Therapeutic value of serum uric acid levels increasing in the treatment of multiple sclerosis

Background/Aim. Uric acid was successfully used in both, prevention and treatment of the animal model of multiple sclerosis (MS). Recently it has been shown that inosine, a ribosylated precursor of uric acid, might be used to elevate serum uric acid levels in MS patients. The aim of this study was to evaluate the safety and efficacy of oral inosine as a single drug treatment in patients with MS. Methods. We administered inosine orally to 32 MS patients from 2001– 2004 year at doses from 1–2 g daily (given twice) depending on the pretreatment serum uric acid levels. The mean follow-up interval was 37.69±6.55 months. The other 32 MS patients, without any treatment except for a relapse period (matched by age, sex, duration of disease and functional disability), were used as controls. The follow-up interval of these patients was 36.39±2.68 months. The neurological disability was evaluated by the Expanded Disability Status Scale score (EDSS). Results. During the observed period the treated MS patients were found to have the lower relapses rate than the non-treated MS patients (Chi-square test, p = 0.001). None of the patients have showed any adverse effect of inosine treatment. The non-treated MS patients were found to have a higher increasing in the mean EDSS score than the treated ones (two-way ANOVArepeated measures/factor times, p = 0.025). Conclusion. Our results suggested that the treatment approaches based on the elevation of serum uric acid levels might prove beneficial for some MS patients.


Introduction
Multiple sclerosis (MS) is a disease with heterogeneity in clinical course, neuroradiological appearance of the lesions, response to therapy, and pathological patterns of demyelination.All of these have already raised the question whether MS is a disease or a neurological syndrome with different immunopathological mechanisms initiating a final pathological process.
There is an increasing evidence that reactive oxygen and reactive nitrogen species play an important role in MS pathogenesis.These species are involved in inflammation, Tončev G. Vojnosanit Pregl 2006; 63 (10): 879-882.blood brain barrier disruption, oligodendrocytes damage, demyelination, and axonal damage.Within the past 10 years, peroxynitrite is has been established as one of the major damaging oxidants produced in humans 1 .Nitrotyrosine, a marker of peroxynitrite forming, has been found to be significantly raised in the serum of multiple sclerosis patients 2 .
On the other hand, uric acid is a natural occurring product of purine metabolism.Recently, uric acid has been shown as a strong natural scavenger of peroxynitrite 3 and/or peroxynitrite-derived reactive species 4 .Uric acid has been successfully used in both prevention and treatment of experimental allergic encephalomyelitis (EAE), an animal model of MS 5 .Significantly lower serum uric acid levels are found in MS patients [5][6][7] .There are studies demonstrating that serum uric acid levels inversely correlate with the disease activity 6, 7 and blood brain barrier dysfunction 7 .The findings that the incidence of MS is significantly reduced in patients with gout (chronic hyperuricemia), and that gout and MS might be virtually self-exclusive 5 , have revealed the possibility that the higher serum uric acid levels may protect against the development of MS.
Due to its destruction in the gut, oral administration of uric acid does not increase humans serum levels.However, inosine is a ribosylated precursor of uric acid that has been used extensively in humans for a number of years as an energy supplement and performance enhancer.Oral administration of inosine leads to an easily controlled hyperuricemia without associated side-effects.
The aim of this study was to evaluate the safety and efficacy of oral inosine as a single drug treatment in patients with MS.

