Abstract
Aims: The effect of uric acid (UA)-lowering therapy with xanthine oxidoreductase (XOR) inhibitors on the development of cardiovascular disease requires further investigation. This study aimed to evaluate the long-term effects of febuxostat on arterial stiffness, focusing on liver function.
Methods: The PRIZE study involved random assignment of patients with asymptomatic hyperuricemia to receive either add-on febuxostat treatment (febuxostat group) or non-pharmacological treatment (control group). Of the 514 participants, 23 and 14 patients in the febuxostat and control groups, respectively, underwent assessment of arterial stiffness using the cardio-ankle vascular index (CAVI). The participants in each group were further grouped on the basis of their baseline alanine aminotransferase (ALT) or aspartate aminotransferase (AST) levels (above or below the media value or 30 U/L). The primary endpoint was the change in the CAVI from baseline to 12 and 24 months.
Results: Overall, no significant differences were found between the control and febuxostat groups in the least-squares mean estimates of changes in CAVI at 24 months (mean between-group difference, −0.41 [95% CI, −1.05 to 0.23];p=0.204). However, there were significant differences in participants with higher baseline ALT or AST levels above 30 U/L at 24 months (mean between-group difference, −1.12 [95% CI, −2.23 to −0.01];p=0.048 for ALT ≥ 30 U/L and −1.08 [95% CI, −2.13 to −0.03];p=0.044 for AST ≥ 30 U/L).
Conclusions: Two-year treatment with febuxostat demonstrated a beneficial effect on CAVI in patients with hyperuricemia and liver dysfunction.
Keywords: Hyperuricemia, Hypertension, Arterial stiffness, Xanthine oxidoreductase inhibitors, Liver dysfunction
Yusuke Kawachi and Yuya Fujishima contributed equally to this manuscript.
Introduction
Hyperuricemia, which is defined by a serum uric acid (UA) concentration of ＞7.0 mg/dL, has been identified as a potential risk factor for cardiovascular disease (CVD) in addition to gout 1 , 2) . The population of patients with hyperuricemia is heterogeneous and can be categorized into reduced renal/extrarenal excretion, overproduction, and mixed types 3) . Specifically, the liver overproduction type is associated with visceral fat-based metabolic syndrome and nonalcoholic fatty liver disease (NAFLD), recently renamed metabolic dysfunction-associated steatotic liver disease (MASLD), in which xanthine oxidoreductase (XOR) is assumed to play a central role 4) .
XOR is the rate-limiting enzyme that catalyzes the production of UA from hypoxanthine and xanthine and is the pharmacological target of anti-hyperuricemic agents such as allopurinol, febuxostat, and topiroxostat. Through purine catabolism, XOR generates reactive oxygen species (ROS), which bind to the apical surface of vascular endothelial cells 5) . Therefore, XOR-dependent vascular ROS have been suggested to be one of the underlying mechanisms responsible for the endothelial dysfunction associated with hyperuricemia 6 , 7) . Human XOR expression is primarily detected in the liver, lungs, and intestines 8) , and XOR protein and activity are also detected in the circulation 9) . We recently reported a significant increase in plasma XOR activity in human liver disease conditions, which may potentially reflect excessive hepatic XOR leakage 10 , 11) . Furthermore, the suppression of elevated plasma XOR activity by XOR inhibitors was associated with attenuation of neointimal proliferation in mice 11) and improvement of arterial stiffness parameters in human clinical settings 12) . Therefore, XOR inhibitors may prevent or delay cardiovascular complications beyond their UA-lowering effects, particularly in patients with NAFLD/MASLD.
Arterial stiffness is a crucial cardiovascular health parameter that reflects the rigidity of the arteries and their ability to accommodate changes in blood pressure 13) . Increased arterial stiffness is an independent predictor of adverse cardiovascular events and an important clinical indicator of cardiovascular risk 13) . Among the existing parameters for arterial stiffness, the cardio-ankle vascular index (CAVI) is a noninvasive indicator of total arterial stiffness from the aorta to the ankle that shows minimal influence of blood pressure at the time of measurement 14) . The Program of Vascular Evaluation under Uric Acid Control by Xanthine Oxidase Inhibitor, Febuxostat: Multicenter, Randomized Controlled (PRIZE) study is a prospective, multicenter study designed to assess the inhibitory effect of febuxostat on the progression of carotid intima-media thickness (IMT) over a 2-year follow-up period 15) . However, the study showed no significant difference in the primary endpoint or change in carotid IMT between patients treated with febuxostat and those not treated with pharmacotherapy. Because plasma XOR activity increases with liver dysfunction, we conducted a sub-analysis of the PRIZE study, dividing the study participants into subgroups on the basis of their baseline alanine aminotransferase (ALT) or aspartate aminotransferase (AST) levels.
