title: Chloroquine potentiates temozolomide cytotoxicity by inhibiting mitochondrial autophagy in glioma cells.
abstract: Mitochondrial autophagy eliminates damaged mitochondria and decreases reactive oxygen species (ROS). The autophagy inhibitor chloroquine (CQ) potentiates temozolomide (TMZ) cytotoxicity in glioma cells, but it is not known whether CQ does this by inhibiting mitochondrial autophagy. The effects of CQ and TMZ on MitoSOX Red fluorescence, a mitochondrial ROS indicator, and cell death were examined in rat C6 glioma cells. Mitochondrial autophagy was monitored by the colocalization of MitoTracker Red fluorescence and EGFP-LC3 dots. Mitochondrial content was measured by MitoTracker Green fluorescence and immunoblotting for a mitochondrial protein. Finally, CQ's effects on tumor cells derived from a glioblastoma patient and human U87-MG glioblastoma cells were assessed. TMZ (100-1,000 μM) alone did not affect mitochondrial ROS or cell death in C6 cells, but when administered with CQ (10 μM), it increased mitochondrial ROS and cell death. Antioxidants significantly suppressed the CQ-augmented cell death in TMZ-treated cells, indicating that mitochondrial ROS were involved in this cell death. TMZ treatment reduced MitoTracker Green fluorescence and mitochondrial protein levels, and these effects were inhibited by CQ. TMZ also increased the colocalization of EGFP-LC3 dots with mitochondria, and CQ enhanced this effect. CQ potentiated TMZ-induced cytotoxicity in patient-derived glioblastoma cells as well as human U87-MG glioblastoma cells. These results suggest that CQ increases cellular ROS and augments TMZ cytotoxicity in glioma cells by inhibiting mitochondrial autophagy.
authors.affiliation: Department of Pharmacology, Sapporo Medical University School of Medicine, South-1, West-17, Chuo-ku, Sapporo, 060-8556, Japan.
authors.forename: Yusuke S
authors.initials: YS
authors.lastname: Hori
authors.forename: Ryusuke
authors.initials: R
authors.lastname: Hosoda
authors.forename: Yukinori
authors.initials: Y
authors.lastname: Akiyama
authors.forename: Rio
authors.initials: R
authors.lastname: Sebori
authors.forename: Masahiro
authors.initials: M
authors.lastname: Wanibuchi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Mikami
authors.forename: Toshiya
authors.initials: T
authors.lastname: Sugino
authors.forename: Kengo
authors.initials: K
authors.lastname: Suzuki
authors.forename: Mitsuhisa
authors.initials: M
authors.lastname: Maruyama
authors.forename: Miki
authors.initials: M
authors.lastname: Tsukamoto
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Mikuni
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Horio
authors.forename: Atsushi
authors.initials: A
authors.lastname: Kuno
mesh_terms: D000818:Animals; D000962:Antimalarials / Q000494:pharmacology; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D001343:Autophagy / Q000187:drug effects*; D002738:Chloroquine / Q000494:pharmacology*; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004357:Drug Synergism; D005910:Glioma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D000077204:Temozolomide; D014407:Tumor Cells, Cultured
chemical_list: D000962:Antimalarials; D018906:Antineoplastic Agents, Alkylating; D017382:Reactive Oxygen Species; D003606:Dacarbazine; D002738:Chloroquine; D000077204:Temozolomide