title: Quantification and modeling of tripartite CD2-, CD58FC chimera (alefacept)-, and CD16-mediated cell adhesion.
abstract: Alefacept is a chimeric protein combining CD58 immunoglobulin-like domain 1 with human IgG1 Fc. Alefacept mediates adhesion by bridging CD2 on T cells to activating Fc receptors on effector cells, but the equilibrium binding parameters have not been determined. Alefacept mediated T cell killing by NK cells and adhesion between CD2- and CD16-expressing cells at an optimum concentration of 100 nM. We introduce novel measurements with supported planer bilayers, from which key two-dimensional and three-dimensional parameters can be determined by data fitting. Alefacept competitively inhibited cell bilayer adhesion mediated by the CD2-CD58 interaction. Alefacept mediated maximal adhesion of CD2(+) T cells to CD16B, an Fc receptor, in planar bilayers at 500 nM. A mechanistic model for alefacept-mediated cell-bilayer adhesion allowed fitting of the data and determination of two-dimensional binding parameters. These included the density of bonds in the adhesion area, which grew to maintain a consistent average bond density of 200 molecules/microm(2) and two-dimensional association constants of 3.1 and 630 microm(2) for bivalently and monovalently bound forms of alefacept, respectively. The maximum number of CD16 bound and the fit value of 4,350 CD2 per cell are much lower than the 40,000 CD2 per cell measured with anti-CD2 Fab. These results suggest that additional information is needed to correctly predict Alefacept-mediated bridge formation.
authors.affiliation: Department of Pathology, New York University School of Medicine and Helen L. and Martin S. Kimmel Center for Biology and Medicine of Skirball Institute of Biomolecular Medicine, New York, New York 10016, USA. dustin@saturn.med.nyu.edu
authors.forename: Michael L
authors.initials: ML
authors.lastname: Dustin
authors.forename: Toby
authors.initials: T
authors.lastname: Starr
authors.forename: Daniel
authors.initials: D
authors.lastname: Coombs
authors.forename: Gerard R
authors.initials: GR
authors.lastname: Majeau
authors.forename: Werner
authors.initials: W
authors.lastname: Meier
authors.forename: Paula S
authors.initials: PS
authors.lastname: Hochman
authors.forename: Adam
authors.initials: A
authors.lastname: Douglass
authors.forename: Ron
authors.initials: R
authors.lastname: Vale
authors.forename: Byron
authors.initials: B
authors.lastname: Goldstein
authors.forename: Adrian
authors.initials: A
authors.lastname: Whitty
mesh_terms: D000077944:Alefacept; D000818:Animals; D018801:CD2 Antigens / Q000096:biosynthesis*; D018968:CD58 Antigens / Q000737:chemistry*; D016466:CHO Cells; D002448:Cell Adhesion; D006224:Cricetinae; D003412:Cricetulus; D003879:Dermatologic Agents / Q000737:chemistry / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D006801:Humans; D019169:Jurkat Cells; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry; D011485:Protein Binding; D011961:Receptors, Fc / Q000737:chemistry; D017452:Receptors, IgG / Q000096:biosynthesis* / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000494:pharmacology; D013601:T-Lymphocytes / Q000378:metabolism
chemical_list: D018801:CD2 Antigens; D018968:CD58 Antigens; D003879:Dermatologic Agents; D008051:Lipid Bilayers; D011961:Receptors, Fc; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D000077944:Alefacept