CP-31398 restores DNA-binding activity to mutant p53 in vitro but does not affect p53 homologs p63 and p73.
The p53 protein plays a major role in the maintenance of genome stability in mammalian cells. Mutations of p53 occur in over 50% of all cancers and are indicative of highly aggressive cancers that are hard to treat. Recently, there has been a high degree of interest in therapeutic approaches to restore growth suppression functions to mutant p53. Several compounds have been reported to restore wild type function to mutant p53. One such compound, CP-31398, has been shown effective in vivo, but questions have arisen to whether it actually affects p53. Here we show that mutant p53, isolated from cells treated with CP-31398, is capable of binding to p53 response elements in vitro. We also show the compound restores DNA-binding activity to mutant p53 in cells as determined by a chromatin immunoprecipitation assay. In addition, using purified p53 core domain from two different hotspot mutants (R273H and R249S), we show that CP-31398 can restore DNA-binding activity in a dose-dependent manner. Using a quantitative DNA binding assay, we also show that CP-31398 increases significantly the amount of mutant p53 that binds to cognate DNA (B(max)) and its affinity (K(d)) for DNA. The compound, however, does not affect the affinity (K(d) value) of wild type p53 for DNA and only increases B(max) slightly. In a similar assay PRIMA1 does not have any effect on p53 core DNA-binding activity. We also show that CP-31398 had no effect on the DNA-binding activity of p53 homologs p63 and p73.
Schering-Plough Research Institute, Kenilworth, New Jersey 07033, USA.
Mark J
MJ
Demma
Serena
S
Wong
Eugene
E
Maxwell
Bimalendu
B
Dasmahapatra
D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D016147:Genes, Tumor Suppressor; D006801:Humans; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D011743:Pyrimidines / Q000494:pharmacology*; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D025521:Tumor Suppressor Proteins
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D011743:Pyrimidines; C115051:TP63 protein, human; C000606460:TP73 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D004247:DNA; C402665:CP 31398