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REDUCED INTIWVENOUS GLUTATHIONE IN THE TREATMENT OF
EARLY PAREINSON’S DISEASE
GIANPIETRO SECHI’, MARIA G. DELEDDA’, GUIDO BUA’, WANDA M.SAl-TA’,
GIOVANNI A. DEIANA’, GIOVANNI M. PES3 and GIULIO ROSATI’
‘Department of Neurology, *Division of Internal Medicine, and 3Chair of Clinical
Biochemistry, University of Sassari, Sassari, Italy.
(Final form, July 1996)
Abstract
Sechi GianPietro, Maria G. Deledda, Guido Bua, Wanda M. Satta, Giovanni A.
Deiana, Giovanni M. Pes, and Giulio Rosati: Reduced intravenous glutathione in
the treatment of early Parkinson’s Disease. Prog. Neuro-Psychopharmacol &
Biol. Psychiat. 1996. 20, pp. 1159-1170
1. Several studies have demonstrated a deficiency in reduced glutathione
(GSH) in the nigra of patients with Parkinson’s Disease (PD). In particular,
the magnitude of reduction in GSH seems to parallel the severity of the
disease. This finding may indicate a means by which the nigra cells could
be therapeutically supported.
2. The authors studied the effects of GSH in nine patients with early,
untreated PD. GSH was administered intravenous, 600 mg twice daily, for
30 days, in an open label fashion. Then, the drug was discontinued and a
follow-up examination carried-out at l-month interval for 2-4 months.
Thereafter, the patients were treated with carbidopa-levodopa.
3. The clinical disability was assessed by using two different rating scale
and the Webster Step-Second Test at baseline and at l-month interval for
4-6 months. All patients improved significantly after GSH therapy, with a
42% decline in disability. Once GSH was stopped the therapeutic effect
lasted for 2-4 months.
4. Our data indicate that in untreated PD patients GSH has symptomatic
efficacy and possibly retards the progression of the disease.
Kevwords: Parkinson’s Disease; reduced glutathione.
Abbreviations: Columbia University Rating Scale (CURS): Parkinson’s Disease
(PD); Patients Global Impressions (PGI); reduced glutathione (GSH); resting
tremor (RT); Webster Step-Second Test (W.S.S.T.).
1159
1160 G.P. Sechi et aL
Introduction
The mechanisms underlying dopamine ceils death in the zona compacta of
substantia nigra in Parkinson’s disease (PD) remain unclear. However, current
concepts of this process indicate that free radicals generated by oxidation
reactions may play a key role (Jenner et al., 1992). Indeed, in postmortem
tissues from patients with PD there is evidence for inhibition of complex 1 of
the mitochondrial respiratory chain, altered iron metabolism and decreased
levels of reduced glutathione (GSH) (Riederer et al., 1989, Jenner, 1993). Of
these defence mechanisms implicated in the prevention of free-radical-
induced tissue damage, only the reduction in the levels of GSH in substantia
nigra appears to be specific to PD (Jenner, 1993, Sian et al., 1994) and,
noteworthy, this reduction has been also found in cases of incidental Lewy
body disease (presymptomatic PD) (Perry et al., 1982, Sian et al., 1992). In
particular, the magnitude of reduction in GSH seems to parallel the severity of
PD and, in advanced stages, in the nigra, GSH is virtually undetectable
(Riederer et al., 1989). In addition, data from animal studies have shown that
an induced GSH depletion in mice produces morphological changes in nigral
dopamine neurons resembling those seen in normal aging and in MPTP (l-
methyl-4-phenyl-1,2,3,6_tetrahydropyridine) neurotoxicity (McNeil et al.,
1986). These observations have led us to determine the effect of intravenous
(i.v.) GSH in patients with early, untreated PD.
