Fig. 4. VHL-based PROTAC 34 of the ‘phenoxy’ subseries (A) induces CDK6-selective degradation in a dose-dependent manner (B). MM.1S cells were treated with 1 μM of the VHL ligand VH298, BSJ-03-123 (8), negative control 32, or PROTAC 34 at the indicated concentrations for 16 h. (C) Quantification of (B) and calculation of the DC₅₀-value. The value for 1 μM was excluded due to a slight hook effect. Mechanistic characterization of 34; the activity is CRL2^VHL- and proteasome-dependent (D). Co-treatment of 34 and VH298 (100-fold excess) abrogated CDK4/6 degradation. MG132 and MLN4924 prevented proteasomal degradation of CDK4/6. MM.1S cells were treated with MG132, MLN4924 or VH298 in the presence or absence of CDK4/6d 34 (0.1 μM). (E) Persisting effects of PROTAC 34 on CDK4/6 degradation after drug washout. MM.1S cells were treated with PROTACs at 0.1 μM for 16 h before washout with PBS (=0 h), then kept in plain media until indicated time points.