Uric Acid Effects on Endothelium and Oxydative Stress

Differential Effects of Uric Acid and Xanthine Oxidoreductase on Endothelial Function and Oxydative Stress

Cardiovascular disease is the leading cause of mortality worldwide. Endothelial dysfunction (ED) is the main mechanism which leads to atherosclerosis, where the balance between pro and antioxidant factors results in a decreased nitric oxide (NO) bioavailability. Xanthine OxidoReductase (XOR) is one of the main generators of reactive oxygen species (ROS). Uric acid (UA), a major antioxidant in human plasma and end product of purine metabolism, is associated with cardiovascular diseases since many years; however the precise mechanisms which relate UA to ED are still not well understood.

The purpose of this study is to unravel the XOR and UA pathways involved in ED. Three groups of participants (young (< 40 y) male healthy participants [1] ; male and female helthy participants (40 to 65 y) [2] and patients with primary hypertension [3]) will be exposed to febuxostat (a strong and selective XOR inhibitor), or recombinant uricase (which oxidizes UA into allantoin) to vary UA levels and concomitantly control for confounding changes in XOR activity. Oxidative stress will be estimated by several markers. Endothelial function will be assessed by a laser Doppler imager in the presence of hyperthermia and endothelium stimulators. This study is specifically designed to untie the respective effects of UA and XOR pathways on oxidative stress and endothelial function in humans.

The investigators will test the following hypothesis:

1. An extremely low level of uric acid after uricase administration induces endothelial dysfunction and oxydative stress,
2. A specific XO inhibitor limits unfavourable effects of the serum UA reduction elicited by uricase administration,
3. Endothelial function and oxydative stress are further improved with febuxostat as compared to placebo,
4. All these observations are more marked in hypertensives then in older participants than in young healthy subjects.

Study Overview

• Status: Completed
• Conditions: Hypertension; Oxidative Stress; Cardiovascular System; Endothelial Function
• Intervention / Treatment: Drug: Placebos; Drug: Febuxostat; Drug: Rasburicase

Detailed Description

The goals of the research protocol are to clearly untie the respective roles of uric acid (UA) and xanthine oxidoreductase (XOR) pathways on endothelial function and oxidative stress in humans.

UA represents the end-product of purine metabolism due to the loss of uricase 15 million years ago in humans. The selective advantage of this mutation could be the strong antioxidant effect of UA (which represents more than 60% of the antioxidant plasmatic capacity). Many recent epidemiological studies have showed a J-shape association between UA levels and cardiovascular risk. An UA level lower than 3 mg/dl could be damageable due to the loss of the antioxidant properties of UA. In contrast, hyperuricemia is associated with an increased inflammation, insulin resistance, ED, platelet aggregation, left ventricle hypertrophy, arterial vasodilatation impairment, aortic stiffness and intima-media thickness. However, the association between UA and cardiovascular disease remains controversial because whether UA is an independent risk factor for these illnesses is unclear.

Interventional studies:

Because the above-mentioned associations do not prove causation, several authors designed interventional studies with the purpose to modify UA levels and determine if this affected endothelial function and oxidative stress. The main limitation of these studies is that they were unable to untie the effects of the synthesis of UA, of UA itself and of the activity of XOR, on ROS production and endothelial function in humans.

This is because:

1. serum UA is a powerful plasmatic antioxidant,
2. but transformation of hypoxanthine to xanthine and xanthine to UA by XO generates intracellular ROS,
3. moreover, in endothelial cells, UA reduces NO bioavailability by many ways (L-arginine blockade and degradation, increased superoxide anion production, NOS inhibition, reduced NOS genes expression and direct NO scavenging),
4. in addition, UA forms crystals in the endothelium wall which create a pro-inflammatory and thrombotic state, increases smooth muscle cells proliferation and also insulin resistance and inflammation in adipocytes,
5. finally, and most importantly, XO is inhibited by physiologic levels of UA (which acts as an uncompetitive XO inhibitor).

