Targeting Notch Signaling and Autophagy Increases Cytotoxicity in Glioblastoma Neurospheres

Brain Pathol. 2016 Nov;26(6):713-723. doi: 10.1111/bpa.12343. Epub 2016 Feb 8.

Abstract

Glioblastomas are highly aggressive tumors that contain treatment resistant stem-like cells. Therapies targeting developmental pathways such as Notch eliminate many neoplastic glioma cells, including those with stem cell features, but their efficacy can be limited by various mechanisms. One potential avenue for chemotherapeutic resistance is the induction of autophagy, but little is known how it might modulate the response to Notch inhibitors. We used the γ-secretase inhibitor MRK003 to block Notch pathway activity in glioblastoma neurospheres and assessed its effects on autophagy. A dramatic, several fold increase of LC3B-II/LC3B-I autophagy marker was noted on western blots, along with the emergence of punctate LC3B immunostaining in cultured cells. By combining the late stage autophagy inhibitor chloroquine (CQ) with MRK003, a significant induction in apoptosis and reduction in growth was noted as compared to Notch inhibition alone. A similar beneficial effect on inhibition of cloogenicity in soft agar was seen using the combination treatment. These results demonstrated that pharmacological Notch blockade can induce protective autophagy in glioma neurospheres, resulting in chemoresistance, which can be abrogated by combination treatment with autophagy inhibitors.

Keywords: autophagy; chloroquine; combination treatment; gamma-secretase inhibitor; glioblastoma.

MeSH terms

  • Antimalarials / pharmacology
  • Autophagy / drug effects
  • Autophagy / physiology*
  • Cell Cycle / drug effects
  • Cell Cycle / physiology
  • Cell Line, Tumor
  • Cell Proliferation / drug effects
  • Cell Proliferation / physiology
  • Chloroquine / pharmacology
  • Colony-Forming Units Assay
  • Cyclic S-Oxides / pharmacology
  • Dose-Response Relationship, Drug
  • Drug Combinations
  • Gene Expression Regulation, Neoplastic / drug effects
  • Glioblastoma / pathology
  • Humans
  • Ki-67 Antigen / metabolism
  • Neoplastic Stem Cells / drug effects
  • Neoplastic Stem Cells / metabolism*
  • Receptors, Notch / antagonists & inhibitors
  • Receptors, Notch / genetics
  • Receptors, Notch / metabolism*
  • Signal Transduction / drug effects
  • Signal Transduction / physiology*
  • Starvation
  • Thiadiazoles / pharmacology

Substances

  • Antimalarials
  • Cyclic S-Oxides
  • Drug Combinations
  • Ki-67 Antigen
  • MRK 003
  • Receptors, Notch
  • Thiadiazoles
  • Chloroquine