Identifying LC3 binders to develop lysosome-targeting chimeras

National Center For Advancing Translational Sciences

Investigators

Abstract

During this reporting period, the project team has implemented a high-throughput screening assay to detect LC3 binders utilizing a reporter construct that monitors the degradation of cargo destined to the lysosome. Virtual screening was performed to identify small molecules that fit into the LC3-interacting region (LIR)-binding site within LC3, which is important for recruiting cargo to autophagosomes. Compounds identified in the high-throughput screen were prioritized for testing in orthogonal assays to examine direct target engagement. Fluorescence polarization, microscale thermophoresis, and cellular thermal shift assays were developed to interrogate direct binding of the molecule to LC3.