Pharmacokinetic/pharmacodynamic modeling and simulation of dotinurad, a novel uricosuric agent, in healthy volunteers

Abstract This study aimed to investigate the pharmacokinetic and pharmacodynamic (PK/PD) profiles of dotinurad, a novel uricosuric agent, and to construct a PK/PD model to predict serum urate (SUA) levels after dotinurad administration in healthy men. PK/PD model was constructed using single‐dose study data considering the physiological features of urate handling. Model validation was performed by comparing the predicted SUA levels with the SUA levels in a multiple‐dose study. Dotinurad was absorbed rapidly, and its exposure increased proportionally in the tested dose ranges (0.5–20 mg) after a single‐dose administration. The PK model after oral administration was described using a one‐compartment model with first‐order absorption. Effects on SUA and renal urate excretion of dotinurad increased with dose escalation but were apparently saturable at a dose >5 mg. The simple maximal effect (E max) model was selected as the PD model of dotinurad on renal urate reabsorption, resulting in an estimated E max of 0.51. The plasma concentration at the half‐maximal effect of dotinurad was 196 ng/mL. Other PD parameters were calculated from the change in SUA level or urinary excretion of urate before and after dotinurad administration. The predicted SUA levels, using the PK/PD model, were well‐fitted with the observed values. The constructed PK/PD model of dotinurad appropriately described the profiles of dotinurad plasma concentrations and SUA level in multiple administration study.


| INTRODUC TI ON
Gout is the most common inflammatory arthritis caused by the deposition of monosodium urate crystals within the joints and around tissues due to chronic hyperuricemia. 1 A recent meta-analysis suggested that hyperuricemia is an independent risk factor of multiple metabolic syndromes, such as hypertension, chronic heart disease, chronic kidney disease (CKD), and type 2 diabetes mellitus. [2][3][4][5] However, it is not clear if hyperuricemia is a cause or a result of these diseases.
In humans, uric acid (urate) is formed as an end product of purine metabolism by catalyzing enzyme xanthine oxidoreductase (XOR).
Approximately two-thirds of the daily turnover of urate is accounted for by renal excretion, with the remaining one-third being excreted by non-renal excretion, mainly into the gut as feces. 6 The serum urate (SUA) level is determined by the balance between biosynthesis and elimination of urate in normal condition, that is, 4-6 mg/ dL (240-360 µmol/L), whereas SUA level in hyperuricemia exceeds 7 mg/dL (>420 µmol/L) due to the overproduction and/or inefficient renal excretion of urate. 6 For the treatment of pain and inflammation associated with acute flares, urate-lowering therapy (ULT) has been adopted as a pharmacological treatment for the prevention of acute gout flares in patients with gout. Several guidelines for the management of gout recommend that SUA level should be lowered sufficiently to improve the signs and symptoms, that is, <6 mg/dL as a minimum target and often <5 mg/dL. [7][8][9][10] Clinically, the available drugs for ULT are XOR inhibitors (XOI) and uricosuric agents. Most guidelines recommend the use of XOI, such as allopurinol, febuxostat, and topiroxostat (labeled only in Japan), as first-line ULT medications. These drugs lower SUA level effectively in most cases; however, many patients do not achieve the recommended SUA level with XOI monotherapy. 11 In such cases, guidelines recommend that an alternative or combination therapy of XOI with a uricosuric agent, such as benzbromarone or probenecid, should be considered. However, in 2003, benzbromarone was withdrawn from the market in many countries because of its risk of severe hepatotoxicity. 12 Although probenecid was approved in many countries such as USA, its widespread use is also limited due to its ineffectiveness, potential nephrotoxicity and significant drug-drug interaction potential with widely used drugs such as nonsteroidal anti-inflammatory drugs and antibiotics. Accordingly, a novel, safer and accessible uricosuric agent is required.
Dotinurad, also called as FYU-981, is a novel Selective Uric Acid Reabsorption Inhibitor (SURI) that shows potent inhibitory effects on the uptake of urate in human renal brush-border membrane vesicles. 13 In 90% of patients with gout, hyperuricemia results from reduced renal urate excretion 14 ; thus, uricosuric agents should effectively lower the level of SUA. However, excessive amounts of renal urate excretion is associated with a higher risk of nephrolithiasis. 15 In addition, excessive urate-lowering effects possibly cause the progression of various neurodegenerative disorders, such as Parkinson's disease, 16 Alzheimer's disease 17 and amyotrophic lateral sclerosis. 18 Given these possibilities, it is important to control SUA levels and renal excretion of urates within an adequate range during ULT, especially with uricosurics. It is important to construct a PK/PD model in order to maintain SUA level within an appropriate range. However, it is unknown if urate handling in patients differs from that in healthy people. In this study, we aimed to construct the PK/PD model of dotinurad and to predict SUA levels in healthy volunteers using phase 1 clinical study of dotinurad. Firstly, the pharmacokinetic (PK) and pharmacodynamic (PD) profiles of dotinurad in healthy male volunteers were evaluated. Secondly, a PK/PD model of dotinurad was constructed using derived data from the single ascending dose (SAD) study to predict SUA profiles in the multiple ascending dose (MAD) study. The models were assessed by comparing the predicted and observed values of dotinurad in the MAD study.

