Inhibition of endoplasmic reticulum (ER) stress sensors sensitizes cancer stem-like cells to ER stress-mediated apoptosis.

Asaha Fujimoto,Aki Yamashita,Yutaka Osuga,Katsuyuki Adachi,Juri Ogishima,Takahide Arimoto,Katsutoshi Oda,Kensuke Tomio,Haruka Nishida,Tomoko Inoue,Takeshi Nagamatsu,Kei Kawana,Hiroe Nakamura,Mitsuyo Yoshida,Tomoyuki Fujii,Osamu Wada-Hiraike,Masakazu Sato,Yoko Matsumoto,Ayumi Taguchi

doi:10.18632/oncotarget.10126

Abstract

Although cancer stem cells (CSC) have been implicated in the development of resistance to anti-cancer therapy including chemotherapy, the mechanisms underlying chemo-resistance by CSC have not yet been elucidated. We herein isolated sphere-forming (cancer stem-like) cells from the cervical cancer cell line, SiHa, and examined the unfolded protein reaction (UPR) to chemotherapeutic-induced endoplasmic reticulum (ER) stress. We revealed that tunicamycin-induced ER stress-mediated apoptosis occurred in monolayer, but not sphere-forming cells. Biochemical assays demonstrated that sphere-forming cells were shifted to pro-survival signaling through the inactivation of IRE1 (XBP-1 splicing) and activation of PERK (elF2α phosphorylation) branches under tunicamycin-induced ER stress conditions. The proportion of apoptotic cells among sphere-forming cells was markedly increased by the tunicamycin+PERK inhibitor (PERKi) treatment, indicating that PERKi sensitized sphere-forming cells to tunicamycin-induced apoptosis. Cisplatin is also known to induce ER stress-mediated apoptosis. A low concentration of cisplatin failed to shift sphere-forming cells to apoptosis, although IRE1 branch, but not PERK, was activated. ER stress-mediated apoptosis occurred in sphere-forming cells by the cisplatin+IRE1α inhibitor (IRE1i) treatment. IRE1i, synergistic with cisplatin, up-regulated elF2α phosphorylation, and this was followed by the induction of CHOP in sphere-forming cells. The results of the present study demonstrated that the inhibition of ER stress sensors, combined with ER stress-inducible chemotherapy, shifted cancer stem-like cells to ER stress-mediated apoptosis.

Highlights

Cancer stem cells (CSC) account for only a small population of disseminated tumor cells, they exhibit several unique properties that are similar to those of adult stem cells, but are not observed in the majority of cancer cells
Previous studies have used cancer sphere-forming cells isolated from various malignant cell lines and clinical samples to investigate the properties of cancer stem cells (CSC) [21, 22]
We isolated sphere-forming cells derived from the cervical cancer cell line, SiHa (HPV16-positive squamous cell carcinoma), according to the standard protocol for sphere formation using a low-attachment plate and serum-free media [23]

Summary

INTRODUCTION

Cancer stem cells (CSC) account for only a small population of disseminated tumor cells, they exhibit several unique properties that are similar to those of adult stem cells, but are not observed in the majority of cancer cells. Once ER stress occurs, the BiP/ unfolded protein complex dissociates from ER stress sensors This accompanies the activation of their signaling pathways and blocks the initiation of protein synthesis until the cell recovers from ER stress, namely, the cell shifts to pro-survival [14]. E6 and E6AP synergistically degrade p53 via a proteasome pathway and E6 activates hTERT [19, 20] These findings imply that p53-dependent apoptosis may be ignored in sphere-forming and monolayer cells derived from SiHa cells. We demonstrated that cancer stem-like (sphereforming) cells derived from SiHa cells resisted ER stress-mediated apoptosis and that signaling activated with chemotherapeutic ER stress was different between sphere-forming and monolayer cells. Our results demonstrated that the inhibition of ER stress sensors combined with chemotherapy has potential as a novel therapeutic strategy

RESULTS AND DISCUSSION

MATERIALS AND METHODS

CONFLICTS OF INTEREST