On‐the‐Spot Immobilization of Quantum Dots,Graphene Oxide,and Proteins via Hydrophobins

Class I hydrophobin Vmh2, a peculiar surface active and versatile fungal protein, is known to self‐assemble into chemically stable amphiphilic films, to be able to change wettability of surfaces, and to strongly adsorb other proteins. Herein, a fast, highly homogeneous and efficient glass functionalization by spontaneous self‐assembling of Vmh2 at liquid–solid interfaces is achieved (in 2 min). The Vmh2‐coated glass slides are proven to immobilize not only proteins but also nanomaterials such as graphene oxide (GO) and quantum dots (QDs). As models, bovine serum albumin labeled with Alexa 555 fluorophore, anti‐immunoglobulin G antibodies, and cadmium telluride QDs are patterned in a microarray fashion in order to demonstrate functionality, reproducibility, and versatility of the proposed substrate. Additionally, a GO layer is effectively and homogeneously self‐assembled onto the studied functionalized surface. This approach offers a quick and simple alternative to immobilize nanomaterials and proteins, which is appealing for new bioanalytical and nanobioenabled applications.     

Discovery of Affinity-Based Probes for Btk Occupancy Assays

Bruton's tyrosine kinase (Btk) is an attractive target for the treatment of a wide array of B-cell malignancies and autoimmune diseases. Small-molecule covalent irreversible Btk inhibitors targeting Cys481 have been developed for the treatment of such diseases. In clinical trials, probe molecules are required in occupancy studies to measure the level of engagement of the protein by these covalent irreversible inhibitors. The result of this pharmacodynamic (PD) activity provides guidance for appropriate dosage selection to optimize inhibition of the drug target and correlation of target inhibition with disease treatment efficacy. This information is crucial for successful evaluation of drug candidates in clinical trials. Based on the pyridine carboxamide scaffold of a novel solvent-accessible pocket (SAP) series of covalent irreversible Btk inhibitors, we successfully developed a potent and selective affinity-based biotinylated probe 12 (2-[(4-{4-[5-(1-{5-[(3aS,4S,6aR)-2-oxo-hexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanamido}-3,6,9,12-tetraoxapentadecan-15-amido)pentanoyl]piperazine-1-carbonyl}phenyl)amino]-6-[1-(prop-2-enoyl)piperidin-4-yl]pyridine-3-carboxamide). Compound 12 has been used in Btk occupancy assays for preclinical studies to determine the therapeutic efficacy of Btk inhibition in two mouse lupus models driven by TLR7 activation and type I interferon.