| Subjects
For the SAD study, a total of 54 subjects were enrolled, and they all completed the study (ClinicalTrials.gov identifier:. NCT02348307) except for one subject who dropped out due to domestic reason; for the MAD study, 18 subjects were enrolled, and all completed the study (NCT02348333). Subject demographics were comparable in both groups. All the subjects were healthy Japanese men (age range, 20-35 years; body mass index, ≥18.5 and <25.0 kg/m 2 ) for both studies.
These studies were conducted at an institute hospital in Japan.
The study protocol was conducted in accordance with the Declaration of Helsinki, the standards of the Japanese Pharmaceutical Affairs

Law, and International Conference on Harmonization and Good
Clinical Practice guidelines. The study protocols were approved by the institutional review board of the institute, and all subjects provided written informed consent to participate prior to the initiation of each study.

| Study design and sample collection
The SAD and MAD studies were randomized, double-blind, placebocontrolled, dose escalation, and parallel group comparison studies.

| Sample analysis
Plasma and urine dotinurad concentrations were analyzed using high-performance liquid chromatography with tandem mass spectrometry using the validation method at FUJI YAKUHIN CO., LTD.

| PK analysis
Plasma PK parameters of dotinurad were derived using Phoenix WinNonlin (version 6.2, Pharsight Corporation as part of Certara, St. Louis, MO). Plasma PK parameters included maximum observed plasma concentration (C max ), time to C max (T max ), area under the plasma concentration-time curve (AUC) from time 0 to 24 hour (AUC 0-24 ), time 0 to infinity (AUC 0-inf ), and terminal phase elimination half-life (t 1/2 ). AUC was calculated using the linear trapezoidal rule, and the apparent oral clearance (CL/F) was determined. The accumulation ratio (R AUC0-24 ) was calculated as the ratio of AUC 0-24 on day 4 or day 7 to AUC 0-24 on day 1.

| PD analysis
The area under the effect curve (AUEC) was calculated as the AUC of the SUA-time curve. The total urinary excretion amount of urate (Xu UA ) was calculated from the urate concentration in urine (Cu UA ) and urine volume during the sampling time. The renal clearance for urate (CLR UA ) was calculated as Xu UA /AUEC.

| PK/PD modeling of dotinurad
Based on the observed mono-exponential decline of the plasma concentrations of dotinurad, the %CV of estimated parameters, and the model comparison analysis using Akaike's information criterion, a one-compartment model with first-order absorption was selected as the PK model of dotinurad.
To predict the SUA level after dotinurad administration in the MAD study, a simple PD model of dotinurad on urate clearance was constructed based on the physiological features of urate handling in the serum and urine. 6 Given the uricosuric effect of dotinurad (see Results), the scheme of the PD model of dotinurad is shown in Figure 1.
This model was constructed under the following assumptions: 1. Before dotinurad administration, the amount of urate production (shown as "Synthesis," involving the absorption of purine body from food intake) is a constant value identical to the sum of the renal and non-renal (mainly into feces from the gut) urate excretion.
2. The ratio of the renal to non-renal urate excretion is 2:1 in the normal state, that is, before the administration of dotinurad. CLR UA and AUEC before and after the administration of dotinurad were calculated as the mean or total value during 24 hours respectively. ∆SUA was the difference between SUA at 0 and that at 24 hours after the administration of dotinurad. ∆Xu UA and ∆Xf UA were calculated from the difference of Xu UA or Xf UA for 24 hours between before and after the administration of dotinurad. The baseline values of Xf UA , CLGut UA and "Synthesis" were estimated from Xu UA and CLR UA before the administration of dotinurad using the following equations:

