A plug-and-play approach to antibody-based therapeutics via a chemoselective dual click strategy.

Although recent methods for the engineering of antibody-drug conjugates (ADCs) have gone some way to addressing the challenging issues of ADC construction, significant hurdles still remain. There is clear demand for the construction of novel ADC platforms that offer greater stability, homogeneity and flexibility. Here we describe a significant step towards a platform for next-generation antibody-based therapeutics by providing constructs that combine site-specific modification, exceptional versatility and high stability, with retention of antibody binding and structure post-modification. The relevance of the work in a biological context is also demonstrated in a cytotoxicity assay and a cell internalization study with HER2-positive and -negative breast cancer cell lines

Dual modification of biomolecules

With the advent of novel bioorthogonal reactions and “click” chemistry, an increasing number of strategies for the single labelling of proteins and oligonucleotides have emerged. Whilst several methods exist for the site-selective introduction of a single chemical moiety, site-selective and bioorthogonal dual modification of biomolecules remains a challenge. The introduction of multiple modules enables a plethora of permutations and combinations and can generate a variety of bioconjuguates with many potential applications. From de novo approaches on oligomers to the post-translational functionalisation of proteins, this review will highlight the main strategies to dually modify biomolecules

Antibody fragments as nanoparticle targeting ligands: a step in the right direction

Recent advances in nanomedicine have shown that dramatic improvements in nanoparticle therapeutics and diagnostics can be achieved through the use of disease specific targeting ligands. Although immunoglobulins have successfully been employed for the generation of actively targeted nanoparticles, their use is often hampered by the suboptimal characteristics of the resulting complexes. Emerging data suggest that a switch in focus from full antibodies to antibody derived fragments could help to alleviate these problems and expand the potential of antibody–nanoparticle conjugates as biomedical tools. This review aims to highlight how antibody derived fragments have been utilised to overcome both fundamental and practical issues encountered during the design and application of antibody–targeted nanoparticles

A mild TCEP-based para-azidobenzyl cleavage strategy to transform reversible cysteine thiol labelling reagents into irreversible conjugates

It has recently emerged that the succinimide linkage of a maleimide thiol addition product is fragile, which is a major issue in fields where thiol functionalisation needs to be robust. Herein we deliver a strategy that generates selective cysteine thiol labelling reagents, which are stable to hydrolysis and thiol exchange

Recent advances in the construction of antibody–drug conjugates

Antibody-drug conjugates (ADCs) are derived from antibodies covalently attached to highly potent drugs using a variety of conjugation and linker technologies. This class of therapeutic conceptually combines the exquisite specificity of antibodies (i.e. enabling discrimination between healthy and diseased tissue) with the cell-killing ability of potent cytotoxic drugs. This powerful and exciting class of targeted therapy has shown considerable promise in the treatment of various cancers with two US Food and Drug Administration (FDA) approved ADCs currently on the market (i.e. Adcetris™ and Kadcyla™) and ca. 40 ADCs currently undergoing clinical evaluation. However, in order for ADCs to deliver their full potential, sophisticated site-specific conjugation technologies to connect the drug to the antibody are vital. This perspective discusses the strategies currently used for the site-specific construction of ADCs and appraises their merits and disadvantages

A Plug-and-Play Approach for the De Novo Generation of Dually Functionalized Bispecifics

Diseases are multifactorial, with redundancies and synergies between various pathways. However, most of the antibody-based therapeutics on the market interact with only one target, thus limiting their efficacy. The targeting of multiple epitopes could improve the therapeutic index of treatment and counteract mechanisms of resistance. To this effect, a new class of therapeutics has emerged: bispecific antibodies. Bispecific formation using chemical methods is rare and low-yielding and/or requires a large excess of one of the two proteins to avoid homodimerization and heterogeneity. In order for chemically prepared bispecifics to deliver their full potential, high-yielding, modular, and reliable cross-linking technologies are required. Herein, we describe a novel approach not only for the rapid and high-yielding chemical generation of bispecific antibodies from native antibody fragments, but also for the site-specific dual functionalization of the resulting bioconjugates. Based on orthogonal clickable functional groups, this strategy enables the assembly of functionalized bispecifics with controlled loading in a modular and convergent manner

A facile, one-pot procedure for the conversion of aromatic aldehydes to esters, as well as thioesters and amides, via acyl hydrazide intermediates

In the present work, an effective and facile one-pot dealloying strategy has been developed to synthesize monolithic asymmetry-patterned nanoporous copper ribbons (AP-NPCRs) from melt-spun bi-phase Al 32 at% Cu alloy with trace α-Al. The microstructure and nanoporosity of these AP-NPCRs were characterized using X-ray diffraction, scanning electron microscopy, energy dispersive X-ray analysis, transmission electron microscopy, high-resolution transmission electron microscopy, and Brunauer–Emmett–Teller measurements. The results show that the cooling rate and dealloying solution have a significant influence on formation, microstructure and nanoporosity of AP-NPCRs. The quenching surface of porous products has regular bimodal channel size distributions regardless of corrosive solution species, while the free surface shows a homogeneous porous network nanostructure in acidic solution and anomalous bimodal nanoporous architecture in alkaline medium. Additionally, the microstructure (surface morphology, ligament/channel sizes and distribution) and nanoporosity of AP-NPCRs can be modulated effectively by simply changing the dealloying solution

Use of pyridazinediones as extracellular cleavable linkers through reversible cysteine conjugation

Herein we report a retro-Michael deconjugation pathway of thiol-pyridazinedione linked protein bioconjugates to provide a novel cleavable linker technology. We demonstrate that the novel pyridazinedione linker does not suffer from off-target modification with blood thiols (e.g., glutathione, human serum albumin (HSA)), which is in sharp contrast to an analogous maleimide linker

Next-generation disulfide stapling: reduction and functional re-bridging all in one

Herein we present a significant step towards next-generation disulfide stapling reagents. A novel class of reagent has been designed to effect both disulfide reduction and functional re-bridging. The strategy has been applied to great success across various peptides and proteins. Moreover, application to a multi-disulfide system resulted in functional re-bridging without disulfide scrambling

A platform for efficient, thiol-stable conjugation to albumin's native single accessible cysteine

Herein we report the use of bromomaleimides for the construction of stable albumin conjugates via conjugation to its native, single accessible, cysteine followed by hydrolysis. Advantages over the classical maleimide approach are highlighted in terms of quantitative hydrolysis and absence of undesirable retro-Michael deconjugation