title: Contribution of Antibody Hydrodynamic Size to Vitreal Clearance Revealed through Rabbit Studies Using a Species-Matched Fab.
abstract: We have developed a tool Fab fragment of a rabbit monoclonal antibody that is useful for early evaluation in rabbit models of technologies for long acting delivery (LAD) of proteins to the eye. Using this Fab we show that vitreal clearance can be slowed through increased hydrodynamic size. Fab (G10rabFab) and Fab' (G10rabFab') fragments of a rabbit monoclonal antibody (G10rabIgG) were expressed in Chinese hamster ovary (CHO) cells and purified using antigen-based affinity chromatography. G10rabFab retains antigen-binding upon thermal stress (37 °C) for 8 weeks in phosphate-buffered saline (PBS) and can be detected in rabbit tissues using an antigen-based ELISA. Hydrodynamic radius, measured using quasi-elastic light scattering (QELS), was increased through site-specific modification of the G10rabFab' free cysteine with linear methoxy-polyethylene glycol(PEG)-maleimide of 20000 or 40000 molecular weight. Pharmacokinetic studies upon intravitreal dosing in New Zealand white rabbits were conducted on the G10rabFab and PEGylated G10rabFab'. Results of single and multidose pharmacokinetic experiments yield reproducible results and a vitreal half-life for G10rabFab of 3.2 days. Clearance from the eye is slowed through increased hydrodynamic size, with vitreal half-life showing a linear dependence on hydrodynamic radius (RH). A linear dependence of vitreal half-life on RH suggests that molecule diffusivity makes an important contribution to vitreal clearance. A method for prediction of vitreal half-life from RH measurements is proposed.
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Whitney
authors.initials: W
authors.lastname: Shatz
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Philip E
authors.initials: PE
authors.lastname: Hass
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Mary
authors.initials: M
authors.lastname: Mathieu
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Hok Seon
authors.initials: HS
authors.lastname: Kim
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Kim
authors.initials: K
authors.lastname: Leach
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Michelle
authors.initials: M
authors.lastname: Zhou
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Yongping
authors.initials: Y
authors.lastname: Crawford
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Amy
authors.initials: A
authors.lastname: Shen
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Kathryn
authors.initials: K
authors.lastname: Wang
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Debby P
authors.initials: DP
authors.lastname: Chang
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Mauricio
authors.initials: M
authors.lastname: Maia
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Susan R
authors.initials: SR
authors.lastname: Crowell
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Leslie
authors.initials: L
authors.lastname: Dickmann
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Justin M
authors.initials: JM
authors.lastname: Scheer
authors.affiliation: Departments of Protein Chemistry, ‡Antibody Engineering, §Early Stage Cell Culture, ∥Bioanalytical Assay Services, ⊥Preclinical and Translational Pharmacokinetics, and #Drug Delivery, Genentech Inc. , 1 DNA Way, South San Francisco, California 94080, United States.
authors.forename: Robert F
authors.initials: RF
authors.lastname: Kelley
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000008:administration & dosage / Q000493:pharmacokinetics*; D016466:CHO Cells; D003412:Cricetulus; D004797:Enzyme-Linked Immunosorbent Assay; D057446:Hydrodynamics; D007140:Immunoglobulin Fab Fragments / Q000008:administration & dosage* / Q000378:metabolism*; D058449:Intravitreal Injections; D007700:Kinetics; D011092:Polyethylene Glycols / Q000737:chemistry; D011817:Rabbits
chemical_list: D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D011092:Polyethylene Glycols
keywords: PEGylation; hydrodynamic radius; long acting delivery; ocular pharmacokinetics