2
Affiliations
  • 1. Department of Biological Science and Technology, Center for Intelligent Drug Systems and Smart Bio-devices (IDS2B), National Yang Ming Chiao Tung University, Hsinchu 300193, Taiwan.
  • 2. Department of Biomedical Science and Environmental Biology, Drug Development and Value Creation Research Center, Kaohsiung Medical University, Kaohsiung 807378, Taiwan.
  • 3. School of Post-Baccalaureate Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 807378, Taiwan.
  • 4. International Center for Wound Repair and Regeneration (iWRR), National Cheng-Kung University, Tainan 701401, Taiwan.
  • 5. Department of Medical Laboratory Sciences, Pharmacological and Diagnostic Research Centre, Faculty of Allied Medical Sciences, Al-Ahliyya Amman University, Amman 19328, Jordan.
  • 6. Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 807378, Taiwan.
PMID: 36320124 DOI: 10.1021/acs.bioconjchem.2c00416
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 mPEG5K and high-molecular-weight (hmw) mPEG20K at 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.

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