title: Detection of phospholipidosis induction: a cell-based assay in high-throughput and high-content format.
abstract: Drug-induced phospholipidosis is characterized by the accumulation of intracellular phospholipids in cells exposed to cationic amphiphilic drugs. The appearance of unicentric or multicentric multilamellar bodies viewed under an electron microscope (EM) is the morphological hallmark of phospholipidosis. Although the EM method is the gold standard for detecting cellular phospholipidosis, this method has its drawbacks, including low throughput, high cost, and unsuitability for screening a large chemical library. In this study, a cell-based phospholipidosis assay has been developed using the LipidTOX Red reagent in HepG2 cells and miniaturized into a 1536-well plate format. To validate this assay for high-throughput screening (HTS), the LOPAC library of 1280 compounds was screened using a quantitative HTS platform. A group of known phospholipidosis inducers, such as amiodarone, propranolol, chlorpromazine, desipramine, promazine, clomipramine, and amitriptyline, was identified by the screen, consistent with previous reports. Several novel phospholipidosis inducers, including NAN-190, ebastine, GR127935, and cis-(Z)-flupentixol, were identified in this study and confirmed using the EM method. These results demonstrate that this assay can be used to evaluate and profile large numbers of chemicals for drug-induced phospholipidosis.
authors.affiliation: 1National Center for Advancing Translational Sciences, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Sampada A
authors.initials: SA
authors.lastname: Shahane
authors.forename: Ruili
authors.initials: R
authors.lastname: Huang
authors.forename: David
authors.initials: D
authors.lastname: Gerhold
authors.forename: Ulrich
authors.initials: U
authors.lastname: Baxa
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Austin
authors.forename: Menghang
authors.initials: M
authors.lastname: Xia
mesh_terms: D056945:Hep G2 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism; D008064:Lipidoses / Q000139:chemically induced / Q000175:diagnosis* / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D010743:Phospholipids / Q000378:metabolism*; D015203:Reproducibility of Results
chemical_list: D010743:Phospholipids
keywords: LipidTOX; phospholipidosis; qHTS