A cell-based quantitative high-throughput image screening identified novel autophagy modulators

Abstract

Macroautophagy is a major cellular degradation pathway for long-lived proteins and cellular organelles to maintain cellular homeostasis. Reduced autophagy has been implicated in neurodegenerative diseases, metabolic syndrome, and tumorigenesis. In contrast, increased autophagy has been shown to protect against tissue injury and aging. Here we employed a cell-based quantitative high-throughput image screening (qHTS) for autophagy modulators using mouse embryonic fibroblasts (MEFs) that are stably expressing GFP-LC3. The library of pharmacologically active compounds (LOPAC) was used to screen for the autophagy modulators in compounds alone or in combination with the lysosome inhibitor chloroquine (CQ). The GFP-LC3 puncta were then quantified to measure autophagic flux. The primary screening revealed 173 compounds with efficacy more than 40%. These compounds were cherry-picked and re-tested at multiple different concentrations using the same assay. A number of novel autophagy inducers, inhibitors, and modulators with dual-effects on autophagy were identified from the cherry-pick screening. Interestingly, we found a group of compounds that induce autophagy are related to dopamine receptors and are commonly used as clinical psychiatric drugs. Among them, indatraline hydrochloride (IND), a dopamine inhibitor, and chlorpromazine hydrochloride (CPZ) and fluphenazine dihydrochloride (FPZ), two dopamine receptor antagonists, were further evaluated. We found that FPZ-induced autophagy through mTOR inhibition but IND and CPZ induced autophagy in an mTOR-independent manner. Our data suggest that image-based autophagic flux qHTS can efficiently identify autophagy inducers and inhibitors.

Enhanced human enterovirus 71 infection by endocytosis inhibitors reveals multiple entry pathways by enterovirus causing hand-foot-and-mouth diseases

“…Thus, our data supported that the endocytic system functioned uniquely in different cell types and another CME-independent pathway was responsible for EV71’s entry in A549 cells. Interestingly, CPZ was recently reported to be a strong autophagy inducer [ 30 , 31 ]. As autophagy was found to be required for efficient EV71 replication [ 32 – 34 ], could CPZ positively affect EV71 activity through inducing autophagy?…”

Combination of an Autophagy Inducer and an Autophagy Inhibitor: A Smarter Strategy Emerging in Cancer Therapy

“…autophagy via genomic interference against autophagic genes (siRNA targeting Atg3, Atg5, Atg7, and Beclin 1) or pharmacological inhibitors of key components within the autophagy pathway in cancer resistance (Kumar et al, 2015) ( Table 1). Additionally, there is also a growing interest in exploring more potent and specific pharmacological autophagy inhibitors (Golden et al, 2015;Li et al, 2016).…”

A novel co-segregatingDCTN1splice site variant in a family with Bipolar Disorder may hold the key to understanding the etiology

“…They achieve this by a common mechanism, namely via negatively regulating phosphoinositide signaling through depletion of inositol and by reducing myo-inositol-1,4,5-trisphosphate (IP3) [65,66]. Apart from the aforementioned psychotropic agents, the majority of antipsychotics (with two notable exceptions, namely being the typical antipsychotic haloperidol and the atypical antipsychotic clozapine) [67][68][69] as well as antidepressants [70,71] are also known to enhance neuronal autophagy. In addition, the autophagy-inducers rapamycin and trehalose have both been shown to have therapeutic effects in rodent models of affective disorders [72,73].…”