Genome-wide RNAi analysis reveals that simultaneous inhibition of specific mevalonate pathway genes potentiates tumor cell death.

Aleksandra A Pandyra,Karl S Lang,Traver Hart,Corey Nislow,Jason Moffat,Carolyn A Goard,Piyush Sharma,Rosemary Yu,Linda Z Penn,Janice T Pong,Manpreet Kalkat,Guri Giaever,Marinella Gebbia,Peter J Mullen,Kevin R Brown,Elke Ericson

doi:10.18632/oncotarget.4817

Abstract

The mevalonate (MVA) pathway is often dysregulated or overexpressed in many cancers suggesting tumor dependency on this classic metabolic pathway. Statins, which target the rate-limiting enzyme of this pathway, 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), are promising agents currently being evaluated in clinical trials for anti-cancer efficacy. To uncover novel targets that potentiate statin-induced apoptosis when knocked down, we carried out a pooled genome-wide short hairpin RNA (shRNA) screen. Genes of the MVA pathway were amongst the top-scoring targets, including sterol regulatory element binding transcription factor 2 (SREBP2), 3-hydroxy-3-methylglutaryl-coenzyme A synthase 1 (HMGCS1) and geranylgeranyl diphosphate synthase 1 (GGPS1). Each gene was independently validated and shown to significantly sensitize A549 cells to statin-induced apoptosis when knocked down. SREBP2 knockdown in lung and breast cancer cells completely abrogated the fluvastatin-induced upregulation of sterol-responsive genes HMGCR and HMGCS1. Knockdown of SREBP2 alone did not affect three-dimensional growth of lung and breast cancer cells, yet in combination with fluvastatin cell growth was disrupted. Taken together, these results show that directly targeting multiple levels of the MVA pathway, including blocking the sterol-feedback loop initiated by statin treatment, is an effective and targetable anti-tumor strategy.

Highlights

Cancer cells undergo drastic metabolic reprogramming to meet their increased demand for energy and macromolecules
Analysis of the short hairpin RNA (shRNA) remaining in the ethanol controltreated but not fluvastatin-treated cells identified genes that were lethal in cells exposed to fluvastatin
Genome-wide shRNA screen, we have demonstrated the utility of targeting more than one member of the MVA pathway to further sensitize cells to fluvastatin

Summary

Introduction

Cancer cells undergo drastic metabolic reprogramming to meet their increased demand for energy and macromolecules. Amongst the metabolic changes occurring in cancer cells is increased de novo lipid and cholesterol synthesis through both the fatty acid synthesis and mevalonate (MVA) pathways [1, 2] The latter leads to the production of cholesterol, and results in important non-sterol end products including farnesyl and geranylgeranyl isoprenoids, dolichol, ubiquinone, and isopentenyladenine (Figure 1A). MVA pathway enzymes can be upregulated by mutant p53 [6] and their elevated expression is associated with poor prognosis and reduced survival in cancer patients [6, 7] Consistent with this observation, over-expression of the rate-limiting enzyme, 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), contributes to oncogenic progression [7]. These multiple levels of MVA pathway dysregulation suggest that cancer cells are dependent on the MVA-derived end products and preferentially sensitive to inhibition of the MVA pathway

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Results

Conclusion