The Mutant p53-Conformation Modifying Drug, CP-31398, Can Induce Apoptosis

Abstract

CP-31398, a styrylquinazoline, emerged from a screen for therapeutic agents that restore a wild-type DNA-binding conformation of mutant p53 to suppress tumors in-vivo (Science 286, 2507, 1999). We investigated the growth inhibitory mechanism of CP-31398 using nine human cancer cell lines containing wild-type, mutant or no p53 expression. Six of nine cell lines underwent apoptosis after exposure to CP-31398, while two cell lines, DLD1 colon cancer and H460 lung cancer, underwent exclusively cell cycle arrest. Cell cycle arrest preceded the apoptosis in some cases. CP-31398 did not inhibit growth of the p53 non-expressing ovarian cancer cell line SKOV3. Interestingly, we found that wild-type p53 protein is stabilized upon CP-31398 exposure. p53 target genes such as p21WAF1/Cip1, and KILLER/DR5 were upregulated by CP-31398, but their expression did not correlate with arrest or apoptosis induction. Combination of CP-31398 and TRAIL or chemotherapeutic agents enhanced cancer cell killing effect possibly through upregulation of p53-regulated genes such as KILLER/DR5. Bax-/-, wild-type p53-expressing cells displayed reduced susceptibility to killing by CP-31398. An Affymetrix GeneChip Array screen revealed that CP-31398 alters expression of non-p53 target genes in addition to p53-responsive genes. Although our preliminary data suggest that CP-31398 does not alter wild-type p53:MDM2 interaction, further efforts are required to elucidate the mechanism of wild-type p53 stabilization by CP-31398. The results increase our understanding of CP-31398 action, and suggest strategies for improving its specificity, possibly through use of microarrays to screen related compounds with higher mutant p53-specificity.

CP-31398, a novel p53-stabilizing agent, induces p53-dependent and p53-independent glioma cell death

“…However, U251MG and U373MG cells nevertheless showed a strong accumulation of p21 when treated with CP-31398 (Figure 3), suggesting that the reporter assay poorly reflects the transcriptional activation, at least of the p21 gene. In this regard, gene chip analysis revealed that most, but not all, p53 response genes are induced when mutant p53 is stabilized through CP-31398 (Takimoto, 2002). Further, in analogy to our protein data (Figure 3), these authors observed a repression of mdm-2 mRNA expression by high concentrations of CP-31398.…”

“…This, however, is the mode of action previously proposed for CP-31398 (Foster et al, 1999). Further support for a direct interaction of CP-31398 and p53 comes from EMSA studies showing restoration of the sequence-specific DNA-binding ability of the 273 (Arg-His) p53 mutant by CP-31398 (Takimoto et al, 2002).…”

Reactivation of mutant p53: molecular mechanisms and therapeutic potential

“…NMR studies failed to detect any binding of CP-31398 to the p53 core domain . Interestingly, CP-31398 increases the levels of wild-type p53 protein in cells by preventing its ubiquitination independently of Mdm-2 and p53 phosphorylation (Luu et al, 2002;Takimoto et al, 2002;Wang et al, 2003). CP-31398 affects gene expression and induces cell death both in a p53-dependent and -independent manner (Takimoto et al, 2002;Wischhusen et al, 2003;Wang et al, 2003).…”

“…This idea has been confirmed in vitro and in vivo in several studies. Synergy has been observed between CP31398 and adriamycin or cisplatin (Takimoto et al, 2002) and between PRIMA-1MET and adriamycin, cisplatin or fludarabine (Nahi et al, 2004;Bykov et al, 2005a). The synergy could result from enhanced expression of mutant p53 induced by chemotherapeutic drugs (Bykov et al, 2005a).…”

The role of p53 in chemosensitivity and radiosensitivity

“…Some strategies have focused on the blockade of p53 to reduce toxicity of drugs or radiation (Komarov et al, 1999), whereas others have focused on blocking interactions between p53 and negative regulators such as MDM2 (Bottger et al, 1996(Bottger et al, , 1997Midgley et al, 2000). Yet other strategies have been targeted at mutated p53 to restore some wildtype p53-related transcriptional responses, for example, CP-31398 (Foster et al, 1999;Takimoto et al, 2002b; or Prima1 , or therapies aimed at preferential killing of p53-deficient cells (Hamid et al, 2003;Hecht et al, 2003). The identification of novel regulators such as HAUSP, Pirh2, Parc, Akt, or Sir2 has suggested other points of intervention to modulate p53 activity (Zhou et al, 2001;Vousden and Lu, 2002;Brooks and Gu, 2003;Grossman et al, 2003;Kastan and Zambetti, 2003;Leng et al, 2003;Nikolaev et al, 2003).…”