Abstract 977: Elevated MDM2 protein contributes to activity of MDM2 antagonists by enabling p53 induction of antitumor microRNAs

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Small-molecule inhibitors of p53-MDM2 binding, the nutlins, can free p53 from MDM2 control and effectively activate its growth suppressive and pro-apoptotic activity in cancer cells (Vassilev et al., Science, 2004, 303, 844). Optimized members of this family of molecules are currently under clinical investigation as novel therapeutic modality for cancers expressing wild-type p53. We have shown previously that MDM2 antagonists are most effective in cancer cells that overproduce MDM2 as a result of mdm2 gene amplification (Tovar et al., PNAS, 2006, 103, 1888). Here, we show that reduction of MDM2 protein by RNA interference decreases the effectiveness of nutlin-3a in osteosarcoma cells with mdm2 gene amplification, suggesting that MDM2 protein contributes to antitumor activity of MDM2 antagonists. In search of the mechanism behind this phenomenon, we found that nutlin-induced p53 activates the transcription of a specific subset of microRNAs in cancer cells with mdm2 amplification but not in cells expressing normal levels of MDM2 protein. Five specific members of the class (miR1, miR133, miR141, miR184, miR375) were studied and showed that their p53-dependent transcription is also dependent on the presence of high levels of MDM2 protein, generated by continuous p53 activation of MDM2 in the presence of nutlin. Weak p53 response elements were found in the promoter regions of all five miRs and p53 binding was conformed by ChIP analysis of cell lysates and p53 reporters in experiments with miR375. Transcriptional cofactor p300 was found elevated in cancer cells with mdm2 gene amplification and dependent on the presence of high MDM2 protein levels. Our experimental data suggests that in the presence of high p300, and high MDM2, generated by nutlin treatment, p53 acquires the ability to activate weak p53-responsive miR promoters in MDM2-amplified cells. Interestingly, miR mimics of two of the five p53-induced miRs showed growth suppressive effects in cancer cells and enhanced the activity of low dose nutlin-3a. These findings reveal a novel mechanism for modulation of p53 transcriptional activity and indicate that the high levels of MDM2, generated in mdm2-amplified cancer cells in the presence of nutlin, contribute to its antitumor. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 977. doi:1538-7445.AM2012-977