Trichostatin A Targets the Mitochondrial Respiratory Chain, Increasing Mitochondrial Reactive Oxygen Species Production to Trigger Apoptosis in Human Breast Cancer Cells

Shujuan Sun,Yuan Tian,Peng Wu,Jia Liu,Jianfeng Zhou,Ding Ma,Yong Fang,Yingyan Han,Dhyan Chandra

doi:10.1371/journal.pone.0091610

Abstract

AimHistone deacetylase inhibitors (HDACIs)-based therapies have stimulated interest via their anti-tumor activities, including apoptosis induction, cell cycle arrest, cell differentiation, and autophagy. However, the mechanisms of HDACI-associated anti-tumor activity are not yet clearly defined. The aim of this study was to explore the key events of Trichostatin A (TSA), a classic HDACI agent, against breast cancer cells.MethodsThe MCF-7, MDA-MB-231 and MCF-10A cell lines were evaluated with colony-forming and cell viability assays. Apoptosis and cell cycle distribution were detected by flow cytometry. Mitochondrial function was measured with biochemical assays, flow cytometry and transmission electron microscopy.ResultsTSA inhibited breast cancer cell viability and proliferation, without affecting MCF-10A cell. TSA-induced breast cancer cell apoptosis was initiated by G2-M arrest and depended on mitochondrial reactive oxygen species (ROS) produced subsequent to reduced mitochondrial respiratory chain activity. The enhanced mitochondrial ROS production and apoptosis in cancer cells were markedly attenuated by antioxidants, such as N-acetyl cysteine (NAC), reduced glutathione (GSH) and Vitamin C.ConclusionThe present study demonstrated that TSA-induced cell death by arresting cell cycle in G2-M phase and was dependent on production of mitochondria-derived ROS, which was derived from impaired mitochondrial respiratory chain.

Highlights

Despite growing advances in targeted therapies and screening techniques, breast cancer remains a leading cause of malignant mortality in women[1]
We examined the effect of Trichostatin A (TSA) on the viability of the MCF10A, MCF-7, MDA-MB-231 cell lines with a MTT assay
During treatment with serial concentrations of TSA, increasing H3 acetylation levels were observed in the MDA-MB-231 and MCF-7 breast cancer cells, but not in MCF-10A cell, indicating the TSA activity (Figure 1B)

Summary

Introduction

Despite growing advances in targeted therapies and screening techniques, breast cancer remains a leading cause of malignant mortality in women[1]. Several investigations have revealed that Trichostatin A (TSA), which was originally identified as a fungicidal antibiotic and was a classic HDACI agent[4], exhibited breast cancer cell toxicity in a dose-dependent manner both in vitro and in vivo[5,6,7]. Vorinostat and TSA have been shown to increase the transcription of BH3-only Bcl-2 family genes[9] and, as demonstrated, to disrupt mitochondrial membrane potential, induce the release of cytochrome c from the mitochondrial inter-membrane space to the cytoplasm and activate caspase-9 [10,11]. Reactive oxygen species (ROS) play an important role in apoptosis, and HDACIs promote ROS production, promoting apoptosis[12,13,14,15]. The mitochondrial respiratory chain is the vital site of ROS production, and complexes I and III have been suggested to be the major sources of ROS[17,18]

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