Methods
Patients
After giving informed consent, 9 consecutive patients with idiopathic PD
were enrolled in the study. There were 6 men and 3 women, age 66 k 9 years
(mean + SD) (range, 49 to 77); Hoehn and Yahr stage of parkinsonism 2 f 0.9
(mean + SD) (range 1 to 4), with a disability duration of 13 + 4.5 months (mean
f SD) (range, 8 to 24). The patients were considered eligible for the study,
provided that they had not been treated previously with any antiparkinsonian
drug or other agents active on the central nervous system including deprenyl or
vitamin E. The patients with dementia (Mini-Mental State Examination), or
depression (Hamilton Rating Scale) were excluded.
Procedures
GSH was administered i.v. 600 mg in 250 ml saline, as l-hour infusion,
twice daily, at 8.00 A.M. and 4.00 P.M, for 30 days, in an open label fashion. The
patients were assessed at baseline and 30 days after the treatment. Then, GSH
Intravenous GSH In PD 1161
was discontinued and, if the patient status improved, a follow-up examination
was carried-out at l-month interval, until the patient’s clinical status
returned to baseline, or when the patient felt he or she was worsened.
Thereafter, the patients were treated with carbidopa-levodopa ( 25250 mg ),
half-tablet three times daily, and a new examination carried-out after 30
days, about two hours after the intake of the drug.
Assessments
On each visit, the clinical disability was assessed according to a modified
Columbia University Rating Scale (CURS) (Yahr et al., 1969), and to the Webster
Step-Second Test (W.S.S.T.) (scoring method: time in seconds to stand and walk
a prescribed course and sit again) (Webster, 1968).
The subscores evaluated at the modified CURS were: speech, hypomimia,
tremor at rest, action or postural tremor of hands, rigidity, finger taps, hand
movements, pronation and supination of hands, foot taping, arising from chair,
posture, gait, balance and hypokinesia.
For the W.S.S.T., three sequential trials were performed for each patient, at
baseline and at each control, with a fixed intertrial interval of 15 s. In the
tabulation of the results the authors used the mean (k SD) of the three W.S.S.T.
values obtained, for each patient, at the beginning of the experiment and after
each of the trial periods.
Clinical response was also self assessed by patients according to Patients
Global Impressions (PGI) (Guy, 1976). This scale (ranging from 1 = very much
better, to 7 = very much worse) was used to assess the change in severity of
the disease from the beginning of the study and from the previous visit.
All patients were evaluated by the same examiner throughout the study. On
each examination, they were observed over two consecutive days. In addition,
in patients with tremor, at baseline and at each examination, at approximately
the same time of day, tremograms were recorded using an accelerometer
transducer attached to the index finger of the hand and recorded on an EEG
polygraph. Tremor frequency (in Hertz) and visual mean amplitude (mean value
of tremor estimated visually in microvolts) were measured from the tracings.
Laboratorv Assessments
The following laboratory tests were performed at entry and after 30 days of
GSH therapy: routine blood chemistries, liver function tests, blood counts,
urinalysis and ECG. A chest X-ray and a brain CT, performed without contrast,
were conducted in all patients before study entry.
1162 G.P. Se&l et cd.
r nalvsrs
A statistical analysis of W.S.S.T. values was made, for each patient, by
Student’s t test for paired samples. A nonparametric statistical method was
used to compare clinical parkinsonian scores (Wilcoxon Matched-pairs Signed-
ranks Test) and the PGI (McNemar’s Test). In addition, the sums of clinical
parkinsonian scores for each patient, at baseline, were correlated either with
the percent improvement calculated through the same scale, or with the
percent improvement at W.S.S.T. after GSH therapy. The percentage of change
was calculated by the following formula:
prestudy value - treatment value / prestudy value x 100 = % change. The level
of significance was p c 0.05.
