Bioconjugation with Maleimides: A Useful Tool for Chemical Biology

Abstract

Maleimide chemistry stands out in the bioconjugation toolbox by virtue of its synthetic accessibility, excellent reactivity, and practicability. The second-generation of clinically approved antibody-drug conjugates (ADC) and much of the current ADC pipeline in clinical trials contain the maleimide linkage. However, thiosuccinimide linkages are now known to be less robust than once thought, and ergo, are correlated with suboptimal pharmacodynamics, pharmacokinetics, and safety profiles in some ADC constructs. Rational design of novel generations of maleimides and maleimide-type reagents have been reported to address the shortcomings of classical maleimides, allowing for the formation of robust bioconjugate linkages. This review highlights the main strategies for rational reagent design that have allowed irreversible bioconjugations in cysteines, reversible labelling strategies and disulfide re-bridging.

N‐Hydroxysuccinimide‐Modified Ethynylphosphonamidates Enable the Synthesis of Configurationally Defined Protein Conjugates

“…Despite being the most widely used cysteine conjugation method, maleimide chemistry is associated with several drawbacks . First, maleimide conjugates are prone to hydrolysis, forming the ring‐opened maleamic acid moiety, which is unreactive to sulfhydryl groups (Scheme ).…”

“…[5] Despite being the most widely used cysteine conjugation method, maleimide chemistry is associated with several drawbacks. [6] First, maleimide conjugates are prone to hydrolysis, forming the ring-opened maleamic acid moiety, which is unreactive to sulfhydryl groups (Scheme 1). This is especially problematic in the context of hetero-bifunctional crosslinking reagents because maleimides that hydrolyze during the installation step on the protein by NHS chemistry can diminish the overall conjugation yield for the final conjugate.…”

Synthesis and Characterization of an Epidermal Growth Factor Receptor‐Selective RuII Polypyridyl–Nanobody Conjugate as a Photosensitizer for Photodynamic Therapy

“…[Ru(phen) 2 (dppz‐7‐maleimidemethyl)](PF 6 ) 2 was coupled to the poly‐glycine chain via a thio‐Michael addition reaction. As recently highlighted, this bioconjugation presents important advantages such as synthetic accessibility, excellent reactivity and, importantly, biocompatibility . Following this synthetic strategy, the thiosuccinimide product [Ru(phen) 2 (dppz‐7‐maleimidemethyl‐S‐Cys‐(Ser) 2 ‐(Gly) 5 ‐NH 3 )] 3+ was prepared by treating the thiol of the (NH 3 ‐(Gly) 5 ‐(Ser) 2 ‐Cys‐CONH 2 )(TFA) peptide chain with the [Ru(phen) 2 (dppz‐7‐maleimidemethyl)](PF 6 ) 2 complex.…”

Stable Pyrrole‐Linked Bioconjugates through Tetrazine‐Triggered Azanorbornadiene Fragmentation

“…Of all proteinogenic amino acids, the sulfhydryl side‐chain of cysteine remains the primary choice to achieve site selectivity because of its enhanced nucleophilicity and low abundance in its reduced form . Although many cysteine bioconjugation strategies have been developed, the thio‐Michael addition reaction to a maleimide acceptor is the most popular for reactions directed at cysteine because it is fast and quantitative at near‐neutral pH, and because the reagents used are commercially available with a maleimide handle for conjugation . The use of maleimides is best illustrated in the generation of clinically used ADCs, such as brentuximab vedotin and ado‐trastuzumab emtansine .…”