We report a quantitative proteomics data analysis pipeline, which coupled to protein‐directed dynamic combinatorial chemistry (DDC) experiments, enables the rapid discovery and direct characterization of protein‐protein interaction (PPI) modulators. A low‐affinity PD‐1 binder was incubated with a library of >100 D‐peptides under thiol‐exchange favoring conditions, in the presence of the target protein PD‐1, and we determined the S‐linked dimeric species that resulted, amplified in the protein samples versus the controls. We chemically synthesized the target dimer candidates and validated them by thermophoresis binding and protein‐protein interaction assays. The results provide a proof‐of‐concept for using this strategy in the high‐throughput search of improved drug‐like peptide binders that block therapeutically relevant protein‐protein interactions. A protein‐directed dynamic combinatorial chemistry (DCC) method, coupled to a proteomics data analysis workflow, was used to discover interface blockers of the PD‐1/PD‐L1 interaction. The S‐linked dimeric species that were significantly amplified in the protein‐templated mixture were identified, synthesized, and subjected to biophysical screening.