Aim
This study examined the differential effect of long-term febuxostat treatment on cardiovascular outcomes, including CAVI, in asymptomatic hyperuricemic patients based on baseline liver transaminase levels.
Methods
Study Design
The PRIZE study was a multicenter, prospective, randomized, open-label, and blinded-endpoint clinical trial. The details of this study have been described previously 15) . The study protocol was approved by the local institutional review boards and independent ethics committees at all sites. The study was conducted in full compliance with the Declaration of Helsinki and in accordance with the Ethical Guidelines for Medical and Health Research Involving Human Subjects established by the Ministry of Health, Labor, and Welfare and the Ministry of Education, Culture, Sports, Science, and Technology in Japan. Before enrollment, all participants were given a full explanation of the study plan and provided written informed consent.
Individuals eligible for the study were adults (age ≥ 20 years) who had both asymptomatic hyperuricemia with serum UA level ＞7.0 mg/dL and maximum common carotid artery (CCA)-IMT ≥ 1.1 mm as measured at eligibility assessment; this CCA-IMT range was defined as a carotid artery plaque (localized protruding lesion) according to the guidelines of the Japan Society of Ultrasonic in Medicine and the Japan Academy of Neurosonology 16) . After confirming their eligibility and reviewing their medical histories, the patients were randomized equally to either the febuxostat group (10–60 mg daily) or the control group (non-pharmacological treatment of hyperuricemia). All patients were followed up with study visits scheduled at 1, 2, 3, 6, 12, and 24 months after their baseline visit. Carotid artery ultrasonography and assessments of arterial stiffness were performed at each clinical site at baseline and after 12 and 24 months (or at premature termination). Details of the inclusion and exclusion criteria have been described previously 15) .
This study was conducted in accordance with the Ethical Guidelines for Medical and Biological Research Involving Human Subjects (Japan) and the Declaration of Helsinki. A series of sub-analyses of the PRIZE study, including this work, were approved by the Ethics Committee of Saga University Hospital (2020-05-R01) and subsequently registered (UMIN000041322). Informed consent was obtained from all participants included in this study.
Randomization and Intervention
Patients were randomized in a 1:1 ratio to the febuxostat or control group at the automatic web-based PRIZE Data Center. All participants in both groups underwent appropriate lifestyle modifications for hyperuricemia, such as healthy diet and exercise therapy, and these modifications continued during the study. Based on the protocol, patients assigned to the febuxostat group received an initial dose of 10 mg daily, which was titrated to 20 mg daily at one month and 40 mg daily at two months. A dose of 40 mg daily was targeted as the maintenance level; however, at 3 months or later, the dosage could be increased to 60 mg daily. If serum UA levels decreased to ≤ 2.0 mg/dL during the study, the maintenance dosage was decreased by 20 mg.
Study Participants
As reported previously 15) , 239 and 244 patients were included in the febuxostat and control groups, respectively, in the PRIZE study. In the PRIZE study, the assessment of CAVI was optional and was performed at participating sites where it was measurable. Measurements were unbiased and independent of baseline characteristics. Then, among the enrolled patients, 54 patients underwent successful CAVI measurements at baseline. Patients without available CAVI data at either 12 or 24 months were excluded. Ultimately, 23 patients in the febuxostat group and 14 patients in the control group were included in this sub-analysis. Patients in each group were further divided into groups according to baseline ALT or AST levels above or below the median value (18 U/L for ALT and 24 U/L for AST) and above or below 30 U/L (the normal-high value).
Study Endpoints
The study endpoint of this sub-analysis was the change in CAVI from baseline to 12 and 24 months; additional endpoints were changes in blood pressure, pulse pressure, CCA mean IMT, serum UA level, and serum ALT level from baseline to 12 and 24 months.
Assessments
CAVI was measured using a CAVI device (Vasera VS3000) at 10 clinical sites participating in the PRIZE study. Examinations were performed after a 5-min rest period. The cuff pressure was maintained at 50 mmHg to minimize its effect on hemodynamics. The blood pressure was then measured. CAVI was determined using the following formula: CAVI=a [(2ρ/ΔP)×ln ( Ps / Pd ) PWV2]＋b, where a and b are constants, ρ is blood density, Ps is systolic blood pressure, Pd is diastolic blood pressure, ΔP is Ps−Pd, and PWV is pulse wave velocity 14) . CAVI was measured on both sides of the body (right and left), and the mean value was used for analysis. To measure CAVI, the ankle-brachial index (ABI) was obtained on both sides. When the ABI was ＜0.95, the CAVI on that side was not included in the analyses.