Results
All 9 patients enrolled completed the study. There were no serious
complications from i.v infusion of GSH. Two patients during the third week of
i.v GSH treatment had fever (axillary, peak temperature, 3&2”C), erithemia of
the skin, irritation and hardness at injection site, likely due to infusion
thrombophlebitis. The irritation and fever cleared up after 5 days of antibiotic
and antiphlogistic therapy. Patient 6 after completion of the wash-out period
suffered a thigh-bone fracture. GSH did not induce clinically significant
changes in any laboratory test compared with basal conditions, The brain CTs
showed a mild cortical atrophy in two patients and no definite abnormalities
in seven of them. The frequency of resting tremor (RT) was 5 to 6 Hz. In patient
1, the mean amplitude of RT, compared with the baseline period, was reduced,
approximately, by 50% after GSH therapy, and by 25% after carbidopa-levodopa
(Fig 1). No definite variations in the mean amplitude of RT were noted in the
other patients, in the various sequences of treatment, with respect to
baseline, or to the wash-out period. At W.S.S.T. all patients improved
significantly after GSH, with respect to baseline (from p < 0.05, to p c 0.01);
instead, after levodopa-carbidopa, only five patients improved significantly,
with respect to the wash-out period (from p < 0.02, to p c 0.01). (Table 1). In
our opinion, for the dosages of levodopa-carbidopa and GSH used, the transient
improvement induced by these drugs was roughly comparable. The total scores
for parkinsonian disability (modified CURS) were significantly lower either
after GSH therapy, with respect to baseline (p < 0.007 ), or after levodopa-
carbidopa with respect to the wash-out period (p c 0.01). (Table 2). A
significant improvement after GSH therapy, with respect to baseline, of
modified CURS subscores, was evidenced for speech, hypomimia, rigidity,
lntravenoue GSH in PD 1163
pronation and supination of hands, foot taping, posture, gait, balance and
hypokinesia (from p < 0.02, to p c 0.007).
Fig 1. Representative resting tremor recordings in patient 1, at baseline (A);
30 days after therapy with reduced glutathione (B); 60 days after withdrawal
of reduced giutathione (C); and 30 days after carbidopa-ievodopa (D): Vertical
calibration is 200 microvolts; horizontal scale is 1 second.
A similar improvement was noted after carbidopa-levodopa, with respect to
the wash-out period (from p c 0.04, to p < 0.01). As seen from Tables, the
values of the modified CURS scores and the values of W.S.S.T., after
withdrawal of GSH therapy, reached the baseline values after 2.6 k 0.7 months
(range, 2 to 4 months). The correlation coefficient between total CURS scores
at baseline, and the percent improvement calculated through the same scale,
after GSH therapy, is shown in Fig 2. in Fig 3, is shown the correlation
coefficient between total CURS scores at baseline and the percent
improvement at the W.S.S.T. after GSH therapy. The slope of this late
correlation is significantly different than zero (r = 0.6813; p = 0.0433), while
the correlation shown in Fig 2 is non-significant.
Table 1
Webster Step-Second Test (W.S.S.T): Scoring Method: Time in Seconds to Stand and Walk a Prescribed
Course and Sit Again (mean + SD, of 3 sequential trials).
W.S.S.T. W.S.S.T. % Wash-out W.S.ST. After WSST. After %
Patient At Baseline After GSH Improv.1 (months) V&&out Lev.+DCI Improv.2
46.8 f 1.6 41.2 f 0.3** 1 2
58.7 f 0.6 57.2 z!z 0.6* 3
34.0 f 0.1 33.0 f 0.5* 3
34.3 + 0.5 33.3 f 0.3* 3
59.0 + 1.8 52.0 + 1.8** 1 2
32.9 f 4.3 25.6 f 1.5’ 22
44.3 f 1.15 39.7 + 0.58** 10
51.7 + 0.8 49.4 f l.O* 4.5
25.0 + 0.1 22.7 + l.l* 9
3 45.5 rt 0.86 43.0 f 0.5** 5.5
4 65.7 _+ 1.1 63.7 _+ 1.1 3
3 37.7 f 0.58 34.7 f 0.57” 8
2 42.7 k 0.58 42.0 + 0.5 2
2 51.2 f 0.3 50.0 f O.l*’ 2.5
2 32.3 + 2.6 -
2 45.0 f 1.2 39.6 f 1.9*** 12
2 52.0 f 1.2 48.2 + 1.3”’ 7
3 24.3 f 0.6 24.0 f 0.1 1
Lev.+DCI=Levodopa+Decarboxylase Inhibitor; W.S.S.T. values after GSH were compared with baseline;
W.S.S.T. values after Lev.+DCI were compared with the wash-out period; *p<O.O5; **p<O.Ol; **‘p<O.O2
(Student’s t Test for paired samples).
