Exploring drug mechanisms through single‐cell proteomics

Drug discovery requires precise mechanistic insights, hindered by traditional bulk omics masking cellular heterogeneity. Single-cell proteomics (SCP) overcomes this by directly quantifying proteins-the cell's functional executors-enabling high-resolution analysis of drug action. This review details how SCP shifts research from the transcriptome to the proteome, covering key methods like high-sensitivity mass spectrometry and data-independent acquisition that now quantify thousands of proteins per cell. We highlight SCP's unique advantages in directly capturing drug targets, modifications, and pathway activity. Translational applications demonstrate its potential in target discovery, elucidation of resistance mechanisms and precision therapy. Finally, the integration of AI with SCP is poised to usher in a new era of protein-centred, high-resolution drug discovery.