Blood pressure was measured in an office setting using either auscultation or oscillometry (the choice of method was left to each facility), and hypertension was defined based on the Japanese Society of Hypertension guidelines for the management of hypertension (JSH 2014) 17) . Mean blood pressure was calculated as (pulse pressure/3 plus diastolic blood pressure). Pulse pressure was defined as systolic blood pressure minus diastolic blood pressure (DBP).
The protocol and method for measuring carotid IMT have been described previously 18) . Based on a standardized protocol 19) , high-resolution carotid ultrasonography using a standard system equipped with a ＞7.5-MHz linear transducer was performed at each site in a blinded manner by a trained expert sonographer who had attended a lecture on acquiring images to capture the carotid IMT. All imaging data were stored as JPEG files and sent to the core imaging laboratory, where an expert analyst measured IMT values in a blinded manner using an automated IMT measurement software program (Vascular Research Tools 5, Medical Imaging Applications LLC, Coralville, IA, USA) 20) .
Statistical Analysis
For the baseline variables, summary statistics were expressed as frequency and proportion for categorical data and median (interquartile range [IQR]) for continuous variables. For the primary analysis, the mean change from baseline to 12 and 24 months and its 95% CI, estimated using a mixed-effects model for repeated measures, were calculated for each parameter, adjusting for age, sex, and corresponding baseline values (and change in mean blood pressure for CAVI), including a cross-product term between the time and treatment groups, and the changes were compared between the treatment groups. Effect modification by baseline AST or ALT concentration was assessed using an interaction analysis, where a cross-product term between treatment groups and a binary variable of AST or ALT values indicating patient subgroups were added to the above multivariable regression models. All p values were two-sided with a significance threshold of 0.05, and no adjustments were performed for multiple comparisons. All statistical analyses were performed using the R 4.2.2 (R Foundation for Statistical Computing, Vienna, Austria)
Results
Baseline Characteristics of the Study Participants
Table 1 summarizes the baseline clinical characteristics of the study participants. For the 37 patients with CAVI data (11 women [29.7%] and 26 men [70.3%]; n=23 in the febuxostat group and n=14 in the control group), the median age was 74 years (IQR, 66–78 years). The median values of baseline CAVI were 9.25 (IQR, 8.15–10.1) in the febuxostat group and 9.70 (IQR, 8.15–11.45) in the control group, respectively. In the febuxostat group, the daily dose of febuxostat was 10 mg in five patients (21.7%), 20 mg in one patient (4.3%), 30 mg in three patients (13.0%), 40 mg in 13 patients (56.5%), and 60 mg in one patient (4.3%); all patients received febuxostat for 24 months.
Table 1. Baseline clinical characteristics of the population with cardio-ankle vascular index data.
| Variable | Overall | Febuxostat (n = 23) | Control (n = 14) | |
|---|---|---|---|---|
| Age (years), median [IQR] | 74 [66, 78] | 73 [64, 77] | 74.5 [70, 78] | |
| Sex, n (%) | female | 11 (29.7) | 8 (34.8) | 3 (21.4) |
| male | 26 (70.3) | 15 (65.2) | 11 (78.6) | |
