Strategic design to create HER2-targeting proteins with target-binding peptides immobilized on a fibronectin type III domain scaffold

  • RSC Adv. 2020 Apr 17;10(26):15154-15162. doi: 10.1039/d0ra00427h.
Wanaporn Yimchuen  1 ,  Tetsuya Kadonosono  1 ,  Yumi Ota  1 ,  Shinichi Sato  2 ,  Maika Kitazawa  1 ,  Tadashi Shiozawa  1 ,  Takahiro Kuchimaru  3 ,  Masumi Taki  4 ,  Yuji Ito  5 ,  Hiroyuki Nakamura  2 ,  Shinae Kizaka-Kondoh  1
Affiliations
  • 1. School of Life Science and Technology, Tokyo Institute of Technology Yokohama 226-8501 Japan [email protected] +81-45-924-5848 +81-45-924-5848.
  • 2. Institute of Innovative Research, Tokyo Institute of Technology Yokohama 226-8501 Kanagawa Japan.
  • 3. Center for Molecular Medicine, Jichi Medical University Shimotsuke 329-0498 Japan.
  • 4. Graduate School of Informatics and Engineering, The University of Electro-Communications Tokyo 182-8585 Japan.
  • 5. Graduate School of Science and Engineering, Kagoshima University Kagoshima 890-0065 Japan.
PMID: 35495466 DOI: 10.1039/d0ra00427h
Abstract

Tumor-binding peptides such as human epidermal growth factor receptor 2 (HER2)-binding peptides are attractive therapeutic and diagnostic options for Cancer. However, the HER2-binding peptides (HBPs) developed thus far are susceptible to proteolysis and lose their affinity to HER2 in vivo. In this report, a method to create a HER2-binding fluctuation-regulated affinity protein (HBP-FLAP) consisting of a fibronectin type III domain (FN3) scaffold with a structurally immobilized HBP is presented. HBPs were selected by phage-library screening and grafted onto FN3 to create FN3-HBPs, and the HBP-FLAP with the highest affinity (HBP sequence: YCAHNM) was identified after affinity maturation of the grafted HBP. HBP-FLAP containing the YCAHNM peptide showed increased proteolysis-resistance, binding to HER2 with a dissociation constant (K D) of 58 nM in ELISA and 287 nM in biolayer interferometry and specifically detects HER2-expressing Cancer cells. In addition, HBP-FLAP clearly delineated HER2-expressing tumors with a half-life of 6 h after intravenous injection into tumor-bearing mice. FN3-based FLAP is an excellent platform for developing target-binding small proteins for clinical applications.

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