In summary, the present protocol aims at testing the following hypothesis:

1. Experimental serum UA variations are correlated with endothelial function and oxydative stress markers : an extremely low level after uricase administration induces ED and oxydative stress,
2. A specific XO inhibitor (FX) limits unfavourable effects of the serum UA reduction elicited by uricase administration, since this will hamper the feedback activation of XO by a low UA level,
3. Endothelial function and oxydative stress are further improved with FX as compared to placebo, because the first experimental condition results in a XO blockade,
4. All these observations are more marked in hypertensive patients then in older participants than in young healthy subjects.

Data collection

Data collection from the participants will be collected informatically through a case report form. The names and personnal data from the patients will be kept in a secret place or in a password-protected file. All the data will be destroyed at the end of the study (including blood and urine samples).

Statistical analysis

Statistical analysis will be performed using SPSS. Baseline characteristics will be compared using a Student t test. Two-way repeated-measures ANOVAs will be used to detect significant changes between sessions and groups. Statistical significance is assumed when p is <0.05. Sample size is not possible due to the lack of data of the effect of acute hypouricemia. We estimate a minimum of 15 participants in each group.

Specific test will be used for non gaussian variables. Correlation test will be used if necessary according the results of the first test.

Placebo-corrected values and comparisons of the delta will be used too.

Subgroups analyses will be performed for the study of population 2 and 3 (enrolled together).

• Study Type: Interventional
• Enrollment (Actual): 53
• Phase: Not Applicable

Contacts and Locations

Study Locations

• Belgium
  • Belgique
    • Brussels, Belgique, Belgium, 1070
      • Erasme Hospital

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 18 years to 65 years (Adult, Older Adult)
• Accepts Healthy Volunteers: Yes
• Genders Eligible for Study: All

Description

Phase 1 :

Inclusion Criteria:

• Age between 18 and 40 years
• Male
• Healthy volunteers
• Non smoker for at least 6 months
• Uric acid level in normal range (normouricemic group)

Exclusion Criteria:

• Any diseases of one of the following systems: cardiovascular, digestive, hormonal, urinary, pulmonary, rheumatic or immune.
• Smoker, alcoholic
• Participants should not take any chronic medicine nor vitamins or other antioxidants.
• A G6PD deficit will be excluded as this is a contraindication to uricase administration (hemolytic anemia).

Phase 2 :

Inclusion Criteria:

• Age between 40 and 65 years
• Male or female (menopaused)
• Healthy volunteers
• Non smoker for at least 6 months
• Uric acid level in normal range (normouricemic group)

Exclusion Criteria:

• Any diseases of one of the following systems: cardiovascular, digestive, hormonal, urinary, pulmonary, rheumatic or immune.
• Smoker, alcoholic
• Participants should not take any chronic medicine nor vitamins or other antioxidants.
• A G6PD deficit will be excluded as this is a contraindication to uricase administration (hemolytic anemia).

Phase 3 :

Inclusion Criteria:

• Age between 40 and 65 years
• History of hypertension for more than 6 months
• Non smoker or smoke stopped for at least for 6 months

Exclusion Criteria:

• Acute coronary syndrome
• Heart failure (LVEJ < 40%)
• Diabetes
• Active smoking
• Gout
• Chronic kidney disease stage superior to 3a
• History of cerebrovascular thrombosis
• Cirrhosis
• Alcohol consumption more than 3 units/day
• Participants should not take any chronic medicine nor vitamins or other antioxidants.
• A G6PD deficit will be excluded as this is a contraindication to uricase administration (hemolytic anemia).

The populations of phases 2 and 3 will be enrolled and studied together with subgroups analyses of the results for the status of hypertension, of treatment, age and gender.

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Basic Science
• Allocation: Randomized
• Interventional Model: Crossover Assignment
• Masking: Double

Number of Arms

4

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Placebo Comparator: Placebos PO and IV; PO : per os IV : intraveinously	Drug: Placebos; Lactose placebo for pills and saline for perfusion.; Other Names: Placebo
Experimental: Febuxostat PO and Placebo IV; 240 mg a day for 3 days	Drug: Placebos; Lactose placebo for pills and saline for perfusion.; Other Names: Placebo; Drug: Febuxostat; Febuxostat tablet.; Other Names: Adenuric
Experimental: Febuxostat PO And Rasburicase IV; Febuxostat : 240 mg a day for 3 days. Uricase : 3 mg once.	Drug: Febuxostat; Febuxostat tablet.; Other Names: Adenuric; Drug: Rasburicase; Rasburicase injectable solution.; Other Names: Fasturtec
Experimental: Placebo PO And Rasburicase IV; Placebo : for 3 days. Uricase : 3 mg once.	Drug: Placebos; Lactose placebo for pills and saline for perfusion.; Other Names: Placebo; Drug: Rasburicase; Rasburicase injectable solution.; Other Names: Fasturtec