| Body mass index (kg/m2), median [IQR] | 26.2 [23.4, 27.5] | 26.3 [23.4, 27.5] | 26.2 [20.9, 28.3] | |
| Smoking habit, n (%) | current | 2 (5.4) | 2 (8.7) | 0 (0) |
| past | 16 (43.2) | 9 (39.1) | 7 (50.0) | |
| never | 18 (48.6) | 12 (52.2) | 6 (42.9) | |
| unknown | 1 (2.7) | 0 (0) | 1 (7.1) | |
| Systolic blood pressure (mmHg), median [IQR] | 126 [116, 134] | 124 [115, 134] | 130 [122, 134] | |
| Diastolic blood pressure (mmHg), median [IQR] | 70 [64, 80] | 69 [64, 78] | 77.5 [64, 86] | |
| CAVI, median [IQR] | 9.25 [8.15, 10.1] | 9.20 [8.15, 10.05] | 9.70 [8.15, 11.45] | |
| ALT (U/L), median [IQR] | 18.0 [15.0, 34.0] | 18.0 [15.0, 33.0] | 19.5 [13.0, 38.0] | |
| AST (U/L), median [IQR] | 24.0 [19.0, 31.0] | 23.0 [19.0, 27.0] | 28.5 [19.0, 41.0] | |
| Fib-4 index, median [IQR] | 2.21 [1.54, 2.70] | 2.14 [1.48, 2.50] | 2.45 [1.68, 3.27] | |
| Hypertension, n (%) | yes | 33 (89.2) | 22 (95.7) | 11 (78.6) |
| no | 4 (10.8) | 1 (4.3) | 3 (21.4) | |
| Diabetes, n (%) | yes | 15 (40.5) | 9 (39.1) | 6 (42.9) |
| no | 22 (59.5) | 14 (60.9) | 8 (57.1) | |
| Dyslipidemia, n (%) | yes | 24 (64.9) | 15 (65.2) | 9 (64.3) |
| no | 13 (35.1) | 8 (34.8) | 5 (35.7) | |
| Previous gouty arthritis, n (%) | yes | 1 (2.7) | 1 (4.3) | 0 (0) |
| no | 36 (97.3) | 22 (95.7) | 14 (100.0) | |
| Myocardial infarction, n (%) | yes | 1 (2.7) | 0 (0) | 1 (7.1) |
| no | 36 (97.3) | 23 (100) | 13 (92.9) | |
| PCI, n (%) | yes | 4 (10.8) | 2 (8.7) | 2 (14.3) |
| no | 33 (89.2) | 21 (91.3) | 12 (85.7) | |
| CABG, n (%) | yes | 2 (5.4) | 1 (4.3) | 1 (7.1) |
| no | 35 (94.6) | 22 (95.7) | 13 (92.9) | |
| Stroke, n (%) | yes | 2 (5.4) | 1 (4.3) | 1 (7.1) |
| no | 35 (94.6) | 22 (95.7) | 13 (92.9) | |
| Heart Failure, n (%) | yes | 4 (10.8) | 3 (13.0) | 1 (7.1) |
| no | 33 (89.2) | 20 (87.0) | 13 (92.9) | |
| Any antihypertensive agent, n (%) | yes | 32 (86.5) | 21 (91.3) | 11 (78.6) |
| no | 5 (13.5) | 2 (8.7) | 3 (21.4) | |
| Renin-angiotensin system inhibitor, n (%) | yes | 24 (64.9) | 16 (69.6) | 8 (57.1) |
| no | 13 (35.1) | 7 (30.4) | 6 (42.9) | |
| ARB, n (%) | yes | 19 (51.4) | 13 (56.5) | 6 (42.9) |
| no | 18 (48.6) | 10 (43.5) | 8 (57.1) | |
| ACE inhibitor, n (%) | yes | 5 (13.5) | 3 (13) | 2 (14.3) |
| no | 32 (86.5) | 20 (87) | 12 (85.7) | |
| Calcium channel blocker, n (%) | yes | 25 (67.6) | 17 (73.9) | 8 (57.1) |
| no | 12 (32.4) | 6 (26.1) | 6 (42.9) | |
| β-Blocker, n (%) | yes | 12 (32.4) | 6 (26.1) | 6 (42.9) |
| no | 25 (67.6) | 17 (73.9) | 8 (57.1) | |
| Diuretic, n (%) | yes | 9 (24.3) | 7 (30.4) | 2 (14.3) |
| no | 28 (75.7) | 16 (69.6) | 12 (85.7) | |
| Any lipid-reducing agent, n (%) | yes | 18 (48.6) | 10 (43.5) | 8 (57.1) |
| no | 19 (51.4) | 13 (56.5) | 6 (42.9) | |
| Statins, n (%) | yes | 17 (45.9) | 9 (39.1) | 8 (57.1) |
| no | 20 (54.1) | 14 (60.9) | 6 (42.9) | |
| Ezetimibe, n (%) | yes | 0 (0) | 0 (0) | 0 (0) |
| no | 37 (100) | 23 (100) | 14 (100) | |
| Any antiplatelet agent, n (%) | yes | 11 (29.7) | 8 (34.8) | 3 (21.4) |
| no | 26 (70.3) | 15 (65.2) | 11 (78.6) | |
| Aspirin, n (%) | yes | 9 (24.3) | 7 (30.4) | 2 (14.3) |
| no | 28 (75.7) | 16 (69.6) | 12 (85.7) |
ACE, angiotensin-converting enzyme inhibitor; ALT, alanine aminotransferase; ARB, angiotensin II receptor blocker; AST, aspartate aminotransferase; CABG, coronary artery bypass grafting; CAVI, cardio-ankle vascular index; eGFR, estimated glomerular filtration rate; Fib-4, fibrosis 4; IQR, interquartile range; PCI, percutaneous coronary intervention.
