Stability of IgG isotypes in serum

Abstract

Drug development from early discovery to late stage commercialization is a long arduous process where a number of factors are taken into consideration when deciding on a particular immunoglobulin isotype for a therapeutic purpose. There are no general rules for which isotype is selected; however, prior experiences, effector function and the specific therapy targeted, as well as extensive testing early in development help in pairing the number of candidates. Over 20 monoclonal antibodies are FDA-approved, and most are IgG1 isotype, although a number of non-IgG1 molecules have been approved recently and the number in development is on the rise. Analytical techniques that examine the physicochemical properties of a molecule provide vital information on the stability and efficacy of candidate antibody therapeutics, but most of these studies are conducted using standard buffers and under well defined storage conditions. It has recently become apparent that analysis of antibody therapeutics recovered after circulation in blood show altered physicochemical characteristics, and in many instances therapeutic molecules recovered from serum show lower potency. This review examines some of these studies, with a focus on the physicochemical changes observed in the molecules. Technologies that can facilitate rapid screening of candidate antibody therapeutics directly from blood are highlighted. The facts indicate that antibody therapeutic development programs must incorporate understanding of the basic biology of the isotype and its stability in serum, which is the intended environment of the therapeutic.

Figure 1

Schematic of the human IgG isotypes and their disulfide linkages.

Figure 2

Deamidation of asparagine residue.

Figure 5

The different Fc glycans present on antibodies.

Figure 4

Exchange of Fab arms between endogenous IgG4 molecules and a therapeutic IgG4 molecule.

Figure 5

Formation of functional covalent dimers between identical as well as different IgG2 molecules.

Figure 6

Shuffling of inter-chain disulfide bonds of IgG2 in serum.

Figure 7

Analysis on LabChip GXII showing recovery of mAbs from blood-precision data (n = 3) after 4 and 24 hours.

Figure 8

(See opposite page). Monitoring aggregate and fragment formation of a mAb in whole blood using the LabChip GXII (T = 0, 4 and 24 hrs).