First, we conducted in vitro ClpP activation assay and pull-down assay using SA-conjugated beads to confirm that compound 1 is capable of ClpP activation and can form a [SA-1-ClpP] ternary complex (Figure ). The results validated our molecular design. Next, in vitro mSA degradation assay revealed that compound 1 decreased mSA in a dose-dependent manner (DC50,24h = 197 nM under the conditions of 50 nM ClpP, 2.5 µM mSA) (Figure ). Excess desthiobiotin inhibited the mSA degradation activity of 1, suggesting that the formation of the [mSA-1-ClpP] ternary complex is required for the activity of 1 (Figure ). Whereas ClpP activators promiscuously enhance protein degradation, chimeric compound 1 is expected to mediate selective degradation of the target protein. Indeed, in cell lysates supplemented with mSA and ClpP, compound 1 led to a decrease of mSA, while other proteins appeared to be unaffected as indicated by CBB staining (Figure ).