The resulting library of 22 fragments was screened for BRD4(1) L94C binding, at a concentration of 50 μM, using a protein LCMS assay (referred to as 'labeling assay 1'). Fragments which showed a mass corresponding to the protein plus bound fragment, with a peak intensity of >2%, were classified as 'hits'. To assess the selectivity of the fragments for labeling of C94, hits were also screened against BRD4(1) WT under the same conditions. In this screen, two 3-methylisoxazole-based fragments, 12 and 14, were observed to label BRD4(1) L94C (Table and Figure ). The acrylamide-based fragment, 12, labeled 97.5% of BRD4(1) L94C after 4 hours. While 94.9% of the protein was labeled once, 2.6% of the protein was labeled twice (Figure ). This fragment showed good selectivity over BRD4(1) WT , with a single labeling event of 3% intensity observed after 4 hours (Figure ). Fragment 14, designed by appending a β-dimethylaminomethyl acrylamide to the structure of 12, led to 74% of singly labeled BRD4(1) L94C after 4 hours, with no labeling of BRD4(1) WT observed (Table and Figure ). Interestingly, the corresponding fragments lacking the phenyl ring at the 4-position showed a lower percentage of BRD4(1) L94C labeling (56% and 52%, for 10 and 13, respectively), suggesting that the addition of the phenyl group increases the fragments affinity for BRD4(1) L94C or induces a conformation that allows for more favorable reactivity between the electrophile and C94.