title: Sequential dosing in chemosensitization: targeting the PI3K/Akt/mTOR pathway in neuroblastoma.
abstract: Breaking resistance to chemotherapy is a major goal of combination therapy in many tumors, including advanced neuroblastoma. We recently demonstrated that increased activity of the PI3K/Akt network is associated with poor prognosis, thus providing an ideal target for chemosensitization. Here we show that targeted therapy using the PI3K/mTOR inhibitor NVP-BEZ235 significantly enhances doxorubicin-induced apoptosis in neuroblastoma cells. Importantly, this increase in apoptosis was dependent on scheduling: while pretreatment with the inhibitor reduced doxorubicin-induced apoptosis, the sensitizing effect in co-treatment could further be increased by delayed addition of the inhibitor post chemotherapy. Desensitization for doxorubicin-induced apoptosis seemed to be mediated by a combination of cell cycle-arrest and autophagy induction, whereas sensitization was found to occur at the level of mitochondria within one hour of NVP-BEZ235 posttreatment, leading to a rapid loss of mitochondrial membrane potential with subsequent cytochrome c release and caspase-3 activation. Within the relevant time span we observed marked alterations in a ∼30 kDa protein associated with mitochondrial proteins and identified it as VDAC1/Porin protein, an integral part of the mitochondrial permeability transition pore complex. VDAC1 is negatively regulated by the PI3K/Akt pathway via GSK3β and inhibition of GSK3β, which is activated when Akt is blocked, ablated the sensitizing effect of NVP-BEZ235 posttreatment. Our findings show that cancer cells can be sensitized for chemotherapy induced cell death - at least in part - by NVP-BEZ235-mediated modulation of VDAC1. More generally, we show data that suggest that sequential dosing, in particular when multiple inhibitors of a single pathway are used in the optimal sequence, has important implications for the general design of combination therapies involving molecular targeted approaches towards the PI3K/Akt/mTOR signaling network.
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Mike-Andrew
authors.initials: MA
authors.lastname: Westhoff
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Najmeh
authors.initials: N
authors.lastname: Faham
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Daniela
authors.initials: D
authors.lastname: Marx
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Lisa
authors.initials: L
authors.lastname: Nonnenmacher
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Jennewein
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Enzenmüller
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Patrick
authors.initials: P
authors.lastname: Gonzalez
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Simone
authors.initials: S
authors.lastname: Fulda
authors.affiliation: Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Ulm, Germany.
authors.forename: Klaus-Michael
authors.initials: KM
authors.lastname: Debatin
mesh_terms: D000970:Antineoplastic Agents / Q000008:administration & dosage; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000008:administration & dosage; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000008:administration & dosage; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000008:administration & dosage*; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D007093:Imidazoles / Q000008:administration & dosage; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D009447:Neuroblastoma / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors*; D011804:Quinolines / Q000008:administration & dosage; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors*; D050995:Voltage-Dependent Anion Channel 1 / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D007093:Imidazoles; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011804:Quinolines; C494762:VDAC1 protein, human; D004317:Doxorubicin; D050995:Voltage-Dependent Anion Channel 1; C546842:MTOR protein, human; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D038362:Glycogen Synthase Kinase 3; C531198:dactolisib