CAVI
Fig.1 shows the estimated time course of changes in CAVI from baseline to 12 and 24 months. Overall, the control and febuxostat groups showed no significant differences in the least-squares means of the estimated changes in CAVI during the study period (mean between-group difference, −0.33 [95% CI, −0.96 to 0.30]; p=0.304 at 12 months and −0.41 [95% CI, −1.05 to 0.23]; p=0.204 at 24 months). ( Fig.1A ) . However, among patients with baseline ALT levels above 30 U/L, CAVI showed a significant durational increase in the control group at 24 months, whereas it remained unchanged in the febuxostat group ( Fig.1B ) . In addition, the change in CAVI from baseline to 24 months for patients with ALT ( Fig.1B , the right panel) and AST ( Fig.1C , the right panel) levels ≥ 30 U/L showed a significant between-group difference (mean between-group difference, −1.12 [95% CI, −2.23 to −0.01]; p=0.048 for ALT ≥ 30 U/L and −1.08 [95% CI, −2.13 to −0.03]; p=0.044 for AST ≥ 30 U/L). Similarly, using the median values as cutoffs, a significant durational increase in CAVI at 24 months was observed in patients in the control group with both higher ALT ( Supplemental Fig.1A , the right panel) and AST ( Supplemental Fig.1B , the right panel) levels. In addition, significant between-group differences in the changes in CAVI at 24 months were observed in patients with AST levels above the median value (mean between-group difference, −0.88 [95% CI, −1.69 to −0.06]; p=0.036) ( Supplemental Fig.1B , the right panel).
Fig.1. Changes in the cardio-ankle vascular index from baseline to month 24.
(A) Changes in the cardio-ankle vascular index (CAVI) in the total population with CAVI data. (B and C) Patients were divided into groups according to baseline alanine aminotransferase (ALT) (B) or aspartate aminotransferase (AST) levels (C) above or below 30 U/L (upper limit of normal). Points and bars represent least-squares mean and 95% confidence interval values, respectively. p values within each graph represent the between-group differences in the change from baseline. ＊p＜0.05, change from baseline.
Supplementary Fig.1. Changes in the cardio-ankle vascular index from baseline to month 24, using the median values as cutoffs for baseline transaminases.
(A and B) Changes in the cardio-ankle vascular index (CAVI) in the population with CAVI data. Patients were divided into groups according to baseline alanine aminotransferase (ALT) (A) or aspartate aminotransferase (AST) levels (B) above or below the median value. Points and bars represent least-squares mean and 95% confidence interval values, respectively. p values within each graph represent the between-group differences in the change from baseline. ＊p＜0.05, change from baseline.
Blood Pressure
Overall, the mean ( Fig.2A ) , systolic ( Supplemental Fig.2A ) , and diastolic ( Supplemental Fig.3A ) blood pressure and pulse pressure ( Supplemental Fig.4A ) did not change significantly in either the control or febuxostat groups at 12 or 24 months, nor did they differ between the two groups. No significant changes in these parameters were observed in any of the ALT and AST subgroups, and between groups ( Fig.2B and C (cutoffs of 30 U/L for mean blood pressure), Supplemental Fig.5A and C (cutoffs of the median values for mean blood pressure), Supplemental Fig.2B and C (systolic blood pressure), Supplemental Fig.3B and C (diastolic blood pressure), and Supplemental Fig.4B and C (pulse pressure)).
Fig.2. Changes in the mean blood pressure from baseline to month 24 in the population with cardio-ankle vascular index data.
(A) Changes in the mean blood pressure (MBP) in the total population with cardio-ankle vascular index (CAVI) data. (B and C) Patients were divided into groups according to baseline alanine aminotransferase (ALT) (B) or aspartate aminotransferase (AST) levels (C) above or below 30 U/L (upper limit of normal). Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Supplementary Fig.2. Changes in systolic blood pressure from baseline to month 24 in the population with cardio-ankle vascular index data.
(A) Changes in the systolic blood pressure (SBP) in the total population with cardio-ankle vascular index (CAVI) data. (B and C) Patients were divided into groups according to baseline alanine aminotransferase (ALT) (B) or aspartate aminotransferase (AST) levels (C) above or below the median value (left) and 30 U/L (upper limit of normal; right). Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Supplementary Fig.3. Changes in diastolic blood pressure from baseline to month 24 in the population with cardio-ankle vascular index data.
(A) Changes in the diastolic blood pressure (DBP) in the total population with cardio-ankle vascular index (CAVI) data. (B and C) Patients were divided into groups according to baseline alanine aminotransferase (ALT) (B) or aspartate aminotransferase (AST) levels (C) above or below the median value (left) and 30 U/L (upper limit of normal; right). Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Supplementary Fig.4. Changes in pulse pressure from baseline to month 24 in the population with cardio-ankle vascular index data.
(A) Changes in the pulse pressure (PP) in the total population with cardio-ankle vascular index (CAVI) data. (B and C) Patients were divided into groups according to baseline alanine aminotransferase (ALT) (B) or aspartate aminotransferase (AST) levels (C) above or below the median value. Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Supplementary Fig.5. Changes in the mean blood pressure from baseline to month 24 in the population with cardio-ankle vascular index data, using the median values as cutoffs for baseline transaminases.
