PRIMA-1 induces apoptosis in p53 mutant breast cancer cell lines by inhibiting the JNK cell survival pathway and promoting the activation of proapoptotic p53 targets Bax and PUMA

3051 The p53 protein plays a major role in the maintenance of genome stability in mammalian cells. Mutations of p53 occur in over 40% of breast cancers and are indicative of tumor resistant to chemotherapeutic agents. Recently, there has been a high degree of interest in pharmacological approaches to restore the normal function to mutant p53. The compound p53-dependent reactivation and induction of massive apoptosis (PRIMA-1) has been shown to induce cytotoxic effects and apoptosis in human tumor cells. We reported (Breast Cancer Res. 7: R765-774, 2005) that PRIMA-1 restores the p53-hsp90α interaction, enhances the translocation of this complex to the nucleus and reactivates p53 transcriptional activity in breast cancer cells. Here we studied the molecular mechanisms of apoptosis in breast cancer cells (MDA-231 and GI-101A) with various types of p53 mutations as compared to MCF-7 with wild-type p53 after treatment with PRIMA-1. We show that PRIMA-1 selectively induces apoptosis in MDA-231 and GI-101A compared to MCF-7 breast cancer cells as measured by the Annexin V assays. This effect was paralleled by an increase in the total levels of p53 and the induction of its phosphorylation at Ser15 as detected by Western blot analysis with the Odyssey™Infrared Imaging System. Using a chromatin immunoprecipitation (ChIP) assay, we show that PRIMA-1 restored the p53 DNA binding activity to the promoters of proapoptotic genes such as PUMA and Bax, but inhibited the binding activity to the promoters of the anti-apoptotic MAP4K4 gene. Hence, our data indicate that PRIMA-1 induces apoptosis in p53 mutant breast cancer cells by promoting the activation of proapoptotic signaling pathways and inhibiting the JNK cell survival pathway. (Supported by Susan G. Komen Breast Cancer Foundation grant BCTR0402398)