title: AL-57, a ligand-mimetic antibody to integrin LFA-1, reveals chemokine-induced affinity up-regulation in lymphocytes.
abstract: Affinity of integrin lymphocyte function-associated antigen 1 (LFA-1) is enhanced by conformational changes from the low-affinity closed form to the high-affinity (HA) open form of the ligand-binding inserted (I) domain as shown by work with purified I domains. However, affinity up-regulation of LFA-1 on the cell surface by physiological agonists such as chemokines has yet to be demonstrated by monovalent reagents. We characterize a mAb, AL-57 (activated LFA-1 clone 57), that has been developed by phage display that selectively targets the HA open conformation of the LFA-1 I domain. AL-57 discriminates among low-affinity, intermediate-affinity, and HA states of LFA-1. Furthermore, AL-57 functions as a ligand mimetic that binds only upon activation and requires Mg2+ for binding. Compared with the natural ligand intercellular adhesion molecule-1, AL-57 shows a tighter binding to the open I domain and a 250-fold slower off rate. Monovalent Fab AL-57 demonstrates affinity increases on a subset (approximately 10%) of lymphocyte cell surface LFA-1 molecules upon stimulation with CXCL-12 (CXC chemokine ligand 12). Affinity up-regulation correlates with global conformational changes of LFA-1 to the extended form. Affinity increase stimulated by CXCL-12 is transient and peaks 2 to 5 min after stimulation.
authors.affiliation: CBR Institute for Biomedical Research and Department of Anesthesia, Harvard Medical School, 200 Longwood Avenue, Boston, MA 02115, USA. shimaoka@cbrinstitute.org
authors.forename: Motomu
authors.initials: M
authors.lastname: Shimaoka
authors.forename: Minsoo
authors.initials: M
authors.lastname: Kim
authors.forename: Edward H
authors.initials: EH
authors.lastname: Cohen
authors.forename: Wei
authors.initials: W
authors.lastname: Yang
authors.forename: Nathan
authors.initials: N
authors.lastname: Astrof
authors.forename: Dan
authors.initials: D
authors.lastname: Peer
authors.forename: Azucena
authors.initials: A
authors.lastname: Salas
authors.forename: Audrey
authors.initials: A
authors.lastname: Ferrand
authors.forename: Timothy A
authors.initials: TA
authors.lastname: Springer
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000378:metabolism*; D000915:Antibody Affinity; D001665:Binding Sites; D002460:Cell Line; D054377:Chemokine CXCL12; D019743:Chemokines, CXC / Q000378:metabolism*; D000939:Epitopes; D006801:Humans; D008024:Ligands; D016169:Lymphocyte Function-Associated Antigen-1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008214:Lymphocytes / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D015854:Up-Regulation
chemical_list: D000911:Antibodies, Monoclonal; C515255:CXCL12 protein, human; D054377:Chemokine CXCL12; D019743:Chemokines, CXC; D000939:Epitopes; D008024:Ligands; D016169:Lymphocyte Function-Associated Antigen-1