(A and B) Changes in the mean blood pressure (MBP) in the population with cardio-ankle vascular index (CAVI) data. Patients were divided into groups according to baseline alanine aminotransferase (ALT) (A) or aspartate aminotransferase (AST) levels (B) above or below 30 U/L (upper limit of normal; right). Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Carotid IMT
Overall, the mean CCA-IMT did not change significantly at 12 or 24 months in the control or febuxostat groups, and no differences were observed between the two groups ( Fig.3A ) . In addition, no between-group differences were observed in the mean CCA-IMT in any of the ALT or AST subgroups ( Fig.3B and C (cutoffs of 30 U/L) and Supplemental Fig.6A and C (cutoffs of the median values)).
Fig.3. Changes in the intima-media thickness of the common carotid artery from baseline to month 24 in the population with cardio-ankle vascular index data.
(A) Changes in the intima-media thickness (IMT) of the common carotid artery in the total population with cardio-ankle vascular index (CAVI) data. (B and C) Patients were divided into groups according to baseline alanine aminotransferase (ALT) (B) or aspartate aminotransferase (AST) levels (C) above or below 30 U/L (upper limit of normal). Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Supplementary Fig.6. Changes in the intima-media thickness of the common carotid artery from baseline to month 24 in the population with cardio-ankle vascular index data, using the median values as cutoffs for baseline transaminases.
(A and B) Changes in the intima-media thickness (IMT) of the common carotid artery in the population with cardio-ankle vascular index (CAVI) data. Patients were divided into groups according to baseline alanine aminotransferase (ALT) (A) or aspartate aminotransferase (AST) levels (B) above or below 30 U/L (upper limit of normal; right). Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Serum UA and ALT
In the overall population, the serum UA levels decreased significantly from baseline to 12 and 24 months in the febuxostat group ( Supplemental Fig.7A ) . Significant differences in the serum UA levels were observed between the control and febuxostat groups at 12 and 24 months, with absolute mean differences of 2.24 mg/dL [95% CI, 1.45 to 3.02] and 3.12 mg/dL [95% CI, 2.33 to 3.91], respectively ( Supplemental Fig.7A ) . In the subgroup analysis, serum UA levels were significantly lower in the febuxostat group than in the control group, and significant between-group differences were observed at 12 and 24 months regardless of baseline ALT or AST levels ( Supplemental Fig.7B and C ) . There were also no differences in changes in serum UA between any of the subgroups at the AST and ALT cutoffs.
Supplementary Fig.7. Changes in serum uric acid levels from baseline to month 24 in the population with cardio-ankle vascular index data.
(A) Changes in the serum uric acid (UA) levels in the toatl population with cardio-ankle vascular index (CAVI) data. (B and C) Patients were divided into groups according to baseline alanine aminotransferase (ALT) (B) or aspartate aminotransferase (AST) levels (C) above or below the median value (left) and 30 U/L (upper limit of normal; right). Points and bars represent least-squares mean and 95% confidence interval values, respectively. p values within each graph represent the between-group differences in the change from baseline. ＊p＜0.001 for changes from baseline.
Supplemental Fig.8 shows the time course of changes in the serum ALT levels. No significant changes were observed in the ALT levels in either the control or febuxostat groups over the study period, both in the overall analysis ( Supplemental Fig.8A ) and in the subgroups based on baseline ALT levels ( Supplemental Fig.8B ) . In addition, no inter-group differences were observed ( Supplemental Fig.8A and B ) .
Supplementary Fig.8. Changes in serum ALT levels from baseline to month 24 in the CAVI population.
(A) Changes in the alanine aminotransferase (ALT) levels in the total population with cardio-ankle vascular index (CAVI) data. (B) Patients were divided into groups according to baseline ALT levels above or below the median value (left) and 30 U/L (upper limit of normal, right). Points and bars represent least-squares mean and 95% confidence interval values, respectively.
Discussion
This sub-analysis of the PRIZE study sheds light on the potential benefits of febuxostat, a non-purine-selective XOR inhibitor, on arterial stiffness in patients with hyperuricemia, particularly those with liver dysfunction.
Previous studies have highlighted the role of XOR in generating ROS and contributing to the endothelial dysfunction associated with hyperuricemia 6 , 7) . Although several experimental studies have suggested a potential link between XOR and the progression of atherosclerosis 21 , 22) , the effectiveness of UA-lowering therapy using XOR inhibitors in improving cardiovascular outcomes remains a topic of debate. Notably, a recent prospective, randomized, open-label, blinded study enrolling 5721 subjects with ischemic heart disease found no difference in the rates of the primary outcomes of non-fatal myocardial infarction, non-fatal stroke, or cardiovascular death between patients receiving allopurinol and those receiving usual care 23) . In addition, the main results of the PRIZE study showed that two-year treatment with febuxostat did not delay the progression of carotid IMT 15) . However, previous clinical trials that enrolled patients with hyperuricemia often did not account for confounding factors such as obesity, hypertension, insulin resistance, and dyslipidemia. These factors may have undermined the potential effectiveness of XOR inhibitors in the cardiovascular system.
