# Enhanced ADCC Activity of a C-Terminal Lysine Variant of an IgG1 Antibody Driven by N-Linked MAN5 Glycan Using a Reporter Gene Assay

## TL;DR

This study shows that a specific antibody variant with a C-terminal lysine boosts ADCC activity due to a MAN5 glycan, not the lysine itself.

## Contribution

The study identifies the MAN5 glycan, not the C-terminal lysine, as the key driver of enhanced ADCC activity in IgG1 antibody variants.

## Key findings

- Variants with C-terminal lysine residues (K1 and K2) showed higher ADCC activity than K0 and acidic variants.
- The enhanced ADCC activity was attributed to the N-linked MAN5 glycan rather than the lysine residue itself.
- The findings challenge previous assumptions about the role of C-terminal lysine in ADCC activity.

## Abstract

Background: Antibody-dependent cellular cytotoxicity (ADCC) is an immune response where antibodies bind to target cells and activate effector cells through Fcγ receptors, ultimately leading to the destruction of the target cells. Methods: This study examined the ADCC activities of charge variants of a therapeutic IgG1, MAB1, using an internally developed reporter gene assay. In this assay, the proprietary target was expressed on DiFi cells, while FcγRIIIa was expressed on Jurkat effector cells. Results: The results revealed that different charge variants had varying levels of ADCC activity, with variants containing C-terminal lysine residues showing enhanced activity. The charge variants arose from modifications such as the presence of sialic acid at the glycan moiety, deamidation, and C-terminal lysine truncation, including K2 (two C-terminal lysine residues), K1 (one C-terminal lysine residue), and K0 (no C-terminal lysine residues) variants. Notably, the K1 and K2 variants demonstrated higher ADCC activity compared to the K0 and acidic variants. However, the observed increase was attributed not to the lysine residue itself, but rather to the MAN5 glycan associated with the lysine-containing variants. Conclusion: These findings challenge previous assumptions about the role of C-terminal lysine in ADCC, suggesting a shift in understanding the functional significance of charge variants and emphasizing the critical influence of glycan composition in therapeutic antibody efficacy.