Structure and activity of a novel DUB inhibitor. (A) Chemical structures of WP1130 and G9. (B) MM1.S cells were treated as described with 5 µM G9 before assessing DUB activity (top) and total protein expression of Usp9x, Usp24, Mcl-1, PARP, and β-actin. The HA-labeled Usp9x/Usp24 band is shown at the top. (C) MM1.S cells were treated with the G9 concentration indicated before assessing changes in DUB activity associated with Usp9x/Usp24 and Usp5. Lysates were also immunoblotted for Mcl-1, p53, and cleaved PARP as a measure of apoptosis. Actin was blotted as a loading control. (D) Cells derived from a plasma cell leukemia (PCL) patient were treated with G9 for the time indicated before DUB activity and specific protein levels were determined by HA-UbVS labeling and immunoblotting.