Plasma XOR activity is reportedly associated with hypertension 24) , vascular endothelial dysfunction 25) , and arterial stiffness 26) independent of UA levels. Importantly, through a series of studies in both humans and mice, we recently demonstrated that, in liver disease conditions, increased plasma XOR activity was directly induced by excessive leakage of hepatic XOR, together with increases in the serum levels of liver enzymes such as ALT and AST 10 , 11) . Moreover, liver-derived XOR induced ROS production by catabolizing hypoxanthine released from vascular endothelial cells, thereby promoting smooth muscle cell proliferation, which could contribute to the augmented neointimal proliferation in NAFLD/MASLD model mice with high plasma XOR activity 11) . On the basis of these findings, we hypothesized that patients with liver dysfunction would benefit the most from XOR inhibitors to prevent atherosclerotic diseases.
In a post-hoc analysis of the PRIZE study, we had previously reported that febuxostat treatment improved arterial stiffness in patients with asymptomatic hyperuricemia, when the results of CAVI and brachial-ankle pulse wave velocity (baPWV) were combined 27) . Furthermore, by subgrouping the study participants on the basis of liver function, the present sub-analysis revealed significant differences in the changes in CAVI between the control and febuxostat groups with higher baseline ALT or AST levels, without between-group differences in blood pressure. Interestingly, this result was derived from a significant increase in CAVI at 24 months in the control group, particularly in patients with ALT/AST levels ≥ 30 U/L, whereas CAVI remained unchanged in the febuxostat group. Accumulating clinical data indicate that NAFLD/MASLD increases the risk of CVD independent of established cardiovascular risk factors 28 - 30) . Moreover, a mild elevation of ALT levels ≥ 30 U/L, which was used as the cutoff value in this study, has been reported to be associated with the incidence of lifestyle-related chronic liver diseases such as NAFLD/MASLD, as well as with hepatic fibrosis, in Japanese patients 31 , 32) . Taken together with the eligibility criteria showing that participants in the PRIZE study were already at risk for atherosclerosis at the time of enrollment, as indicated by the presence of a carotid artery plaque (CCA-IMT ≥ 1.1 mm), the significant increase in CAVI observed in patients with ALT ≥ 30 U/L in the control group may reflect the susceptibility to arteriosclerosis associated with liver disease conditions. Thus, UA-lowering therapy with febuxostat may attenuate the pathogenic link between NAFLD/MASLD and CVDs. Because the changes in serum ALT levels did not differ between the two groups ( Supplemental Fig.5 ) , the effect of febuxostat on liver function did not seem to affect arterial stiffness. In contrast, unlike CAVI, we did not observe any differences in changes in carotid IMT between the control and febuxostat groups, regardless of the baseline ALT or AST levels. Arterial stiffness reflects the structural and functional changes in the diffuse inner layer of the vessel wall that can lead to atherogenesis 33) . Since we have previously shown that liver-derived XOR could promote proliferation and dedifferentiation of vascular smooth muscle cells 11) , it is possible that suppression of plasma XOR by febuxostat had a beneficial effect on CAVI. On the other hand, it may take longer for changes in the CAVI to affect carotid atherosclerosis progression. Further long-term studies are warranted to clarify the effects of febuxostat on atherosclerotic cardiovascular outcomes.
The results of the present study indicate that febuxostat has a favorable effect on arterial stiffness in patients with hyperuricemia and liver dysfunction. These findings are consistent with those of a previous post-hoc analysis of the Beneficial Effect by Xanthine Oxidase Inhibitor on Endothelial Function Beyond Uric Acid (BEYOND-UA) study 34) . In that study, 24-week treatment with the selective XOR inhibitor topiroxostat significantly improved CAVI and baPWV in patients with higher baseline ALT levels 12) . Furthermore, ALT-high participants in the BEYOND-UA study exhibited increased plasma XOR activity, which was markedly suppressed after topiroxostat treatment. Additionally, in the febuxostat group of the present study, there were no significant associations between changes in serum UA levels and changes in CAVI, either in the overall participant or in any of the subgroups of ALT and AST cutoffs (data not shown). Thus, the improvement in arterial stiffness parameters with these XOR inhibitors was not solely due to reductions in serum UA levels but also attributable to reductions in plasma XOR activity. Given the lack of a placebo control group in the BEYOUND-UA study, the strength of the present study is that the beneficial effect of febuxostat on CAVI was demonstrated in comparison with non-pharmacological therapy. Collectively, these findings imply that both XOR inhibitors, febuxostat and topiroxostat, have similar positive effects on arterial stiffness in patients with hyperuricemia and liver dysfunction. Additional basic research is required to clarify the mechanisms underlying these differential effects of XOR inhibitors on hepatic function, such as the reduction of UA, inhibition of plasma XOR activity, or other factors.
