Homo-PROTAC cereblon degrader 1

Homo-PROTAC cereblon degrader 1 (OUN20985), a highly potent and efficient Cereblon (CRBN) degrader with only minimal effects on pomalidomide neosubstrates IKZF1 and IKZF3, has no effects on proliferation of different cell lines, prevents pomalidomide-induced degradation of IKZF1 and IKZF3, and antagonizes the effects of pomalidomide on multiple myeloma cellslidomide resistance in different multiple myeloma cells.^[1]

• Cell lines

  Multiple myeloma cell line NCI-H929, MM1S, HEK293T

• Concentrations

  0.1 µM

• Incubation Time

  3 h

• Method

  HEK293T cells were transfected with a FLAG-CRBN-expressing plasmid. Forty-eight hours after transfection, cells were treated with DMSO or compound 15a for 3 h. Cells were then washed twice with ice-cold phosphate-buffered saline (PBS), lysed under denaturing conditions using 2% sodium dodecyl sulfate  (SDS)-containing lysis buffer, and boiled for 10 min at 95 ℃. The SDS was diluted via the addition of 10×IP lysis buffer (Pierce) containing HALT Protease and Phosphatase Inhibitor Cocktail, MG132 (10 μM), and PR619 (50 nM) and incubated at 4 ℃ for 30 min. Immunoprecipitation was performed using anti-FLAG M2 Affinity Gel according to the manufacturer's protocol, and protein was eluted with FLAG peptide. CRBN was detected by a FLAG-specific antibody, and ubiquitinated CRBN by the FK2 antibody.