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Oncotarget. 2018 Jan 3;9(6):6883-6896. doi: 10.18632/oncotarget.23855. eCollection 2018 Jan 23.

Temozolomide, sirolimus and chloroquine is a new therapeutic combination that synergizes to disrupt lysosomal function and cholesterol homeostasis in GBM cells.

Author information

1
Department of Neurosurgery, Neurological Institute, Taipei Veterans General Hospital, Taipei, Taiwan.
2
School of Medicine, National Yang Ming University, Taipei, Taiwan.
3
Department of Neurological Surgery, School of Medicine and Public Health, University of Wisconsin, Madison, WI, USA.
4
JohnPro Biotech Inc., Taipei, Taiwan.
5
Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, Taipei, Taiwan.
6
Department of Radiation Therapy and Oncology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan.
7
Miami Cancer Institute, Miami, FL, USA.
#
Contributed equally

Abstract

Glioblastoma (GBM) cells are characterized by high phagocytosis, lipogenesis, exocytosis activities, low autophagy capacity and high lysosomal demand are necessary for survival and invasion. The lysosome stands at the cross roads of lipid biosynthesis, transporting, sorting between exogenous and endogenous cholesterol. We hypothesized that three already approved drugs, the autophagy inducer, sirolimus (rapamycin, Rapa), the autophagy inhibitor, chloroquine (CQ), and DNA alkylating chemotherapy, temozolomide (TMZ) could synergize against GBM. This repurposed triple therapy combination induced GBM apoptosis in vitro and inhibited GBM xenograft growth in vivo. Cytotoxicity is caused by induction of lysosomal membrane permeabilization and release of hydrolases, and may be rescued by cholesterol supplementation. Triple treatment inhibits lysosomal function, prevents cholesterol extraction from low density lipoprotein (LDL), and causes clumping of lysosome associated membrane protein-1 (LAMP-1) and lipid droplets (LD) accumulation. Co-treatment of the cell lines with inhibitor of caspases and cathepsin B only partially reverse of cytotoxicities, while N-acetyl cysteine (NAC) can be more effective. A combination of reactive oxygen species (ROS) generation from cholesterol depletion are the early event of underling mechanism. Cholesterol repletion abolished the ROS production and reversed the cytotoxicity from QRT treatment. The shortage of free cholesterol destabilizes lysosomal membranes converting aborted autophagy to apoptosis through either direct mitochondria damage or cathepsin B release. This promising anti-GBM triple therapy combination severely decreases mitochondrial function, induces lysosome-dependent apoptotic cell death, and is now poised for further clinical testing and validation.

KEYWORDS:

autophagy; chloroquine; cholesterol; lysosome cell death; rapamycin

Conflict of interest statement

CONFLICTS OF INTEREST MM has served as a consultant to Varian, Agenus, Insys, and Remedy and serves on the data safety monitoring committee for Monteris. JSK is a consultant to Kiyatec, Inc. KHC has stock ownership and leadership position in JohnPro Biotech Inc.

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