Elevated Production of Mitochondrial Reactive Oxygen Species via Hyperthermia Enhanced Cytotoxic Effect of Doxorubicin in Human Breast Cancer Cell Lines MDA-MB-453 and MCF-7.

Azusa Terasaki,Hiromu Ito,Hiroko Bando,Daisuke Matano,Hirofumi Matsui,Yoshiki Komatsu,Hisato Hara,Masahiko Terasaki,Hiromi Kurokawa

doi:10.3390/ijms21249522

Abstract

Hyperthermia (HT) treatment is a noninvasive cancer therapy, often used with radiation therapy and chemotherapy. Compared with 37 °C, 42 °C is mild heat stress for cells and produces reactive oxygen species (ROS) from mitochondria. To involve subsequent intracellular accumulation of DOX, we have previously reported that the expression of ATP-binding cassette sub-family G member 2 (ABCG2), an exporter of doxorubicin (DOX), was suppressed by a larger amount of intracellular mitochondrial ROS. We then hypothesized that the additive effect of HT and chemotherapy would be induced by the downregulation of ABCG2 expression via intracellular ROS increase. We used human breast cancer cell lines, MCF-7 and MDA-MB-453, incubated at 37 °C or 42 °C for 1 h to clarify this hypothesis. Intracellular ROS production after HT was detected via electron spin resonance (ESR), and DOX cytotoxicity was calculated. Additionally, ABCG2 expression in whole cells was analyzed using Western blotting. We confirmed that the ESR signal peak with HT became higher than that without HT, indicating that the intracellular ROS level was increased by HT. ABCG2 expression was downregulated by HT, and cells were injured after DOX treatment. DOX cytotoxicity enhancement with HT was considered a result of ABCG2 expression downregulation via the increase of ROS production. HT increased intracellular ROS production and downregulated ABCG2 protein expression, leading to cell damage enhancement via DOX.

Highlights

The number of cancer patients is increasing worldwide
Cell viability of MDA-MB-453 and MCF-7 after combination treatment decreased significantly in 1 and 10 μM DOX-treated groups, compared with that in the non-HT group that had only DOX treatment. Cell viability of both cell lines decreased dose-dependently of DOX. These results indicated that the combination treatment of HT and DOX showed an additive effect
TuhseeseMrietsouSlOtsXinTMditcoateddetect thatmthiteRROOSS, dfleutoercetescdebnyceESinRtewnesirteieosf wmeitroechsiognndifriicaalnotrliygienn. hanced in the HT group (Figure 3). These reAsluthltosuinghditchaetemdetchhaatnthisemRoOfSindtreateccetlleudlabrysEigSnRalwtrearnesodfumctiitoonchboynRdOriSalfroormigimn.itochondria remains unclear, we previously reported that mitROS increase was associated with ATP-binding cassette sub-family G member 2 (ABCG2) transporter downregulation [10,11]

Summary

Introduction

The number of cancer patients is increasing worldwide. In 2016, there were 17.2 million cancer cases and 8.9 million deaths globally. HT treatment is a noninvasive cancer therapy that, when combined with chemotherapy and radiation therapy, may have an additive effect. We previously reported that the treatment of gastric cancer with cisplatin elevated intracellular ROS generation and downregulated ABCG2 transporter expression [10]. We reported that ABCG2 downregulation by HT-induced elevation of ROS, derived from mitochondria (mitROS), induced an additive effect for photodynamic therapy in gastric cancer cells [11]. The ABC transporter excretes the drug extracellularly and reduces intracellular accumulation of the drug This mechanism of drug resistance and the cause of anticancer drug resistance are reported to be the overexpression of ABC transporters [13]. We hypothesized that HT-induced elevation of intracellular mitROS might have an additive effect with intracellular DOX accumulation by the downregulation of ABCG2 transporter expression. We investigated DOX cytotoxicity on breast cancer cells by adding DOX to HT, using two types of human breast cancer cells with different subtypes

Methods

Results

Conclusion