Xanthine oxidoreductase inhibitor topiroxostat ameliorates podocyte injury by inhibiting the reduction of nephrin and podoplanin.

BACKGROUND: Topiroxostat, an inhibitor of xanthine oxidoreductase (XOR) was shown to reduce urinary albumin excretion of hyperuricemic patients with chronic kidney disease. However, its pharmacological mechanism is not well understood. In this study, we examined the effects of topiroxostat on glomerular podocytes. Podocyte is characterized by foot process and a unique cell-cell junction slit diaphragm functioning as a final barrier to prevent proteinuria. METHODS: The effects of topiroxostat on the expressions of podocyte functional molecules were analysed in db/db mice, a diabetic nephropathy model, anti-nephrin antibody-induced rat podocyte injury model and cultured podocytes treated with adriamycin. RESULTS: Topiroxostat treatment ameliorated albuminuria in db/db mice. The expression of desmin, a podocyte injury marker was increased, and nephrin and podocin, key molecules of slit diaphragm, and podoplanin, an essential molecule in maintaining foot process were downregulated in db/db mice. Topiroxostat treatment prevented the alterations in the expressions of these molecules in db/db mice. XOR activity in kidney was increased in rats with anti-nephrin antibody-induced podocyte injury. Topiroxostat treatment reduced XOR activity and restored the decreased expression of nephrin, podocin and podoplanin in the podocyte injury. Furthermore, topiroxostat enhanced the expression of podoplanin in injured human cultured podocytes. CONCLUSIONS: Podocyte injury was evident in db/db mice. Topiroxostat ameliorated albuminuria in diabetic nephropathy model by preventing podocyte injury. Increase of XOR activity in kidney contributes to development of podocyte injury caused by stimulation to slit diaphragm. Topiroxostat has an effect to stabilize slit diaphragm and foot processes by inhibiting the reduction of nephrin, podocin and podoplanin.

Solid Nanocrystals of Rebamipide Promote Recovery from Indomethacin-Induced Gastrointestinal Bleeding.

Indomethacin (IMC)-induced gastrointestinal (GI) injuries are more common in rheumatoid arthritis (RA) patients than in other IMC users, and the overexpression of nitric oxide (NO) via inducible NO synthase (iNOS) is related to the seriousness of IMC-induced GI injuries. However, sufficient strategies to prevent IMC-induced GI injuries have not yet been established. In this study, we designed dispersions of rebamipide (RBM) solid nanocrystals (particle size: 30-190 nm) by a bead mill method (RBM-NDs), and investigated whether the oral administration of RBM-NDs is useful to prevent IMC-induced GI injuries. The RBM nanocrystals were spherical and had a solubility 4.71-fold greater than dispersions of traditional RBM powder (RBM-TDs). In addition, the RBM-NDs were stable for 1 month after preparation. The RBM contents in the stomach, jejunum, and ileum of rats orally administered RBM-NDs were significantly higher than in rats administered RBM-TDs. Moreover, the oral administration of RBM-NDs decreased the NO levels via iNOS and area of the GI lesions in IMC-stimulated RA (adjuvant-induced arthritis rat) rats in comparison with the oral administration of RBM-TDs. Thus, we show that the oral administration of RBM-NDs provides a high drug supply to the GI mucosa, resulting in a therapeutic effect on IMC-induced GI injuries. Solid nanocrystalline RBM preparations may offer effective therapy for RA patients.

Multicenter, Open-Label Study of Long-Term Topiroxostat (FYX-051) Administration in Japanese Hyperuricemic Patients with or Without Gout.

BACKGROUND AND OBJECTIVES: Topiroxostat-a novel selective xanthine oxidoreductase inhibitor-has been reported to reduce serum urate levels. The purpose of this study was to assess the efficacy and safety of long-term topiroxostat administration in Japanese hyperuricemic patients with or without gout. METHODS: This multicenter, open-label study evaluated the efficacy and safety of long-term twice-daily oral topiroxostat administration in patients with or  without gout. The initial topiroxostat dosage was 40-80 mg/day, and the maintenance dosage was 120 mg/day, which was increased to 240 mg/day at 40 mg increments if the serum urate level exceeded 6.0 mg/dL. RESULTS: Serum urate level, which was the primary endpoint, decreased stably over time and showed significant reduction on the final visit (38.44% ± 13.34%) compared with that at the baseline. Both urinary albumin/creatinine ratio and mean blood pressure significantly improved. The overall incidence rate of adverse drug reactions to topiroxostat was 67.8%; on the final visit, the rate of adverse drug reactions was 66.7% with 120 mg/day, 72.2% with 160 mg/day, 53.8% with ≥ 200 mg/day, and 100% with the other dosages. On the final visit, the incidence of gouty arthritis, for which a causal relationship with topiroxostat could not be ruled out, was 4.1% overall, 4.8% with 120 mg/day, 0% with 160 mg/day, and 7.7% with ≥ 200 mg/day. CONCLUSIONS: We verified the efficacy and safety of 58-week oral topiroxostat administration at stepwise increments to up to 240 mg/day. STUDY REGISTRATION: JAPIC CTI-101068.

