Fab-dsFv: A bispecific antibody format with extended serum half-life through albumin binding

Abstract

An antibody format, termed Fab-dsFv, has been designed for clinical indications that require monovalent target binding in the absence of direct Fc receptor (FcR) binding while retaining substantial serum presence. The variable fragment (Fv) domain of a humanized albumin-binding antibody was fused to the C-termini of Fab constant domains, such that the VL and VH domains were individually connected to the Cκ and CH1 domains by peptide linkers, respectively. The anti-albumin Fv was selected for properties thought to be desirable to ensure a durable serum half-life mediated via FcRn. The Fv domain was further stabilized by an inter-domain disulfide bond. The bispecific format was shown to be thermodynamically and biophysically stable, and retained good affinity and efficacy to both antigens simultaneously. In in vivo studies, the serum half-life of Fab-dsFv, 2.6 d in mice and 7.9 d in cynomolgus monkeys, was equivalent to Fab'-PEG.

Antibody Structure and Function: The Basis for Engineering Therapeutics

“…In cases where transient drug exposure is favorable (such as diagnostics), this apparent defect can lead to a desired increase in clearance. On the other hand, the half-life can be prolonged by incorporating albumin-binding functionality into one of the antigen-binding domains [494].…”

Preclinical optimization of antibody‐based radiopharmaceuticals for cancer imaging and radionuclide therapy—Model, vector, and radionuclide selection

“…The use of serum albumin binding moieties to increase the blood half-life of small molecules or affibodies found promising applications in radiotherapy and can be similarly applied to antibody fragments (131,132). Davé et al recently designed an antibody format referred as Fab-dsFv, composed of the variable fragment of a humanized albumin-binding antibody fused to the C-termini of Fab constant domain (Figure 7A) (133). The anti-albumin variable fragment resulted in an increase serum half-life in both mice (t 1/2 = 2.9 days) and cynomolgus monkeys (t 1/2 = 7.9 days) (133).…”

“…Davé et al recently designed an antibody format referred as Fab-dsFv, composed of the variable fragment of a humanized albumin-binding antibody fused to the C-termini of Fab constant domain (Figure 7A) (133). The anti-albumin variable fragment resulted in an increase serum half-life in both mice (t 1/2 = 2.9 days) and cynomolgus monkeys (t 1/2 = 7.9 days) (133). A sequential delivery approach, referred as pretargeting, has also been developed in order to improve therapeutic indices (Figure 7B) (25).…”

“…A) Schematic representation of an Fab-dsFv (133). An disulfide bonded Fab is conjugated to an anti-HSA variable fragment (Fv) domain containing an incorporated disulfide bond via an amino acid linker.…”

Fabs-in-tandem immunoglobulin is a novel and versatile bispecific design for engaging multiple therapeutic targets

“…11,12,28,29 The design of mutation and linkers was critical to their proper function and in vitro/in vivo stability. 30 Although the mutation can be generic and applied to any antibody couple with the same family germline, optimization and new design are still needed for different germline families. 31 Structurally, when a variable domain is connected to anther Iglike domain at its N-terminus, a linker is always required to provide enough space and flexibility for antigen binding, otherwise a strong steric hindrance may occur.…”