Label-free Fab and Fc affinity/avidity profiling of the antibody complex half-life for polyclonal and monoclonal efficacy screening

Abstract

A unified approach to affinity screening for Fab and Fc interactions of an antibody for its antigen and FcγR receptor has been developed. An antigen array is used for the Fab affinity and cross-reactivity screening and protein A/G proxy is the FcγR receptor. The affinities are derived using a simple 1:1 binding model with a consistent error analysis. The association and dissociation kinetics are measured over optimised times for accurate determination. The Fab/Fc affinities are derived for ten antibodies: mAb-actin (mouse), pAb-BSA (sheep), pAb-collagen V (rabbit), pAb-CRP (goat), mAb-F1 (mouse), mAbs (mouse) 7.3, 12.3, 29.3, 36.3 and 46.3 raised against LcrV in Yersinia pestis. The rate of the dissociation of antigen-antibody complexes relates directly to their immunological function as does the Fc-FcγR complex and a new half-life plot has been defined with a Fab/Fc half-life range of 17-470 min. The upper half-life value points to surface avidity. Two antibodies that are protective as an immunotherapy define a Fab half-life >250 min and an Fc half-life >50 min as characteristics of ideal interactions which can form the basis of an antibody screen for immunotherapy.

Antibody Structure and Function: The Basis for Engineering Therapeutics

“…However, Fc region clustering due to FcγR interactions can contribute to Fab target avidity [322][323][324]. The avidity due to Fc region-FcγR crosslinking can affect the immune cell effector function that can contribute to autoimmune disorders [325][326][327][328][329].…”

Site-specific PEGylation of an anti-CEA/CD3 bispecific antibody improves its antitumor efficacy

“…Based on the serum protein concentration up to 144 h, S-Fab had an in vivo half-life of ~3.0 h, similar to the reported half-life of other Fabs in vivo. [36][37][38] PEGylation significantly extended the half-life of PEG-S-Fab to 36.0 h, a 12-fold increase over that of S-Fab ( Figure 6B).…”

Covalent nano delivery systems for selective imaging and treatment of brain tumors

“…The rate of interaction of CDS with cell surface receptors is governed by the slow dissociation of the receptor complexes with high affinity binding antibody modules. Their replacement by peptides decreases the size and also binding affinity and accelerates dissociation [32, 33], (Table 2). In our recent preclinical study, the replacement resulted in the reduction of hydrodynamic diameter from 22 nm to 7.8 nm while retaining excellent therapeutic efficacy [23] (Fig.…”