This study had several limitations that require consideration. This study was post-hoc and exploratory in nature, and the sample size of participants with CAVI data was small due to the optional nature of the CAVI measurement in the PRIZE study. Therefore, the results should be interpreted with caution. However, this study did not intentionally select sites, and we believe that there is no systematic bias in the selection of subjects for CAVI measurement. While we acknowledge the possibility of a statistical type I error due to the small sample size, the results of this study should provide important preliminary evidence that warrants further investigation. Although the data used in this study were generated from a randomized controlled clinical trial, a sub-analysis focusing on liver function was not performed. Finally, the plasma XOR activity was not measured during the study period. Thus, the findings cannot clarify whether the between-group differences in changes in CAVI observed in individuals with higher ALT/AST levels are related to the suppression of plasma XOR by febuxostat.
Conclusions
Two-year treatment with febuxostat showed a beneficial effect on CAVI in patients with hyperuricemia and increased ALT or AST levels. Although a long-term study of cardiovascular outcomes is warranted, this result indicates the clinical usefulness of XOR inhibitors for cardiovascular protection, especially in patients with NAFLD/MASLD.
Conflicts of Interest
A.T. has received honoraria from Boehringer Ingelheim Japan and Mochida; research funding from GlaxoSmithKline, Takeda, Bristol Myers Squibb, and Novo Nordisk. A.T. is an Editorial Board member of the journal, and he was not involved in handling this manuscript during the submission and review processes. H.Y. received lecture fees from Kyowa Kirin and outsourcing fees from the Organization for Clinical Medicine Promotion. IS has received lecture fees from Ono Pharmaceutical Co., Kowa Company, Ltd., Sumitomo Pharma Co., Eli Lilly Japan K.K, Novo Nordisk Pharma; research funds from Japan Agency for Medical Research and Development (AMED), Cancerscan Inc., Kubarahonke Co. Ltd., Kowa Company, Ltd., Kobayashi Pharmaceutical Co. Ltd., Rohto Pharmaceutical Co. Ltd.; and scholarship donations from Kowa Company, Ltd., Daiichi Sankyo Co., Sumitomo Pharma Co., Takeda Pharma K.K., Mitsubishi Tanabe Pharma Co., Teijin Pharma, Novo Nordisk Pharma, Mochida Pharmaceutical Co., Suzuken Memorial Foundation, Manpei Suzuki Diabetes Foundation, Midori Health Care Center, McSYL, Hakuhokai Central Hospital. N.K. has received honoraria from AstraZeneca, Bayer Yakuhin, Boehringer Ingelheim Japan, Daiichi Sankyo, Eli Lilly Japan, Kowa, Mitsubishi Tanabe Pharma, Mochida Pharmaceutical, MSD, Novartis Pharma, Novo Nordisk Pharma, Ono Pharmaceutical, Otsuka, Tsumura & Co; Research grant from Astellas, Bayer Yakuhin, Boehringer Ingelheim Japan, Fujiyakuhin, Mitsubishi Tanabe Pharma, Mochida Pharmaceutical, Novartis Pharma; Scholarship from Abbott, Boehringer Ingelheim Japan, Daiichi Sankyo, Mitsubishi Tanabe Pharma, Teijin Pharma. ADD FOR ALL OTHER AUTHORS, or state that “All other authors declare no conflict of interest.”.
Funding
The PRIZE study was funded by Teijin Pharma Limited (to K.N.). The funding agencies had no role in the study design, data collection, analysis, decision to publish, or manuscript preparation.
Author Contributions
All authors contributed to the study design and conception. K.Y., Y.F., and H.N. wrote the manuscript. A.T. was involved in data analyses and interpretation. H.Y. performed statistical analyses. S.N., M.S., I.S., and K.N. reviewed the manuscript. All the authors have read and approved the final version of the manuscript.
Acknowledgements
The authors are grateful to all participants and staff for their contributions to the PRIZE study. This work was supported in part by the Suzuken Memorial Foundation (to I.S.), a research grant from the Gout and Uric Acid Foundation of Japan (to H.N.), a Grant-in-Aid for Scientific Research (C) no. 23K08006 (to H.N.), and the Taiju Life Social Welfare Foundation (to A.T.).
Data Availability Statement
The data will be made available on reasonable request from researchers. Inquiries are to be addressed to the corresponding authors or the secretariat of the PRIZE study (contacted via prizesub-secre@clin-med.org).
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Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The data will be made available on reasonable request from researchers. Inquiries are to be addressed to the corresponding authors or the secretariat of the PRIZE study (contacted via prizesub-secre@clin-med.org).