Abstract 1218: Synthetic lethal targeting of p53 mutant cells with prenylation inhibitors
Authors: Ferdosi Shay R. , Katchman Benjamin , Loo Jia , Grewal Harneet , Eaton Seron , Yang Shanshan , Park Jin , Labaer Joshua , Anderson Karen S. 
Published: July 2017

Abstract
Introduction: TP53 is mutated in 30-40% of breast cancers and is associated with poor prognosis. Mutation of TP53 causes increased cellular proliferation, migration and invasion, and downstream activation of multiple pathways. HMG CoA reductase inhibitors, such as simvastatin, inhibit tumorigenic properties induced by TP53 mutation. Mechanism of this response is an important question in targeted cancer therapy. It is well known that HMG CoA reductase inhibitors block cholesterol production and prenylation. Therefore, we hypothesized that prenylation inhibitors would target p53 mutant cells primarily by inhibiting activation of the ras family induced by TP53 mutation.
Method: We generated 5 MCF10A stably transduced cell lines over-expressing each of the common TP53 point mutations, R273H, G245S, R248Q, Y234C, or wild type p53. Total Ras, Ras-GTP, total RhoA and RhoA-GTP was measured by immunoprecipitation and immunoblot. Proliferation was measured using CellTiterGlo. The functional effects of a panel of prenylation inhibitors was measured using a caspase3 reporter assay, and migration was measured with a scratch assay. Illumina mRNA sequencing was performed to measure gene expression of mutant and wild type cells before and after simvastatin treatment (2.5µl, 48 h). The RNAseq was analyzed with the DAVID Enrichment tool to evaluate ras pathway activation induced by TP53 mutation.
Results: Mutation in TP53 was associated with significant activation of both wild-type Ras and RhoA. Proliferation of cells expressing mutant TP53 was sensitive to a panel of prenylation inhibitors, including simvastatin. Growth inhibition by simvastatin correlated with induction of apoptosis, and could be fully rescued by addition of farnesylpyrophosphate or geranylgeranylpyrophosphate, suggesting that simvastatin functionally blocks both prenylation pathways which are activated in the presence of mutant p53 (IC50(wt/mut) = 4). RNAseq analysis confirms that Ras signaling pathways, including GAPs and GEFs are enriched in the presence of mutant TP53, and FOXO signaling is significantly targeted post statin treatment (P&lt;0.01).
Conclusion: p53 mutant breast cancer cells are highly sensitive to prenylation inhibition due to activation of both ras and rhoA. Simvastatin inhibits both farnesylation and geranylgeranylation, effectively blocking ras pathway activation and induction of proliferation in p53 mutant cell lines. A gene expression panel associated with ras pathway activation was identified and may be predictive biomarkers for sensitivity to statin therapy.
Citation Format: Shay R. Ferdosi, Benjamin Katchman, Jia Loo, Harneet Grewal, Seron Eaton, Shanshan Yang, Jin Park, Joshua Labaer, Karen S. Anderson. Synthetic lethal targeting of p53 mutant cells with prenylation inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1218. doi:10.1158/1538-7445.AM2017-1218