Tagged for thiolation: A novel glycoconjugation strategy utilizes a non‐natural olefin‐containing amino acid (homoallylglycine, Hag) as a “tag” for modification and a photoinitiated hydroglycothiolation reaction that is selective only for the Hag olefinic “tag”. Application of this method to a number of model proteins allowed complete and precise site‐selective glycosylation generating glycoconjugates that include, for example, virus‐like particles displaying up to 180 glycans at preselected positions (see scheme).

cMet is a well‐characterized oncogene that is the target of many drugs including small molecule and biologic pathway inhibitors, and, more recently, antibody–drug conjugates (ADCs). However, the clinical benefit from cMet‐targeted therapy has been limited. We developed a novel cMet‐targeted ‘third‐generation’ ADC, TR1801‐ADC, that was optimized at different levels including specificity, stability, toxin–linker, conjugation site, and in vivo efficacy. Our nonagonistic cMet antibody was site‐specifically conjugated to the pyrrolobenzodiazepine (PBD) toxin–linker tesirine and has picomolar activity in cancer cell lines derived from different solid tumors including lung, colorectal, and gastric cancers. The potency of our cMet ADC is independent of MET gene copy number, and its antitumor activity was high not only in high cMet‐expressing cell lines but also in medium‐to‐low cMet cell lines (40 000–90 000 cMet/cell) in which a cMet ADC with tubulin inhibitor payload was considerably less potent. In vivo xenografts with low–medium cMet expression were also very responsive to TR1801‐ADC at a single dose, while a cMet ADC using a tubulin inhibitor showed a substantially reduced efficacy. Furthermore, TR1801‐ADC had excellent efficacy with significant antitumor activity in 90% of tested patient‐derived xenograft models of gastric, colorectal, and head and neck cancers: 7 of 10 gastric models, 4 of 10 colorectal cancer models, and 3 of 10 head and neck cancer models showed complete tumor regression after a single‐dose administration. Altogether, TR1801‐ADC is a new generation cMet ADC with best‐in‐class preclinical efficacy and good tolerability in rats.

The lipid carrier specificity of the protein N-glycosylation enzyme C. jejuni PglB was tested using a logical, synthetic array of natural and unnatural C10, C20, C30, and C40 polyisoprenol sugar pyrophosphates, including those bearing repeating cis-prenyl units. Unusual, short, synthetically accessible C20 prenols (nerylnerol 1d and geranylnerol 1e) were shown to be effective lipid carriers for PglB sugar substrates. Kinetic analyses for PglB revealed clear K(M)-only modulation with lipid chain length, thereby implicating successful in vitro application at appropriate concentrations. This was confirmed by optimized, efficient in vitro synthesis allowing >90% of Asn-linked β-N-GlcNAc-ylated peptide and proteins. This reveals a simple, flexible biocatalytic method for glycoconjugate synthesis using PglB N-glycosylation machinery and varied chemically synthesized glycosylation donor precursors.

Unnatural, NMR- and MRI-active fluorinated sugar probes, designed and synthesised to bind to the pathogenic protein TgMIC1 from Toxoplasma gondii, were found to display binding potency equal to and above that of the natural ligand. Dissection of the binding mechanism and modes, including the first X-ray crystal structures of a fluoro-oligosaccharide bound to a lectin, demonstrate that it is possible to create effective fluorinated probe ligands for the study of, and perhaps intervention in, sugar-protein binding events.