Antibody structure and engineering considerations for the design and function of Antibody Drug Conjugates (ADCs)

Abstract

Antibody-drug conjugates (ADCs) are emerging as effective tools in cancer therapy, combining the antibody's exquisite specificity for the target antigen-expressing cancer cell together with the cytotoxic potency of the payload. Much success stems from the rational design of "toxic warheads", chemically linked to antibodies, and from fine-tuning the intricate properties of chemical linkers. Here, we focus on the antibody moiety of ADCs, dissecting the impact of Fab, linkers, isotype and Fc structure on the anti-tumoral and immune-activating functions of ADCs. Novel design approaches informed by antibody structural attributes present opportunities that may contribute to the success of next generation ADCs.

Keywords: Antibodies; Antibody Drug Conjugate (ADC); IgG1; IgG4; Immunoglobulin Fc; biodistribution; effector functions; kinetics; stability.

Figure 1.

Schematic of ADC components and their role in ADC design, engineering and functions. The Fab region (A) is responsible for antigen recognition and binding, and can lead to ADC internalization. Therefore, the Fab region needs to be targeted to tumor-associated antigens that are homogenously expressed on tumor cells, ideally with little or no expression on normal cells. The payload is attached to the antibody via a cleavable or non-cleavable linker (B). Non-cleavable linkers rely on the complete degradation of the antibody after internalization of the ADC, whereas most cleavable linkers are cleaved by different mechanisms depending on the linker (i.e. proteases, reduction) and some cleavable linkers do not depend on ADC internalization for payload release and can result in higher off-target cytotoxicities. The hydrophobicity of linkers can play a vital role in the biodistribution of an ADC. Linkers can be attached non-selectively via lysines or the hinge thiols of cysteines, or antibody engineering can be performed for site-specific linking. The payload (C) is responsible for ADC toxicity and is usually a small hydrophobic molecule, able to cross cell membranes and cause cell death by targeting the cytoskeleton or DNA. Once cleaved from the antibody payloads can enter other (tumor) cells, resulting in further tumor killing (i.e. bystander effect) as well as off-target cytotoxicity when entering normal cells. The Fc region of the antibody (D) can trigger immune effector functions such as Antibody-Dependent Cytotoxicity through binding to Fcγ-receptors. However, if the ADC is internalized into non-malignant cells, it can cause off-target cytotoxicity. Antibody engineering can enhance or impair immune effector functions through, for example, single point mutations, Thiomabs, glycoengineering or incorporation of unnatural amino acids.