Development of A Image-based High-content Screening Assay Based on Clickable Probes for Identification of XPB Inhibitors

ABSTRACT: High-content screening (HCS) has become a powerful tool in drug discovery; however, their reliance on indirect readouts and surrogate markers limits their ability to directly assess drug-protein interactions at endogenous levels, particularly in subcellular contexts. Here, we report the development of a confocal image-based HCS platform that addresses these limitations by combining with bio-orthogonal labelling. We synthesized a clickable probe TL-alkyne that rapidly and specifically labels xeroderma pigmentosum type B (XPB), a critical protein in the nucleotide excision repair (NER). The colorimetric HCS assay enabled direct and precise measurement of drug occupancy rates in XPB specifically within the nucleus of live cells. Out of 1,953 FDA-approved drugs, pelitinib was identified as a novel ligand to bind XPB. It formed a covalent bond with Cys342 of XPB, suppressed XPB's ATPase activity, impaired nucleotide excision repair, and synergistically enhanced chemotherapy. This study demonstrates the transformative potential of integrating click chemistry with advanced HCS technologies, offering a robust framework for high-throughput drug discovery targeting challenging protein systems

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Cervix Carcinoma

SUBMITTER: Yang Yang  

LAB HEAD: Qian Zhao

PROVIDER: PXD059271 | Pride | 2025-09-08

REPOSITORIES: Pride