Development of a human antibody that exhibits antagonistic activity toward CC chemokine receptor 7.

Abstract

Background: CC chemokine receptor 7 (CCR7) is a member of G-protein-coupled receptor family and mediates chemotactic migration of immune cells and different cancer cells induced via chemokine (C-C motif) ligand 19 (CCL19) or chemokine (C-C motif) ligand 21 (CCL21). Hence, the identification of blockade antibodies against CCR7 could lead to the development of therapeutics targeting metastatic cancer.

Methods: CCR7 was purified and stabilized in its active conformation, and antibodies specific to purified CCR7 were screened from the synthetic M13 phage library displaying humanized scFvs. The in vitro characterization of selected scFvs identified two scFvs that exhibited CCL19-competitive binding to CCR7. IgG₄'s harboring selected scFv sequences were characterized for binding activity in CCR7⁺ cells, inhibitory activity toward CCR7-dependent cAMP attenuation, and the CCL19 or CCL21-dependent migration of CCR7⁺ cells.

Results: Antibodies specifically binding to purified CCR7 and CCR7⁺ cells were isolated and characterized. Two antibodies, IgG₄(6RG11) and IgG₄(72C7), showed ligand-dependent competitive binding to CCR7 with K_D values of 40 nM and 50 nM, respectively. Particularly, IgG₄(6RG11) showed antagonistic activity against CCR7, whereas both antibodies significantly blocked the ligand-induced migration and invasion activity of CCR7⁺ cancer cells.

Conclusions: Two antibody clones were successfully identified from a synthetic scFv-displaying phage library using purified recombinant CCR7 as an antigen. Antibodies specifically bound to the surface of CCR7⁺ cells and blocked CCR7⁺ cell migration. Particularly, 6RG11 showed antagonist activity against CCR7-dependent cAMP attenuation.