Abstract P1-03-02: Preclinical efficacy of a novel and potent inhibitor of the MDM2-p53 axis AMG 232 in ER+ breast cancer model
Authors: De PK , Carlson JH , Jepperson T , Krie A , Williams K , Klein J , Canon J , Williams C , Dey N , Leyland-Jones B 
Published: February 2018

Background: Approximately 70% of breast cancers (BC) express estrogen receptors (ER) and/or progesterone receptors (PR) which define their degrees of estrogen dependence. Although mutations in the tumor suppressor TP53 gene are thought to be the most abundant genetic alterations occurring in cancers, the relative prevalence of TP53 mutations in ER+ BC is low (luminal A 12% and luminal B 29%) compared to HER2+ or TNBC, and total MDM2/4 alterations are about 12% (Cancer Genome Atlas Network Nature. 2012). In our (Avera Cancer Institute, Sioux Falls, SD) ER+ BC cohort TP53 and MDM2/4 alterations are 25% and 14% respectively. Purpose: Blocking the interaction between MDM2 and p53 may generate a novel treatment opportunity in TP53 wild-type ER+ BC patients. Methods: Comprehensive genomic profiles from 160 ER+ BC patients (February 2014 through May 2017) were analyzed. Patients were biopsied after consultation and samples were analyzed for genomic [FoundationOne] and proteomic analyses [Theranostics]). We also evaluated mutation distribution in cell-free DNA via digital NGS using the Guardant360 panel. We tested the anti-tumor efficacy of MDM2 inhibitor (AMG 232) alone or a combination of an aromatase inhibitor, letrozole plus AMG 232 in ER+ BC model (MCF7, Zr-75-1, & MDA-MB415). Results: 1) AMG 232 binds the MDM2 protein, blocks the MDM2-p53 interaction and induces p53 expression and activity, 2) p53 effector molecule p21 is robustly induced following the treatment of AMG 232, 3) the anti-proliferative activity of AMG 232 was observed by 3D-ON-TOP clonogenic assay and real-time monitoring in an IncuCyte Zoom, 4)ER+ /TP53 wild-type (WT) cell lines exhibited an increase in annexin V positivity (initiation of apoptotic activity) following AMG 232 treatment.AMG 232 also induced the apoptotic markers cleaved-CASPASE3 and cleaved PARP1 protein expression in ER+/PR+/TP53 WT and PTEN null breast cancer cell lines, 5) AMG 232 treatment induced G1 cells cycle arrest (Flow cytometric analysis of PI staining) dose dependently which may be associated with upregulation of p21. 6)we also assessed mRNA expression (several pro-apoptotic and anti-apoptotic molecules) by RT-qPCR following the treatment of AMG232 in MCF7 (ER+/TP53 WT) and Zr-75-1 (ER+/PR+/TP53 WT and PTEN null) cell lines. Similar to our protein expression data, mRNA data also showed that pro-apoptotic/cell cycle inhibitor transcripts such as p21 and PUMA mRNA expression were significantly increased following the treatment of AMG 232. On the contrary anti-apoptotic/ pro-survival transcripts such as survivin (BIRC5) and stathmin (STMN1) mRNA expression were significantly decreased following the treatment of AMG 232 and 7) importantly, normoxic-proteasomal degradation of HIF1α (responsible for the development of chemotherapy and targeted therapy resistance) was not rescued by prior treatment of this novel MDM2-p53 interacting inhibitor AMG 232 in Zr-75-1 cell line.Conclusion:Taken together, our data suggest that AMG 232 effectively inhibited proliferation and enhanced apoptosis via inhibition of the MDM2-p53 interaction and p53-mediated downstream signaling events in ER+/TP53 wild-type BC model.

Citation Format: De PK, Carlson JH, Jepperson T, Krie A, Williams K, Klein J, Canon J, Williams C, Dey N, Leyland-Jones B. Preclinical efficacy of a novel and potent inhibitor of the MDM2-p53 axis AMG 232 in ER+ breast cancer model [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P1-03-02.