Folate-Targeted Monodisperse PEG-Based Conjugates Made by Chemo-Enzymatic Methods for Cancer Diagnosis and Treatment

This paper focuses on preliminary in vitro and in vivo testing of new bivalent folate-targeted PEGylated doxorubicin (DOX) made by modular chemo-enzymatic processes (FA₂-dPEG-DOX₂). A unique feature is the use of monodisperse PEG (dPEG). The modular approach with enzyme catalysis ensures exclusive γ-conjugation of folic acid, full conversion and selectivity, and no metal catalyst residues. Flow cytometry analysis showed that at 10 µM concentration, both free DOX and FA₂-dPEG-DOX₂ would be taken up by 99.9% of triple-negative breast cancer cells in 2 h. Intratumoral injection to mice seemed to delay tumor growth more than intravenous delivery. The mouse health status, food, water consumption, and behavior remained unchanged during the observation.     

DNA as a Platform to Program Assemblies with Emerging Functions in Chemical Biology

The programmability of oligonucleotide recognition offers an attractive platform for directing the assembly of ligands that can interact cooperatively with a target or the assembly of reactive partner that can engage in chemical reactions. In both cases, an oligonucleotide directs the assembly, which yields a functional output. In terms of the controlled display of ligands, several examples have shown a significant increase in binding upon ligand assembly using small molecule fragments, peptides, glycans, and protein fragments. Combinatorial approaches to identifying the optimal pairing have also been reported. In terms of nucleic acid triggered reactions, several robust chemistries have been reported, leading to the unmasking of different fluorophores or bioactive molecules as well as the synthesis of a bioactive compound in response to DNA or even cellular RNA. Herein we present the historical context of this work and summarize recent developments in this area.     

Multivalent carbohydrate recognition on a glycodendrimer-functionalized flow-through chip

Dendrimers were fitted out with up to eight mannose moieties by "click" chemistry. They were subsequently attached to aluminum oxide chips via a spacer that was linked to the dendrimer core; this resulted in a microarray of glycodendrimers. Binding of the glycodendrimers to the fluorescent lectins ConA and GNA was observable in real time. In a single experiment it was possible to observe the multivalency enhancement or cluster effect in the binding event. This effect was small for ConA, in agreement with its widely spaced binding sites, whereas it was large for GNA, with its twelve much more closely spaced binding sites. The dendrimer-fitted chip represents a valuable screening tool for multivalency effects. Furthermore kinetic and thermodynamic data on binding events can be deduced. Inhibition experiments are also possible with the system as was shown for ConA with alpha-methyl mannose as the inhibitor.     

Reactivity Analysis of New Multivalent Electrophilic Probes for Affinity Labeling of Carbohydrate Binding Proteins

We have designed and synthesized six different multivalent electrophiles as carbohydrate affinity labeling probes. Evaluation of the reactivity of the electrophiles against peanut agglutinin (PNA) and Ricinus communis agglutinin (RCA) showed that p- and m-aryl sulfonyl fluoride are effective protein reactive groups that label carbohydrate binding lectins in a ligand-dependent fashion at a nanomolar probe concentration. Analysis of the selectivity of affinity labeling in the presence of excess BSA as a nonspecific protein indicated that m-arylsulfonyl fluoride is a more selective protein-reactive group, albeit with attenuated reactivity. Further analysis showed that the labeling efficiency of the multivalent electrophilic probes can be improved by employing reaction conditions involving 25 °C instead of typically employed 4 °C. Both isomers of arylsulfonyl fluoride groups together represent promising affinity labels for target identification studies that could serve as more efficient alternatives to photoreactive groups.     

Recruitment of Receptors and Ligands in a Weakly Multivalent System with Omnipresent Signatures of Superselective Binding (Small 23/2023)

Recruitment of receptors at membrane interfaces is essential in biological recognition and uptake processes. The interactions that induce recruitment are typically weak at the level of individual interaction pairs, but are strong and selective at the level of recruited ensembles. Here, a model system is demonstrated, based on the supported lipid bilayer (SLB) that mimics the recruitment process induced by weakly multivalent interactions. The weak (mm range) histidine-nickel-nitrilotriacetate (His₂-NiNTA) pair is employed owing to its ease of implementation in both synthetic and biological systems. The recruitment of receptors (and ligands) induced by the binding of His₂-functionalized vesicles on NiNTA-terminated SLBs is investigated to identify the ligand densities necessary to achieve vesicle binding and receptor recruitment. Threshold values of ligand densities appear to occur in many binding characteristics: density of bound vesicles, size and receptor density of the contact area, and vesicle deformation. Such thresholds contrast the binding of strongly multivalent systems and constitute a clear signature of the superselective binding behavior predicted for weakly multivalent interactions. This model system provides quantitative insight into the binding valency and effects of competing energetic forces, such as deformation, depletion, and entropy cost of recruitment at different length scales.