Multimodal biopanning of T7 phage-displayed peptides reveals angiomotin as a potential receptor of the anti-angiogenic macrolide Roxithromycin

  • Eur J Med Chem. 2015 Jan 27:90:809-21. doi: 10.1016/j.ejmech.2014.12.015.
Kaori Takakusagi  1 ,  Yoichi Takakusagi  2 ,  Takahiro Suzuki  3 ,  Aya Toizaki  3 ,  Aiko Suzuki  1 ,  Yaichi Kawakatsu  1 ,  Madoka Watanabe  1 ,  Yukihiro Saito  4 ,  Ryushi Fukuda  4 ,  Atsuo Nakazaki  3 ,  Susumu Kobayashi  3 ,  Kengo Sakaguchi  1 ,  Fumio Sugawara  5
Affiliations
  • 1. Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
  • 2. Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan. Electronic address: [email protected].
  • 3. Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
  • 4. Meiwafosis Co. Ltd., 1-14-2 Shinjuku, Tokyo 160-0022, Japan.
  • 5. Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan. Electronic address: [email protected].
PMID: 25528335 DOI: 10.1016/j.ejmech.2014.12.015
Abstract

Roxithromycin (RXM) is a semi-synthetic fourteen-membered macrolide Antibiotic that shows anti-angiogenic activity in solid tumors. In the present study, we conducted biopanning of T7 phage-displayed peptides either on a 96-well formatted microplate, a flow injection-type quartz-crystal microbalance (QCM) biosensor, or a cuvette-type QCM. RXM-selected peptides of different sequence, length and number were obtained from each mode of screening. Subsequent bioinformatics analysis of the RXM-selected peptides consistently gave positive scores for the extracellular domain (E458-T596) of angiomotin (Amot), indicating that this may comprise a binding region for RXM. Bead pull down assay and QCM analysis confirmed that RXM directly interacts with Amot via the screen-guided region, which also corresponds to the binding site for the endogenous anti-angiogenic inhibitor angiostatin (Anst). Thus, multimodal biopanning of T7PD revealed that RXM binds to the extracellular domain on Amot as a common binding site with Anst, leading to inhibition of angiogenesis-dependent tumor growth and metastasis. These data might explain the molecular basis underlying the mechanism of action for the anti-angiogenic activity of RXM.

Keywords
Angiogenesis; Angiomotin; QCM-D; RELIC; Roxithromycin; T7 phage display.
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