Dual actions of norathyriol as a new candidate hypouricaemic agent: uricosuric effects and xanthine oxidase inhibition

Abstract

Hyperuricaemia, which results from the overproduction and underexcretion of uric acid, has been linked with chronic renal dysfunction, cardiovascular diseases, diabetes and metabolic syndrome. However, available clinical drugs cannot simultaneously target the production and excretion of uric acid. Norathyriol, a natural xanthone, was expected to have the dual actions. We previously reported that norathyriol possessed potent anti-hyperuricaemic activity related to the inhibition of uric acid production. Here, we investigated the uricosuric actions in hyperuricaemic animals and explored the possible molecular mechanisms associated with renal urate transporters and xanthine oxidase (XO). The results showed that norathyriol (0.5–4.0 mg/kg) dose- and time-dependently decreased serum urate levels in uric acid-induced hyperuricaemic mice and markedly increased the fractional excretion of urate in oxonate-induced hyperuricaemic rats, demonstrating a promotion of urate excretion in the kidney. Further evidence showed that norathyriol markedly increased renal mRNA and protein expression of the secretory organic anion transporter 1 (OAT1) in hyperuricaemic mice and strengthened its transport function in vitro. Moreover, norathyriol also upregulated the mRNA expression of the secretory transporters OAT3, ATP-binding cassette transporter G2 and multidrug resistance protein 4, but not their protein expression. Changes in the expression of the reabsorptive transporters were not observed. This is the first report that norathyriol has uricosuric effects by targeting OAT1. Moreover, norathyriol significantly inhibited XO activity in an uncompetitive manner. Taken together, these findings suggest that norathyriol has the potential to be developed as a new anti-hyperuricaemic agent with dual actions that activate OAT1 and inhibit XO activity.

Introduction

Uric acid, a final product of purine metabolism in humans, is generated from the oxidation of hypoxanthine and xanthine by xanthine oxidase (XO) and is excreted by the kidney and intestinal tract. It exists largely as urate in body fluids at physiological pH. Its homeostasis depends on the balance of the production and excretion of uric acid. Overproduction and/or insufficient excretion of uric acid result in hyperuricaemia. Currently, hyperuricaemia is believed to be a key risk factor not only for gout but also for chronic nephritis, renal dysfunction, cardiovascular diseases, hypertension, diabetes and metabolic syndrome (Dalbeth et al., 2016; Zoccali and Mallamaci, 2013). Therefore, maintaining normal levels of this key factor is considered as an important strategy for the prevention and treatment of these diseases.

Management approaches for hyperuricaemia in gout commonly include XO inhibitors (e.g., allopurinol and febuxostat), uricosuric drugs (e.g., benzbromarone and probenecid) and uricase therapy. However, existing anti-hyperuricaemic agents possess some undesirable side effects, such as hypersensitivity towards allopurinol and hepatotoxicity associated with benzbromarone, which limit their clinical uses (Chaichian et al., 2014). More recently, lesinurad, a new uricosuric agent targeting urate transporter 1 (URAT1) in the kidney, has been approved by the U.S. Food and Drug Administration (Hoy, 2016) and the European Commission for monotherapy or in combination with XO inhibition for the management of gout. It has been suggested that the potential to combine two mechanisms of treatment, the inhibition of uric acid production with XO inhibitors and the promotion of uric acid excretion with uricosuric drugs, represents a powerful approach for the reduction of serum uric acid (Fleischmann et al., 2014). Currently, there are no anti-hyperuricaemic agents in clinical use that can target both the production and excretion of uric acid. Therefore, the development of novel compounds with dual hypouricaemic actions is highly warranted.

Norathyriol (1,3,6,7-tetrahydroxy-9H-xanthen-9-one, Fig. 1), a natural bioactive xanthone, is widely present in many medicinal plants including Mangifera indica L., Hypericum elegans, Gentianaceae and Tripterospermum lanceolatum, etc (Ko et al., 1991; Li et al., 2012; Lin et al., 1982). It has been reported that norathyriol possesses a wide variety of biological activities, such as antioxidation, anti-inflammation, antitumour, antiradiation, protein tyrosine phosphatase 1B inhibition and AMP-activated protein kinase activation (Chieli et al., 2009; Ding et al., 2014; Hsu et al., 2004; Lee et al., 1998; Lei et al., 2012; Li et al., 2012; Librowski et al., 2005; Wang et al., 2014). We recently reported that norathyriol has potent hypouricaemic actions in hyperuricaemic mice. Intragastrical administration of norathyriol at doses of 0.92, 1.85 and 3.7 mg/kg decreased serum urate levels by 27, 34 and 37%, respectively, which might be related to inhibiting the production of uric acid (Niu et al., 2016). However, the uricosuric actions and underling mechanisms have not been completely elucidated. In other words, we wonder whether norathyriol has the dual actions of inhibiting the generation and enhancing the excretion of uric acid. Moreover, the kidney plays an important role in the excretion of uric acid because it accounts for approximately two-thirds of urate elimination. Insufficient renal excretion is the dominant cause of hyperuricaemia in approximately 90% of affected individuals (Perez-Ruiz et al., 2015). Emerging evidence indicates that the reabsorption and secretion of urate in the kidney is controlled by a suite of apically and basolaterally expressed urate transporters that coexist along the length of the proximal renal tubule (Dalbeth et al., 2016). The altered expression and function of the urate transporters are suggested to be associated with hyperuricaemia (Terkeltaub, 2010). Therefore, in the current study, we studied the uricosuric actions in uric acid-induced hyperuricaemic mice and potassium oxonate (an uricase inhibitor)-induced hyperuricaemic rats; we also explored the possible molecular mechanisms, focusing on the renal urate transporters and XO.