title: Endoplasmic reticulum stress and cell death in mTORC1-overactive cells is induced by nelfinavir and enhanced by chloroquine.
abstract: Inappropriate activation of mammalian/mechanistic target of rapamycin complex 1 (mTORC1) is common in cancer and has many cellular consequences including elevated endoplasmic reticulum (ER) stress. Cells employ autophagy as a critical compensatory survival mechanism during ER stress. This study utilised drug-induced ER stress through nelfinavir in order to examine ER stress tolerance in cell lines with hyper-active mTORC1 signalling. Our initial findings in wild type cells showed nelfinavir inhibited mTORC1 signalling and upregulated autophagy, as determined by decreased rpS6 and S6K1 phosphorylation, and SQTSM1 protein expression, respectively. Contrastingly, cells with hyper-active mTORC1 displayed basally elevated levels of ER stress which was greatly exaggerated following nelfinavir treatment, seen through increased CHOP mRNA and XBP1 splicing. To further enhance the effects of nelfinavir, we introduced chloroquine as an autophagy inhibitor. Combination of nelfinavir and chloroquine significantly increased ER stress and caused selective cell death in multiple cell line models with hyper-active mTORC1, whilst control cells with normalised mTORC1 signalling tolerated treatment. By comparing chloroquine to other autophagy inhibitors, we uncovered that selective toxicity invoked by chloroquine was independent of autophagy inhibition yet entrapment of chloroquine to acidified lysosomal/endosomal compartments was necessary for cytotoxicity. Our research demonstrates that combination of nelfinavir and chloroquine has therapeutic potential for treatment of mTORC1-driven tumours.
authors.affiliation: Institute of Cancer and Genetics, Cardiff University, Heath Park, Cardiff CF14 4XN, UK.
authors.forename: Charlotte E
authors.initials: CE
authors.lastname: Johnson
authors.affiliation: Institute of Cancer and Genetics, Cardiff University, Heath Park, Cardiff CF14 4XN, UK.
authors.forename: David K
authors.initials: DK
authors.lastname: Hunt
authors.affiliation: Institute of Cancer and Genetics, Cardiff University, Heath Park, Cardiff CF14 4XN, UK.
authors.forename: Marie
authors.initials: M
authors.lastname: Wiltshire
authors.affiliation: Department of Cell Physiology and Pharmacology, University of Leicester, The Henry Wellcome Building, University Road, Leicester LE1 9HN, UK.
authors.forename: Terry P
authors.initials: TP
authors.lastname: Herbert
authors.affiliation: Institute of Cancer and Genetics, Cardiff University, Heath Park, Cardiff CF14 4XN, UK.
authors.forename: Julian R
authors.initials: JR
authors.lastname: Sampson
authors.affiliation: Institute of Cancer and Genetics, Cardiff University, Heath Park, Cardiff CF14 4XN, UK.
authors.forename: Rachel J
authors.initials: RJ
authors.lastname: Errington
authors.affiliation: Institute of Cancer and Genetics, Cardiff University, Heath Park, Cardiff CF14 4XN, UK.
authors.forename: D Mark
authors.initials: DM
authors.lastname: Davies
authors.affiliation: Institute of Cancer and Genetics, Cardiff University, Heath Park, Cardiff CF14 4XN, UK. Electronic address: teea@cardiff.ac.uk.
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Tee
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001343:Autophagy / Q000187:drug effects; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D002738:Chloroquine / Q000494:pharmacology*; D004622:Embryo, Mammalian / Q000166:cytology; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D018942:Macrolides / Q000494:pharmacology; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D008954:Models, Biological; D046912:Multiprotein Complexes / Q000378:metabolism*; D019888:Nelfinavir / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins / Q000172:deficiency / Q000378:metabolism
chemical_list: D018942:Macrolides; D046912:Multiprotein Complexes; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins; C025946:3-methyladenine; D002738:Chloroquine; C040929:bafilomycin A1; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases; D019888:Nelfinavir; D000225:Adenine
keywords: Autophagy; Cancer; Chloroquine; ER stress; Nelfinavir; TSC; mTOR