Characterization of the p53-rescue drug CP-31398 in vitro and in living cells

Abstract

The Pfizer compound CP-31398 has been reported to stabilize the core domain of the tumour suppressor p53 in vitro and be an effective anti-cancer drug by virtue of rescuing destabilized mutants of p53. We did not detect any interaction between the p53 core domain and CP-31398 in vitro by a wide range of quantitative biophysical techniques over a wide range of conditions. CP-31398 did not stabilize p53 in our experiments. However, we found that CP-31398 intercalated with DNA and also altered and destabilized the DNA-p53 core domain complex. We analysed by NMR TROSY the interaction of the domain with a DNA oligomer and identified the changes in the complex on the binding of CP-31398. CP-31398 also decreased sequence-specific DNA binding of wild-type p53 and His-273 mutant p53. CP-31398 had a non-specific toxic effect independent of mutant p53 expression in several cell lines carrying Tet-regulated mutant p53. CP-31398 caused a small increase in MDM-2 expression and a more pronounced p53-independent increase in Bax expression. CP-31398 did, however, induce the PAb1620 epitope (characteristic of native p53) in cells expressing His-175 mutant p53. This was prevented by cycloheximide, suggesting that any stabilizing action of CP-31398 would have to be on newly synthesized p53. One of the unstable mutants that was reported to have been rescued by CP-31398, R249S, does not bind DNA when folded at lower temperatures.