| Model simulation
Using the fixed PK model of dotinurad and estimated PD parameters, the SUA-time profile was predicted after the multiple administrations of dotinurad. The mean observed baseline SUA and CLR UA were used as initial values. Additionally, the initial UA pool was calculated using Equation (1). CLR UA was estimated every hour, and Xu UA and Xf UA during −24 to 0 hours, 0-24 hours and 24-48 hours were estimated using the SUA and CLR UA or CLGut UA values before 1 hour of dotinurad administration. Finally, SUA level was estimated from Equation (1), using "Synthesis" in the scheme and Vd UA as the constant value. Model confirmation was performed by comparing the predicted SUA-time curve with the observed values in the MAD study.

| Statistical analysis
Observed PK and PD parameters were summarized using descriptive statistics. Each datum was represented as the mean + or ±standard deviation (SD). Estimated PK and PD parameters were represented as the mean and %CV. SAS software (version 9.2, SAS Institute Inc, Cary, NC) was used to perform statistical analyses. Dose-proportional assessment for C max and AUC was performed using a power model as follows: where slope b is close to unity, and the relationship between dose and PK parameters was concluded to be dose proportional within the dose range studied. The effect of food intake on PK parameters was analyzed by paired t-test.

| PK of dotinurad in healthy male volunteers
In the SAD study, plasma concentrations of dotinurad in fasted volunteers increased in a dose-proportional manner ( Figure 2, Table 1).  (Table 2). Other PK parameters were comparable between day 4 and day 7 at each dose.

| SUA-lowering effect and uricosuric effect of dotinurad in the SAD study
The mean (SD) baseline SUA, Xu UA, and CLR UA values of all the subjects were 5.71 (0.75) mg/dL, 575 (100) mg/day, and 7.10 (1.51)    For PD parameters, the mean value (%CV) of Vd UA was estimated as 16.0 (36.6) L from Equation (4) and f reabs as 0.943 (0.6) from

| PK/PD parameter estimation and prediction of SUA level after the multiple administrations of dotinurad
Equation (9). The effect of dotinurad on CLR UA were also estimated with 196 (11.9) ng/mL as the EC 50 and 0.51 (4.6) as the E max .
In this study, the observed and predicted SUA levels were compared, and the constructed PK/PD model was considered able to predict the observed SUA level after multiple dose administrations of dotinurad, although the predicted values were slightly underestimated at the 2-mg dose (Figure 4).

| D ISCUSS I ON
This study evaluated the PK and PD profiles of single increasing doses of dotinurad, a novel uricosuric agent, in healthy male subjects. It also established a PK/PD model for the prediction of SUA levels after multiple administration of dotinurad, based on the results of a single administration study.  been identified, 19 and one of such transporters is URAT1. 20 URAT1 is involved primarily in the renal apical uptake of urate, and it is the primary target of most recent uricosuric drugs. In fact, dotinurad inhibited urate transport in a concentration-dependent manner in human URAT1-overexpressed MDCKII cells (IC 50 = 37.2 nmol/L) and exhibited little inhibitory effect on XOR activity in vitro. 21 In the present study, after the single oral administration of dotinurad, SUA level was decreased rapidly, and the Xu UA and CLR UA were increased at all the tested doses. From these results, dotinurad effectively lowered SUA levels in human, and this effect was attributed to its uricosuric effect.
In the SAD study, it is worthy of note that although the C max and AUC of dotinurad, at up to 20-mg dose, were increased dose proportionally, a decrease in SUA level and an increase in urinary urate excretion were almost maximum at the 5-mg dose. Therefore, the uricosuric effect of dotinurad could be represented by a saturable and simple E max model.

ACK N OWLED G EM ENTS
We thank Kazuma Sekine, a former employee of FUJI YAKUHIN Co., Ltd, for contributing to PK/PD model analysis. We also appreciate the staff of Kitasato University Kitasato Institute Hospital.

D I SCLOS U R E S
Dr Ikumi Tami serve as a consultant for FUJI YAKUHIN CO, LTD. This work was supported by FUJI YAKUHIN CO, LTD.

AUTH O R CO NTR I B UTI O N S
Takako Igarashi and Koichi Omura analyzed plasma dotinurad con-

DATA AVA I L A B I L I T Y S TAT E M E N T
The data that support the findings of this study are available from the corresponding author upon reasonable request.