A novel immunotoxin reveals a new role for CD321 in endothelial cells

PLoS One. 2017 Oct 13;12(10):e0181502. doi: 10.1371/journal.pone.0181502. eCollection 2017.

Abstract

There are currently several antibody therapies that directly target tumors, and antibody-drug conjugates represent a novel moiety as next generation therapeutics. Here, we used a unique screening probe, DT3C, to identify functional antibodies that recognized surface molecules and functional epitopes, and which provided toxin delivery capability. Accordingly, we generated the 90G4 antibody, which induced DT3C-dependent cytotoxicity in endothelial cells. Molecular analysis revealed that 90G4 recognized CD321, a protein localized at tight junctions. Although CD321 plays a pivotal role in inflammation and lymphocyte trans-endothelial migration, little is known about its mechanism of action in endothelial cells. Targeting of CD321 by the 90G4 immunotoxin induced cell death. Moreover, 90G4 immunotoxin caused cytotoxicity primarily in migratory endothelial cells, but not in those forming sheets, suggesting a critical role for CD321 in tumor angiogenesis. We also found that hypoxia triggered redistribution of CD321 to a punctate localization on the basal side of cells, resulting in functional impairment of tight junctions and increased motility. Thus, our findings raise the intriguing possibility that endothelial CD321 presented cellular localization in tight junction as well as multifunctional dynamics in several conditions, leading to illuminate the importance of widely-expressed CD321 as a potential target for antitumor therapy.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Death / drug effects
  • Cell Line
  • Cell Movement / drug effects
  • Endothelial Cells / cytology
  • Endothelial Cells / drug effects*
  • Endothelial Cells / metabolism*
  • Humans
  • Immunotoxins / immunology
  • Immunotoxins / toxicity*
  • Junctional Adhesion Molecule A / chemistry
  • Junctional Adhesion Molecule A / immunology
  • Junctional Adhesion Molecule A / metabolism*

Substances

  • Immunotoxins
  • Junctional Adhesion Molecule A

Grant support

This study was supported in part by KAKENHI, a Grant-in-Aid for Encouragement of Young Scientists (B) (TF as PI, 25830123), Grant-in-Aid for Scientific research (C) (TF as PI, 16K07184) from Japan Society for the Promotion of Science (JSPS), the Ministry of Education, Culture, Sports, Science and Technology (MEXT)-Supported Program for the Strategic Research Foundation at Private Universities (TF and TW as co-PI, S1411014) at Tokyo University of Pharmacy and Life Sciences, and The Vehicle Racing Commemorative Foundation (TF and TW as PI).