Table 2
Modified Columbia University Rating Scale: Total Scores in 9 Patients with Parkinson’s Disease Treated
with GSH (600 mg/day, I.V.) or Levodopa (375 mg/day + DCI, per OS).
Baseline T. Scores % Wash-out T.Scores T. Scores %
Patient T. Scores After GSH Improv.1 (months) After W&-cut After Lev.+DCI Impr0v.p
25 15 40 3 24
12 3 75 4 16
29 20 31 3 27
15 7 53 2 18
35 17 51 2 30
42 26 38 2 42
20 15 25 2 21
31 19 39 2 32
34 25 27 3 35
14 42
3 81
20 26
7 61
14 53
14 33
21 34
26 26
Mean + SD 27 +lO 16 f 8* 42 f 16 2.6 f 0.7 27 f 8 15 f 7** 44.5 f 19
Lev.+DCI=Levodopa+Decarboxylase Inhibitor; T. Scores=Total Scores; T. Scores values after GSH were
compared with baseline; T. Scores values after Lev.+DCI were compared with the wash-out period;
*pcO.O07; **p<O.Ol ; (Wilcoxon Matched-pair Signed-ranks Test).
1166 G.P. Se& et aL
Total Scores of CURS at Baseline
Fig 2. Correlation coefficient between total CURS scores at baseline and the
percent improvement calculated through the same scale after therapy with
reduced glutathione (r= - 0.5027; p=n.s.).
25
20
15
10
5
-
. .
/ .
Oi I I L
10 20 30 40 50
Total Score8 of CURS at Baaeltne
I I I 7
.
Fig 3. Correlation coefficient between total CURS scores at baseline and the
percent improvement at the W.S.S.T. after therapy with reduced glutathione
(r=0.6813; p=O.O433).
Intravenous GSH in PD 1167
Total scores of PGI were significantly lower either after GSH therapy, with
respect to wash-out period (or, baseline) (p < 0.0039) or after levodopa-
carbidopa, with respect to the wash-out period (p c 0.0039), where, no
significant difference was found at PGI between GSH therapy and levodopa-
carbidopa. One patient (n.9) with a marked sialorrhea, reported the
disappearance of this symptom after GSH therapy. After withdrawal of GSH the
benefit lasted for about 3 months. Levodopa-carbidopa therapy was ineffective
on sialorrhea in the same patient.
Discusslon
The crucial observation, that In idiopathic PD the magnitude of reduction in
GSH in substantia nigra seems to parellel the severity of the disease, may
indicate a means by which the nigra cells could be therapeutically supported
(Riederer et al., 1989).
GSH and the Blood-Brain Bar&r
GSH is a tripeptide (gamma-glutamyl-cysteinyl-glycine) which in
physiological conditions is belleved to be extracted in minimal amount at the
blood-brain barrier (Cornford et al., 1978). In addition, as it is a naturally
occurring peptide, the possibility exists that there may be a breakdown of
glutathione in plasma by peptidases, as in other tissues and at blood-brain
barrier itself (Meister and Tate, 1976). Therefore, its clinical value as a
therapeutic agent, if administered by a peripheral route, should be minimal.
