High-throughput screening based on label-free detection of small molecule microarrays

Based on small-molecule microarrays (SMMs) and oblique-incidence reflectivity difference (OI-RD) scanner, we have developed a novel high-throughput drug preliminary screening platform based on label-free monitoring of direct interactions between target proteins and immobilized small molecules. The screening platform is especially attractive for screening compounds against targets of unknown function and/or structure that are not compatible with functional assay development. In this screening platform, OI-RD scanner serves as a label-free detection instrument which is able to monitor about 15,000 biomolecular interactions in a single experiment without the need to label any biomolecule. Besides, SMMs serves as a novel format for high-throughput screening by immobilization of tens of thousands of different compounds on a single phenyl-isocyanate functionalized glass slide. Based on the high-throughput screening platform, we sequentially screened five target proteins (purified target proteins or cell lysate containing target protein) in high-throughput and label-free mode. We found hits for respective target protein and the inhibition effects for some hits were confirmed by following functional assays. Compared to traditional high-throughput screening assay, the novel high-throughput screening platform has many advantages, including minimal sample consumption, minimal distortion of interactions through label-free detection, multi-target screening analysis, which has a great potential to be a complementary screening platform in the field of drug discovery.

[1]  Stuart L Schreiber,et al.  A robust small-molecule microarray platform for screening cell lysates. , 2006, Chemistry & biology.

[2]  I. Shin,et al.  Fabrication of chemical microarrays by efficient immobilization of hydrazide-linked substances on epoxide-coated glass surfaces. , 2005, Angewandte Chemie.

[3]  Lan Mi,et al.  Calibration of oblique-incidence reflectivity difference for label-free detection of a molecular layer. , 2016, Applied optics.

[4]  Xiangdong Zhu,et al.  Discovering small molecule ligands of vascular endothelial growth factor that block VEGF-KDR binding using label-free microarray-based assays. , 2013, Assay and drug development technologies.

[5]  A. Koehler,et al.  Ligand discovery using small-molecule microarrays. , 2012, Methods in molecular biology.

[6]  Yiyan Fei,et al.  Characterization of protein expression levels with label-free detected reverse phase protein arrays. , 2016, Analytical biochemistry.

[7]  Lan Mi,et al.  Developing an Efficient and General Strategy for Immobilization of Small Molecules onto Microarrays Using Isocyanate Chemistry , 2016, Sensors.

[8]  Angela N Koehler,et al.  A method for the covalent capture and screening of diverse small molecules in a microarray format , 2006, Nature Protocols.

[9]  K. Lam,et al.  A novel high-throughput scanning microscope for label-free detection of protein and small-molecule chemical microarrays. , 2008, The Review of scientific instruments.

[10]  Y.S. Sun,et al.  Effect of fluorescently labeling protein probes on kinetics of protein-ligand reactions , 2008, 2008 Conference on Lasers and Electro-Optics and 2008 Conference on Quantum Electronics and Laser Science.

[11]  K. Lam,et al.  Oblique-incidence reflectivity difference microscope for label-free high-throughput detection of biochemical reactions in a microarray format. , 2006, Applied optics.

[12]  M. Lesaicherre,et al.  Developing site-specific immobilization strategies of peptides in a microarray. , 2002, Bioorganic & medicinal chemistry letters.