Fig. 3
Effects of MDM2 inhibition on SETD2 expression and function. Co-immunoprecipitation assays showed that (A) p53 interacts with SETD2 in the SETD2-proficient ROSAKIT D816V cell line and, after bortezomib treatment, in SETD2-deficient HMC-1 cells, and that (B) ubiquitinated SETD2 binds MDM2 in the same context. C and D Silencing or inhibiting MDM2 by siRNA (for 72 h) or by SP141 (5 µM for 24 h), respectively, rescued SETD2/H3K36Me3 expression in HMC-1 cells. A negative control (NC) obtained by the immunoprecipitation of protein lysates using a resin conjugated with an anti-IgG1 antibody was loaded for each immunoprecipitation experiment. E SP-141 administration in HMC-1 cells (red: 1 µM; green: µM) maintained in liquid medium and counted every 24 h up to 96 h of treatment showed cytostatic effects. F Clonogenic assays performed in semisolid medium using scalar doses of the drug (0.075–0.3 µM) supported the cytostatic effects of SP141 in HMC-1.1 and -1.2 cell lines