However, since recent investigations support the concept of a selective
transcytosis for many peptides across an intact blood-brain barrier (Pardridge,
1986) and since the finding that in idiopathic PD the locus coeruleus, which
helps to preserve the integrity of blood-brain barrier functions, is damaged
(Tomonaga, 1983, Harik and McGunigal, 1984) the authors administered GSH as
l-hour infusion two times daily for 30 days in PD patients, to investigate a
possible therapeutic effect of this peptide, after peripheral administration.
Actually, recent experimental evidences have shown blood-brain extractlon of
circulating GSH in a brain perfusion model, and the transcytosis of intact GSH
into the brain parenchyma without breakdown (Zlokovic et al., 1994).
Effects of Intravenous GSH in Parkinson’s Disease
The results of our open study indicate that in PD this peptide given i.v. may
reach its specific target in the brain (i.e, the nigra cells) and may have a
significant beneficial effect on several parkinsonian signs. In particular, as
1168 G.P. Sechi et cd.
shown in Fig 3, the therapeutic effect of GSH on hypokinesia appears to be
correlated to the severity of the symptom. This peptide was also effective in
reducing the RT in one patient, but failed in other four. In this patient, GSH
apparently improved the RT more than levodopa-carbidopa. In our opinion, since
the dosages of levodopa-carbidopa and GSH used are not comparable, to draw
this conclusion is incorrect. Once GSH was stopped the therapeutic effect
lasted for 2-4 months. This finding, in our opinion, is a strong evidence against
a placebo effect and this may indicate a protective effect of the drug on the
rate of progression of PD. However, this does not necessarily exclude a
symptomatic effect of GSH. These concepts are supported by the results of two
double-blind studies on the use of selegine, or bromocriptine versus placebo in
PD (Teychenne et al., 1982, Myllyla et al., 1992). Indeed, in these studies, the
mean CURS scores in the placebo group returned to baseline after about l-
month, and the symptomatic effect of bromocriptine, once stopped, did not last
for more than four weeks (Teychenne et al., 1982, Myllyla et al., 1992).
Hvoothetical Mechanisms Underlying the TheraDeutic Fffect of GSH
The mechanism underlying the therapeutic effect of GSH in PD is unknown.
According to the most basic neurochemical abnormality in the brain of PD
patients (i.e., the marked loss of dopamine in the nigrostriatal neuron system)
(Ehringer and Hornykiewiez, 1960), an action of GSH at dopaminergic synapses
(presynaptically or postsynaptically) can be hypothesized. In particular, based
on the theoretical notion that decreasing the oxidative load in substantia nigra
may slow disease progression, it would seem that GSH, because of its
antioxidant properties (e.g., reduced formation of hydrogen peroxide), may be
able to protect the striatonigral cells and foster dopaminergic activity
(Jenner, 1993). Recent evidences of glutamate uptake inhibition by oxygen free
radicals in rat cortical astrocytes fit this hypothesis (Volterra et al., 1994).
Another important biological function that has been ascribed to glutathione is
the role in translocation of amino acids, and possibly also peptide, across cell
membranes (Meister and Tate, 1976). This function may be important for the
transport of substrate and specific proteic neurotrophic factors into the
dopaminergic neurons of the substantia nigra (Tooyama et al., 1993). Given the
reduction in the levels of GSH in cells of substantia nigra, in PD this function
is likely impaired. A replacement therapy with exogenous GSH may contribute
to its reinstatement. A controlled study of this peptide for the treatment of PD
seems warranted.
intravenous GSH in PD 1169
Conclusion
The findings indicate that in PD GSH given i.v. may reach its specific target
in the brain (i.e., the nigra cells), it has a significant beneficial effect on
several parkinsonian signs and possibly retards the progression of the disease.
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Inquiries and reprint requests should be addressed to:
GianPietro Sechi, M.D.
Neurological Clinic
Viale S. Pletro, 10
07lOOSassari
Italy.
