All-in-one disulfide bridging enables the generation of antibody conjugates with modular cargo loading.

Antibody-drug conjugates (ADCs) are valuable therapeutic entities which leverage the specificity of antibodies to selectively deliver cytotoxins to antigen-expressing targets such as cancer cells. However, current methods for their construction still suffer from a number of shortcomings. For instance, using a single modification technology to modulate the drug-to-antibody ratio (DAR) in integer increments while maintaining homogeneity and stability remains exceptionally challenging. Herein, we report a novel method for the generation of antibody conjugates with modular cargo loading from native antibodies. Our approach relies on a new class of disulfide rebridging linkers, which can react with eight cysteine residues, thereby effecting all-in-one bridging of all four interchain disulfides in an IgG1 antibody with a single linker molecule. Modification of the antibody with the linker in a 1 : 1 ratio enabled the modulation of cargo loading in a quick and selective manner through derivatization of the linker with varying numbers of payload attachment handles to allow for attachment of either 1, 2, 3 or 4 payloads (fluorescent dyes or cytotoxins). Assessment of the biological activity of these conjugates demonstrated their exceptional stability in human plasma and utility for cell-selective cytotoxin delivery or imaging/diagnostic applications.

Divinylpyrimidine reagents generate antibody-drug conjugates with excellent in vivo efficacy and tolerability.

The development of divinylpyrimidine (DVP) reagents for the synthesis of antibody-drug conjugates (ADCs) with in vivo efficacy and tolerability is reported. Detailed structural characterisation of the synthesised ADCs was first conducted followed by in vitro and in vivo evaluation of the ADCs' ability to safely and selectively eradicate target-positive tumours.

Site-selective modification strategies in antibody-drug conjugates.

Antibody-drug conjugates (ADCs) harness the highly specific targeting capabilities of an antibody to deliver a cytotoxic payload to specific cell types. They have garnered widespread interest in drug discovery, particularly in oncology, as discrimination between healthy and malignant tissues or cells can be achieved. Nine ADCs have received approval from the US Food and Drug Administration and more than 80 others are currently undergoing clinical investigations for a range of solid tumours and haematological malignancies. Extensive research over the past decade has highlighted the critical nature of the linkage strategy adopted to attach the payload to the antibody. Whilst early generation ADCs were primarily synthesised as heterogeneous mixtures, these were found to have sub-optimal pharmacokinetics, stability, tolerability and/or efficacy. Efforts have now shifted towards generating homogeneous constructs with precise drug loading and predetermined, controlled sites of attachment. Homogeneous ADCs have repeatedly demonstrated superior overall pharmacological profiles compared to their heterogeneous counterparts. A wide range of methods have been developed in the pursuit of homogeneity, comprising chemical or enzymatic methods or a combination thereof to afford precise modification of specific amino acid or sugar residues. In this review, we discuss advances in chemical and enzymatic methods for site-specific antibody modification that result in the generation of homogeneous ADCs.