Reactivation of mutant p53 by SLMP53-2: a promising therapeutic option against hepatocellular carcinoma

In order to pursue the search for new reactivators of mutant p53, a small library of molecules was synthesized starting from a previously established lead (SLMP53-1). The molecules were tested in various cell-based assays and one, herein referred to as SLMP53-2, displayed a marked reduction of the IC50 values in NCI-H1299 cells expressing the p53 mutants (mutp53) R175H, Y220C, G245S, R248Q, R273C, and R273H. The activity of SLMP53-2 was then investigated using cancer cells lines expressing endogenous mutant p53 and focusing on hepatocellular carcinoma HuH-7 and breast ductal carcinoma HCC1419 cells endogenously expressing mutp53-Y220C. In HuH-7 cells, SLMP53-2 displayed a concentration-dependent growth inhibitory effect on colony formation, restored wt-like p53 protein conformation and transcriptional activity and displayed in vivo antitumour activity in a xenograft mouse model, with no apparent toxicity for normal tissues. To investigate more deeply the impact of SLMP53-2 treatments on gene expression in HuH-7 cells we performed microarray-based analysis using two different concentrations of the compound, corresponding to 2x and 3x IC50. We also employed the MDM2 inhibitor Nutlin alone or in combination with 2x IC50 SLMP53-2. The treatment were: 2x and 3x IC50 SLMP53-2 (28 and 42 µM), for 24 hours, in quadruplicate. Cells were processed for RNA extraction and gene expression was assessed by Agilent gene expression microarrays. Direct comparisons were performed in order to obtain differential gene expression gene lists.