title: Long-term acquired everolimus resistance in pancreatic neuroendocrine tumours can be overcome with novel PI3K-AKT-mTOR inhibitors.
abstract: BACKGROUND
  The mTOR-inhibitor everolimus improves progression-free survival in advanced pancreatic neuroendocrine tumours (PNETs). However, adaptive resistance to mTOR inhibition is described.
  
  
  METHODS
  QGP-1 and BON-1, two human PNET cell lines, were cultured with increasing concentrations of everolimus up to 22 weeks to reach a dose of 1 μM everolimus, respectively, 1000-fold and 250-fold initial IC50. Using total DNA content as a measure of cell number, growth inhibitory dose-response curves of everolimus were determined at the end of resistance induction and over time after everolimus withdrawal. Response to ATP-competitive mTOR inhibitors OSI-027 and AZD2014, and PI3K-mTOR inhibitor NVP-BEZ235 was studied. Gene expression of 10 PI3K-Akt-mTOR pathway-related genes was evaluated using quantitative real-time PCR (RT-qPCR).
  
  
  RESULTS
  Long-term everolimus-treated BON-1/R and QGP-1/R showed a significant reduction in everolimus sensitivity. During a drug holiday, gradual return of everolimus sensitivity in BON-1/R and QGP-1/R led to complete reversal of resistance after 10-12 weeks. Treatment with AZD2014, OSI-027 and NVP-BEZ235 had an inhibitory effect on cell proliferation in both sensitive and resistant cell lines. Gene expression in BON-1/R revealed downregulation of MTOR, RICTOR, RAPTOR, AKT and HIF1A, whereas 4EBP1 was upregulated. In QGP-1/R, a downregulation of HIF1A and an upregulation of ERK2 were observed.
  
  
  CONCLUSIONS
  Long-term everolimus resistance was induced in two human PNET cell lines. Novel PI3K-AKT-mTOR pathway-targeting drugs can overcome everolimus resistance. Differential gene expression profiles suggest different mechanisms of everolimus resistance in BON-1 and QGP-1.
authors.affiliation: Center of Oncological Research (CORE), University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.|Section of Endocrinology, Department of Internal Medicine, Erasmus Medical Center, Dr Molewaterplein 40, 3015GD Rotterdam, The Netherlands.
authors.forename: Timon
authors.initials: T
authors.lastname: Vandamme
authors.affiliation: Center of Oncological Research (CORE), University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.|Center of Medical Genetics, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.
authors.forename: Matthias
authors.initials: M
authors.lastname: Beyens
authors.affiliation: Center of Oncological Research (CORE), University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.|Center of Medical Genetics, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.
authors.forename: Ken Op
authors.initials: KO
authors.lastname: de Beeck
authors.affiliation: Section of Endocrinology, Department of Internal Medicine, Erasmus Medical Center, Dr Molewaterplein 40, 3015GD Rotterdam, The Netherlands.
authors.forename: Fadime
authors.initials: F
authors.lastname: Dogan
authors.affiliation: Section of Endocrinology, Department of Internal Medicine, Erasmus Medical Center, Dr Molewaterplein 40, 3015GD Rotterdam, The Netherlands.
authors.forename: Peter M
authors.initials: PM
authors.lastname: van Koetsveld
authors.affiliation: Department of Pathology, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.
authors.forename: Patrick
authors.initials: P
authors.lastname: Pauwels
authors.affiliation: Center of Medical Genetics, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.
authors.forename: Geert
authors.initials: G
authors.lastname: Mortier
authors.affiliation: Department of Molecular Imaging, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.
authors.forename: Christel
authors.initials: C
authors.lastname: Vangestel
authors.affiliation: Section of Endocrinology, Department of Internal Medicine, Erasmus Medical Center, Dr Molewaterplein 40, 3015GD Rotterdam, The Netherlands.
authors.forename: Wouter
authors.initials: W
authors.lastname: de Herder
authors.affiliation: Center of Medical Genetics, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.
authors.forename: Guy
authors.initials: G
authors.lastname: Van Camp
authors.affiliation: Center of Oncological Research (CORE), University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium.
authors.forename: Marc
authors.initials: M
authors.lastname: Peeters
authors.affiliation: Section of Endocrinology, Department of Internal Medicine, Erasmus Medical Center, Dr Molewaterplein 40, 3015GD Rotterdam, The Netherlands.
authors.forename: Leo J
authors.initials: LJ
authors.lastname: Hofland
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm; D000068338:Everolimus / Q000494:pharmacology*; D020869:Gene Expression Profiling; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D018358:Neuroendocrine Tumors / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics; D010190:Pancreatic Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors*; D011804:Quinolines / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors*; D014227:Triazines / Q000494:pharmacology
chemical_list: D000970:Antineoplastic Agents; D007093:Imidazoles; C568605:OSI 027; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D011804:Quinolines; D014227:Triazines; D000068338:Everolimus; C546842:MTOR protein, human; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; C531198:dactolisib