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Cutaneous perfusion by Laser Doppler (perfusion unit)	Perfusion unit (Laser Doppler Imager + iontophoresis of (ACh and SNP and hyperemia with ou without L-NAME). Assessment of endothelial function.	24 hours after infusion of Uricase or Placebo

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Change in Oxydative stress biomarkers from baseline to 30 min or 24 hours after infusion of Uricase or Placebo		Baseline, 30 minutes and 24 hours after infusion of Uricase or Placebo
Arterial stiffness	Carotido-femoral and carotido-radial pulse wave velocity and pulse wave analysis	24 hours after infusion of Uricase or Placebo
Blood pressure (mmHg)	Beat-to-beat measurements with Finapres and manually	24 hours after infusion of Uricase or Placebo
Cardiac output (l-min)	Beat-to-beat measurements with Finapres	24 hours after infusion of Uricase or Placebo
Change in enzymes activity	Xanthin oxydase activity	30 min and 24 hours after infusion of Uricase or Placebo
Change in enzymes expression	eNOS, NOX, XO we will incubate endothelial cells with plasma or serum from subjects. Then we will measure the sus-mentionned enzymes' expression.	30 min and 24 hours after infusion of Uricase or Placebo
Change in proteomic or metabolomic analysis	We will perform proteomics or metabolomics analysis directly on serum from participants and also on lysate of endothelial cells pre-incubated with plasma or serum from participants.	30 min and 24 hours after infusion of Uricase or Placebo
Change in renin-angiotensin activity		Baseline, 30 minutes and 24 hours after infusion of Uricase or Placebo
Change in urinary excretion of sodium		Baseline, 30 minutes and 24 hours after infusion of Uricase or Placebo

Collaborators and Investigators

• Sponsor: Erasme University Hospital
• Collaborators: Fonds National de la Recherche Scientifique; Fonds Erasme
• Investigators: Study Director: Philippe van de Borne, Erasme Hospital

Publications and helpful links

General Publications

• De Becker B, Hupkens E, Dewachter L, Coremans C, Delporte C, van Antwerpen P, Franck T, Zouaoui Boudjeltia K, Cullus P, van de Borne P. Acute effects of hypouricemia on endothelium, oxidative stress, and arterial stiffness: A randomized, double-blind, crossover study. Physiol Rep. 2021 Sep;9(17):e15018. doi: 10.14814/phy2.15018.
• De Becker B, Coremans C, Chaumont M, Delporte C, Van Antwerpen P, Franck T, Rousseau A, Zouaoui Boudjeltia K, Cullus P, van de Borne P. Severe Hypouricemia Impairs Endothelium-Dependent Vasodilatation and Reduces Blood Pressure in Healthy Young Men: A Randomized, Placebo-Controlled, and Crossover Study. J Am Heart Assoc. 2019 Dec 3;8(23):e013130. doi: 10.1161/JAHA.119.013130. Epub 2019 Nov 22.

Study record dates

Study Major Dates

• Study Start (Actual): January 3, 2018
• Primary Completion (Actual): December 31, 2019
• Study Completion (Actual): February 27, 2020

Study Registration Dates

• First Submitted: December 29, 2017
• First Submitted That Met QC Criteria: January 4, 2018
• First Posted (Actual): January 10, 2018

Study Record Updates

• Last Update Posted (Actual): February 28, 2020
• Last Update Submitted That Met QC Criteria: February 27, 2020
• Last Verified: February 1, 2020

More Information

Terms related to this study

• Keywords: uric acid; febuxostat; xanthin oxydoreductase; uricase
• Additional Relevant MeSH Terms: Cardiovascular Diseases; Vascular Diseases; Hypertension; Antirheumatic Agents; Gout Suppressants; Rasburicase; Febuxostat
• Other Study ID Numbers: 30309647

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: NO

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No