A Class of Diacylglycerol Acyltransferase 1 Inhibitors Identified by a Combination of Phenotypic High-throughput Screening, Genomics, and Genetics

Kirsten Tschapalda,Ya-Qin Zhang,Li Liu,Kseniya Golovnina,Thomas Schlemper,Thomas O Eichmann,Madhu Lal-Nag,Urmila Sreenivasan,John Mclenithan,Slava Ziegler,Carole Sztalryd,Achim Lass,Douglas Auld,Brian Oliver,Herbert Waldmann,Zhuyin Li,Min Shen,Matthew B Boxer,Mathias Beller

doi:10.1016/j.ebiom.2016.04.014

Abstract

Excess lipid storage is an epidemic problem in human populations. Thus, the identification of small molecules to treat or prevent lipid storage-related metabolic complications is of great interest. Here we screened >320.000 compounds for their ability to prevent a cellular lipid accumulation phenotype. We used fly cells because the multifarious tools available for this organism should facilitate unraveling the mechanism-of-action of active small molecules. Of the several hundred lipid storage inhibitors identified in the primary screen we concentrated on three structurally diverse and potent compound classes active in cells of multiple species (including human) and negligible cytotoxicity. Together with Drosophila in vivo epistasis experiments, RNA-Seq expression profiles suggested that the target of one of the small molecules was diacylglycerol acyltransferase 1 (DGAT1), a key enzyme in the production of triacylglycerols and prominent human drug target. We confirmed this prediction by biochemical and enzymatic activity tests.

Highlights

Cells of organisms from all phyla balance fluctuations in energy demand and supply by the regulated deposition and remobilization of energy rich storage molecules
While we focused on the role of CT2 in TAG metabolism, we looked for evidence of adverse effects on cell function because excess oleic acid (OA) can be toxic
Our phenotypic screen identified a significant number of small molecules that reduce cellular lipid storage with little to no toxicity and we explored three of them in greater detail

Summary

Introduction

Cells of organisms from all phyla balance fluctuations in energy demand and supply by the regulated deposition and remobilization of energy rich storage molecules. Neutral lipid synthesis relies on the conversion of surplus fatty acids by multiple biochemical pathways into triacylglycerols (TAG) and sterol esters (SE). TAG and SE are stored in specialized cellular organelles, called lipid droplets (LDs) (Farese and Walther, 2009; Murphy, 2001). In addition to the energy buffering function, the conversion of fatty acids to mono-, di-, and TAG protects cells from lipotoxicity. Lipids stored within LDs serve as anabolic building blocks for membrane, hormone, and other synthesis processes. While the molecular details of the regulated lipid storage remobilization in adipocytes are emerging (Lass et al, 2011), many details of cellular lipid metabolism remain elusive

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Conclusion