Uric acid-lowering and renoprotective effects of topiroxostat, a selective xanthine oxidoreductase inhibitor, in patients with diabetic nephropathy and hyperuricemia: a randomized, double-blind, placebo-controlled, parallel-group study (UPWARD study).

BACKGROUND: Hyperuricemia is supposed to be an independent risk factor for kidney dysfunction in diabetic patients. We attempted to examine the uric acid-lowering effect and the renoprotective effect of topiroxostat, a selective xanthine oxidoreductase inhibitor, in patients with diabetic nephropathy and hyperuricemia in this pilot study. METHODS: The study design was randomized, double-blind, placebo-controlled, parallel-group study. A total of 65 patients with hyperuricemia and diabetic nephropathy with microalbuminuria were enrolled and assigned to either the topiroxostat group or the placebo group. Topiroxostat (stepwise dosing from 40 to 160 mg/day) or matching placebo was administered BID for 28 weeks. The primary endpoint was a change in the urinary albumin-to-creatinine ratio in the first-morning-void urine sample. Secondary endpoints were changes in the estimated glomerular filtration rate and the serum uric acid level. RESULTS: At 28 weeks, there was no significant difference in the percent change from baseline in the urinary albumin-to-creatinine ratio between the two groups (topiroxostat: 0 vs. placebo: 17%, p = 0.3206), but the changes in the estimated glomerular filtration rate (- 0.2 vs. - 4.0 mL/min/1.73 m2, p = 0.0303) and the serum uric acid level (- 2.94 vs. - 0.20 mg/dL, p < 0.0001) were significantly different between the topiroxostat and placebo groups. Gouty arthritis occurred in 1 patient in the placebo group and no patients in the topiroxostat group. CONCLUSION: These findings support that diabetic nephropathy combined with hyperuricemia may be associated with kidney dysfunctions. Topiroxostat provides strict control of the serum uric acid level preventing decline of eGFR in these patients.

Effects of topiroxostat and febuxostat on urinary albumin excretion and plasma xanthine oxidoreductase activity in db/db mice.

Topiroxostat, a xanthine oxidoreductase (XOR) inhibitor, has been shown to decrease the urinary albumin-to-creatinine ratio compared with placebo in hyperuricemic patients with stage 3 chronic kidney disease. Thus, we aimed to ascertain the albuminuria-lowering effect of topiroxostat in diabetic mouse. Db/db mice were fed standard diets with or without topiroxostat (0.1, 0.3, 1, and 3mg/kg/day) and febuxostat (0.1, 0.3, and 1mg/kg/day) for four weeks. Urinary albumin and purine bodies levels, XOR activities, and drug concentrations in the liver, kidney, and plasma were measured. Moreover, the XOR inhibitory activity of each XOR inhibitor was evaluated with or without an exogenous protein in vitro. Topiroxostat decreased dose-dependently the urinary albumin excretion, but febuxostat did not show such a tendency. Treatment with topiroxostat inhibited plasma XOR activity with dose-dependent increase in plasma purine levels, which was not observed by febuxostat. Pharmacokinetic/pharmacodynamic analysis revealed that topiroxostat and febuxostat concentration in each tissue showed a good correlation with both the hypouricemic effect and plasma drug concentration, whereas the change in albuminuria correlated neither with the change in uric acid nor with drug concentration in plasma. However, the change in urinary albumin and plasma XOR activity showed good correlation in topiroxostat group. The 50% inhibitory concentration (IC50 value) of febuxostat against plasma XOR in vitro was 12-fold higher than that of topiroxostat, and increased by approximately 13-fold by interfering with an exogenous protein. Topiroxostat caused reduced urinary albumin excretion, in which potent inhibition of the plasma XOR activity might be involved.

Amino Acid esters prevent thermal inactivation and aggregation of lysozyme.

Small potent inhibitors of aggregation are eagerly demanded for preventing the inactivation of proteins. This paper shows that amino acid esters (AAEs) prevent heat-induced aggregation and inactivation of hen egg lysozyme. Lysozyme was completely inactivated (<1% original activity) during heat treatment at 98 degrees C for 30 min in a solution containing 0.2 mg/mL lysozyme in 50 mM Na-phosphate buffer (pH 6.5). The residual activities only slightly increased (<5%) in the presence of 100 mM commonly used additives such as arginine, guanidine, urea, and sugars. However, in the presence of 100 mM AAEs, the residual activities were >60% and no aggregates were observed during the heat treatment at 98 degrees C for 30 min. This fact provides new information on the scaffold for designing additives to prevent heat-induced aggregation.

High temperature increases the refolding yield of reduced lysozyme: implication for the productive process for folding.

Misfolding poses a serious problem in the biotechnological field in obtaining the active protein from inclusion bodies. Here we show that high temperature increases the refolding yield of reduced lyosyzme by a simple dilution method. The refolding yields at 98 degrees C were three times higher than those at 20 degrees C in the solutions tested, which is related to the fact that the thermally unfolded state of lysozyme is a more productive form for folding than the denaturant-induced fully unfolded state. The thermal-assisted refolding could be used for various reduced and denatured proteins as a result of its simplicity and low cost.