Efficacy of atovaquone on EpCAM<sup>+</sup>CD44<sup>+</sup> HCT‑116 human colon cancer stem cells under hypoxia

Fu,Changhao; Xiao,Xu; Xu,Hao; Lu,Weifei; Wang,Yi

doi:10.3892/etm.2020.9416

Efficacy of atovaquone on EpCAM⁺CD44⁺ HCT‑116 human colon cancer stem cells under hypoxia

Authors:
- Changhao Fu
- Xu Xiao
- Hao Xu
- Weifei Lu
- Yi Wang
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Affiliations: Department of Regenerative Medicine, School of Pharmaceutical Sciences, Jilin University, Changchun, Jilin 130021, P.R. China, Key Laboratory of Organ Regeneration and Transplantation of Ministry of Education, Cancer Center, The First Hospital of Jilin University, Changchun, Jilin 130021, P.R. China, Department of Animal Biotechnology, College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, Henan 450002, P.R. China
Published online on: October 29, 2020 https://doi.org/10.3892/etm.2020.9416
Article Number: 286
Copyright: © Fu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Tumor hypoxia contributes to the development of resistance to chemotherapeutic drugs in several human cancer cell lines. Atovaquone, an anti‑malaria drug approved by the US Food and Drug Administration, has recently demonstrated anti‑cancer effects in vitro and in vivo in several cancer models. To assess the potential of atovaquone as an anti‑cancer agent under hypoxia in colorectal carcinoma, EpCAM⁺CD44⁺ colon cancer stem cells were isolated from HCT‑116 human colon cancer cells through magnetic‑activated cell sorting. The efficacy of atovaquone on cytotoxicity, tumorsphere formation, apoptosis, invasion and cell‑cycle progression under hypoxic conditions were evaluated. MTS assays indicated that atovaquone inhibited the proliferation of EpCAM⁺CD44⁺ HCT‑116 cells with a half‑maximal inhibitory concentration of 15 µM. Atovaquone inhibited tumorsphere formation and cell proliferation by causing cell‑cycle arrest in S‑phase, which induced apoptosis of EpCAM⁺CD44⁺ HCT‑116 cells, as detected by Annexin V‑FITC/PI double staining assays, and caused mitochondrial membrane potential depolarization, as determined by a JC‑1 staining assay. Reverse transcription‑quantitative PCR demonstrated increased expression of Bax and downregulation of Bcl‑2. Transwell invasion assays indicated that atovaquone inhibited the invasiveness of EpCAM⁺CD44⁺ HCT‑116 cells under hypoxia, which was associated with upregulation of MMP‑2 and ‑9 and increased expression of tissue inhibitor of MMPs (TIMP)‑1. Taken together, atovaquone reduced the tumorsphere formation and invasion ability of EpCAM⁺CD44⁺ HCT‑116 cells, at least in part by increasing the expression of TIMP‑1 and downregulating the expression of MMP‑2 and ‑9, as well as the cells' viability by inducing cell‑cycle arrest in S‑phase and induction of apoptosis via the Bcl‑2/Bax pathway under hypoxic conditions. Further studies are warranted to explore the mechanisms of action of atovaquone as a promising anticancer agent in the treatment of colorectal carcinoma.

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