Methods
Sixty four MS patients with the confirmed diagnosis of relapsing MS, according to the McDonalds criteria 8 , were included in this study.The patients were diagnosed in the Center for Neurology, Clinical Center Kragujevac.The patients have never been sabmitted to immunomodulatory or immunosuppressive treatment and had the Expanded Disability Status Scale score (EDSS) ≤ 5 9 .
We administered inosine orally as a single drug to 32 MS patients at doses from 1-2 g daily (given twice) depending on the pretreatment serum uric acid levels in order to elevate these levels over the mean normal values: more than 315 µmol/l for men (the normal range is 210 to 420 µmol/l) and more than 250 µmol/l for women (the normal range is 150 to 350 µmol/l) (1 µmol/l = 0.0166 mg/dl).Serum uric acid levels were measured every two moths by using a quantitative enzymatic assay (Elitech diagnostics, Sees, France) according to the manufacturer's protocol, and the results were standardized using a commercial uric acid standard solution.The doses of inosine were corrected every two months according to the serum uric acid levels.The other 32 MS patients were used as controls.The follow-up interval for these patients was 36.39±2.68months.
The EDSS score served as a measure for the disease worsening and was measured before and after the follow-up interval.In case of relapse, the patients received IV methylprednisolone (1g daily for 5 days).
All MS patients and controls agreed to be involved in the study.The local Ethical Committee approved this.
Statistical analysis was performed by using the SYSTAT program.Proportions were compared by using Chi-square tests with a continuity correction or Fisher's exact test when appropriate.Two-way ANOVA (repeated measures-factor times) was used to compare changes in EDSS score during the follow-up interval.The mean serum uric acid levels were compared by using t test.
None of the patients showed any adverse effect of the inosine treatment.During the period observed, the treated MS patients were found to have significantly lower relapses rate than the non-treated MS patients (Chi-square test, p = 0.001) (Figure 1).We noticed 16 relapses in 12 treated MS patients vs. 56 relapses in 31 non-treated MS patients.In the treated group, 9 patients had the experience of one relapse, 2 of two relapses and 1 of three relapses.In the non-  The non-treated MS patients were found to have a significantly higher increasing in progression of disability measured by the mean EDSS score (from 1.84±1.05 to 3.28±1.01)than the treated MS patients (from 2.62±1.48 to 3.03±1.66)(two-way ANOVA-repeated measures-factor times, p = 0.025) (Figure 2).
Before the treatment starting there were no differences in serum uric acid levels between the treated and non-treated MS patients (210.63±39.23 vs. 208.96±23.96).After the follow-up interval the treated MS patients without relapses were found to have significantly higher serum uric acid levels than the treated MS patients with the experience of one or more relapses (t-test, p = 0.005) and than the non-treated MS patients (t-test, p = 0.001) (Table 3).The treated MS patients with relapses were also found to have significantly higher serum uric acid levels than the non-treated patients after the follow-up period (t test, p = 0.028) (Table 3).

Discussion
This study was an attempt to evaluate the safety and efficacy of inosine as a single drug in MS patients.The treated MS patients were found to have a significantly lower relapses rate and a significantly lower progression of disability than the non-treated MS patients.Unfortunately, despite the treatment, one group of MS patients had the relapses and progression of functional disability measured by the EDSS score.
To our knowledge, this was the second attempt to evaluate the safety and efficacy of oral inosine, but the first with inosine as a single drug treatment in patients with MS.The first treatment attempt in human MS with uric acid has been recently reported 10,11 .Inosine has been administered orally to MS patients starting at doses of 1 g daily and increasing to 3 g daily (given twice).After nearly a year of the oral administration of either 2 or 3 g of inosine, 3 of 11 patients showed some evidence of clinical improvement with-  out any sign of the disease progression in the remaining patients during the observed period.Moreover, a significant reduction in lesion activity was seen in one of the two patients with active lesions detected by magnetic resonance imaging at the beginning of the trial.In our study, we did not find any the patients with the evidence of clinical improvement but we found 20 patients (62.5% of the treated patients) with no relapse and sign of the disease progression.Unfortunately, we were not able to measure the disease activity by magnetic resonance imaging during the study (before and after the treatment).
There has been a lot of evidence that uric acid is involved in MS pathology.The incidence of MS is substantially reduced in patients with gout, a disease most often resulting from high serum uric acid levels 5 .There has been a similar finding that MS patients have lower serum uric acid despite the fact that both the MS and control groups were heterogeneous in up to the already published studies: MS patients with the different clinical types and activity of disease, patients with other neurological diseases, patients with other inflammatory neurological diseases, healthy subjects 5,7,8,12 .Consistently, in the pairs of mono-and dizygotic twins where only one had MS, serum uric acid levels were lower in the individuals with MS than their healthy siblings 13 .Serum uric acid levels were also lower in patients with optic neuritis 14 .
On the other hand, increased serum uric acid levels are demonstrated after the glatiramer acetate therapy 15 , and after methylprednisolone therapy 16 implying that a beneficial therapeutic effect of these drugs might be due to the elevation of uric acid.In the light of these findings, uric acid is proposed as one of the possible bio-markers of disease activity and a response to the therapy in multiple sclerosis 17 .
Uric acid was successfully used in both prevention and treatment of EAE, the animal model of MS 5 but the lower mammalians have urate oxidase, the enzyme that catalyses the oxidation of uric acid to allantoin.Allantoin has no antioxidant properties.However, humans have a different purine degradation pathway from the lower mammalians.In humans, uric acid is a final product of purine metabolism because enzyme urate oxidase is lost in the evolution.Consequently, in humans, there are higher serum uric acid levels 18 .It is believed that the loss of urate oxidase can have strong evaluative superiority in humans.

Conclusion
The results of the current study suggest that the treatment approaches based on the elevation of serum uric acid levels by using a ribosylated precursor of uric acid may have a benefit for some MS patients but it has to be confirmed in double-blind, placebo-controlled studies.

Table 1 Baseline demographic and disease characteristics
treated group, 12 patients had the experience of one relapse, 13 of two relapses and 6 of three relapses (Table2).