Trovafloxacin Enhances Lipopolysaccharide-Stimulated Production of Tumor Necrosis Factor-αby Macrophages: Role of the DNA Damage Response

Abstract

Trovafloxacin (TVX) is a drug that has caused idiosyncratic, druginduced liver injury (IDILI) in humans. In a murine model of IDILI, otherwise nontoxic doses of TVX and the inflammagen lipopolysaccharide (LPS) interacted to produce pronounced hepatocellular injury. The liver injury depended on a TVX-induced, small but significant prolongation of tumor necrosis factor-a (TNF) appearance in the plasma. The enhancement of TNF expression by TVX was reproduced in vitro in RAW 264.7 murine macrophages (RAW cells) stimulated with LPS. The current study was designed to identify the molecular target of TVX responsible for this response in RAW cells. An in silico analysis suggested a favorable binding profile of TVX to eukaryotic topoisomerase II-a (TopIIa), and a cellfree assay revealed that TVX inhibited eukaryotic TopIIa activity. Topoisomerase inhibition is known to lead to DNA damage, and TVX increased the DNA damage marker phosphorylated histone 2A.X in RAW cells. Moreover, TVX induced activation of the DNA damage sensor kinases, ataxia telangiectasia mutated (ATM) and Rad3-related (ATR). The ATR inhibitor NU6027 [6-(cyclohexylmethoxy)-5-nitrosopyrimidine-2,4-diamine] prevented the TVX-mediated increases in LPS-induced TNF mRNA and protein release, whereas a selective ATM inhibitor [2-(4-morpholinyl)-6-(1-thianthrenyl)-4H-pyran-4-one (KU55933)] was without effect. TVX prolonged TNF mRNA stability, and this effect was largely attenuated by NU6027. These results suggest that TVX can inhibit eukaryotic topoisomerase, leading to activation of ATR and potentiation of TNF release by macrophages, at least in part through increased mRNA stability. This off-target effect might contribute to the ability of TVX to precipitate IDILI in humans.

Metabolomic Study on Idiosyncratic Liver Injury Induced by Different Extracts of Polygonum multiflorum in Rats Integrated with Pattern Recognition and Enriched Pathways Analysis

“…Moreover, predicting idiosyncratic hepatotoxicity reactions has been a formidable challenge. The LPS model has been employed to evaluate IDILI successfully for several drugs in humans, such as trovafloxacin, sulindac, halothane, chlorpromazine, monocrotaline, ranitidine, diclofenac, and amiodarone in toxicological experiment assessment on the basis of the inflammatory stress hypothesis (Waring et al, 2006; Shaw et al, 2010; Roth and Ganey, 2011; Poulsen et al, 2014). The previous research constructed Heshouwu idiosyncratic hepatotoxicity, based on LPS animal models, has manifested Heshouwu could induce acute liver injury in rats cotreated with LPS approximate to the clinical equivalent dose, and the animal model should make it possible to assess idiosyncratic hepatotoxicity of the herb (Li et al, 2015).…”

“…The mechanism of IDILI is very complex, among which inflammation response plays a critical role. The inflammatory stress hypothesis has provided some of the first animal models of idiosyncratic hepatotoxicity (Roth and Ganey, 2011; Poulsen et al, 2014). Nontoxic dose of LPS precipitate modest inflammatory responses in mammals, resulting in increased susceptibility to toxicity from numerous hepatotoxic chemicals (Deng et al, 2009).…”

Trovafloxacin-induced replication stress sensitizes HepG2 cells to tumor necrosis factor-alpha-induced cytotoxicity mediated by extracellular signal-regulated kinase and ataxia telangiectasia and Rad3-related

“…In a cell-free system, TVX inhibited eukaryotic topoisomerase-IIα (Poulsen et al 2014), which is involved in DNA replication and cell cycle regulation (Larsen et al 1996). This can create a replication stress that initiates events involved in cell death and inhibition of proliferation.…”

The role of hepatic cytochrome P450s in the cytotoxicity of dronedarone

“…7), suggesting the parent dronedarone is more likely than its metabolites to interfere with topoisomerase IIα expression. It has been reported that some drugs and naturally occurring compounds inhibit topoisomerases II, leading to DNA damage and liver toxicity (Chen et al 2013; Poulsen et al 2014; Zhang et al 2015). In agreement with these studies, our current study highlights the role of topoisomerase IIα in drug-induced liver toxicity.…”