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Vol. 51, Issue 4, 593-628, December 1999
Department of Pharmacology and Toxicology, University of Kuopio, Kuopio, Finland (P.T.M.); and Department of Neurology, University of Helsinki, Helsinki, Finland (S.K.)
I. Introduction
II. COMT Gene and Proteins
A. One COMT Gene and Two Proteins
B. Three-Dimensional Structure of COMT
C. Kinetic Reaction Mechanism of COMT
D. Other Enzymological Aspects
E. Genetic Polymorphism: Variants with Different Thermostability
F. Distribution of COMT
1. Brain COMT.
2. COMT in Other Tissues.
G. General Importance of COMT
1. Substrates of COMT.
2. Quantitative Role of COMT in Metabolism of Catecholamines.
3. COMT Knockout Mice.
III. COMT Inhibitors
A. First-Generation COMT Inhibitors
B. Second-Generation COMT Inhibitors
C. Properties of New Compounds
1. COMT Inhibition.
2. Effects on L-Dopa and Catecholamine Metabolism.
3. Microdialysis Studies.
4. Voltammetric Studies.
5. Estrogen Metabolism and Role of COMT and COMT Inhibitors.
6. Behavior.
7. S-Adenosyl-L-Methionine-Saving Effect of COMT Inhibitors.
8. Other Effects of COMT Inhibitors.
9. Physicochemical Properties and Animal Pharmacokinetics.
10. Toxicity.
11. Conclusions from Animal Studies.
IV. Positron Emission Tomography Studies
V. Practical and Theoretical Clinical Uses of COMT Inhibitors
VI. Human Studies with COMT Inhibitors
A. Human Pharmacokinetics of COMT Inhibitors
B. COMT Inhibition
C. Effect on Levodopa Pharmacokinetics
D. Effect on 3-OMD Levels
E. Effect on Plasma Catecholamine Metabolism
F. Clinical Efficacy
G. Safety
H. Drug Interactions
VII. Summary
VIII. Future Aspects
Acknowledgments
References
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I. Introduction |
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 Top NextReferences
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Axelrod et al. (1958)
first described the enzyme-catalyzed
O-methylation of catecholamines and other catechols in the
late 1950s. The enzyme responsible for the O-methylation,
catechol-O-methyltransferase (COMT; EC
2.1.1.6),2 was
partly purified and characterized by the same group (Axelrod and
Tomchick, 1958). The subsequent basic research on COMT and the first
COMT inhibitors introduced between 1958 and 1975 have been extensively
reviewed by Guldberg and Marsden (1975).
The interest in COMT was rekindled in the late 1980s when the
potent and selective second-generation COMT inhibitors were developed
(Männistö and Kaakkola, 1989, 1990), and soon the structures of the two isoforms of COMT and the gene were characterized and COMT polypeptide cDNAs were cloned (Salminen et al., 1990; Bertocci
et al., 1991; Lundström et al., 1991). Several review articles
have recently dealt with this development (Männistö and
Kaakkola, 1989, 1990; Männistö et al., 1992b, 1994; Roth, 1992; Kaakkola et al., 1994a; Dingemanse, 1997), culminating in the
marketing of two new COMT inhibitors. This review concentrates on the
recent information on the biochemistry and molecular biology of COMT
and on the pharmacology and clinical efficacy of the new selective and
relative nontoxic COMT inhibitors.
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II. COMT Gene and Proteins |
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TopPreviousNextReferences
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A. One COMT Gene and Two Proteins
The structural organization of the COMT gene has been reviewed in
detail by Lundström et al. (1995). There is one single gene for
COMT, which codes for both soluble COMT (S-COMT) and membrane-bound
COMT (MB-COMT; Salminen et al., 1990; Lundström et al., 1991). In
the application of the in situ hybridization and cell hybrid
techniques, the COMT gene in humans was localized to chromosome 22, band q11.2 (Grossman et al., 1992a; Winqvist et al., 1992). The human
COMT gene contains six exons with the two first exons being noncoding.
The expression of the COMT gene is controlled by two distinct promoters
located in exon 3 (Salminen et al., 1990; Lundström et al.,
1991). The distal 5' promoter (P2) regulates synthesis of 1.9-kb (rat)
and 1.5-kb (human) mRNA species. This mRNA can code for both MB-COMT
and S-COMT proteins by using the leaky scanning mechanism of
translation initiation (Tenhunen and Ulmanen, 1993; Tenhunen et al.,
1993, 1994). The expression of the shorter transcript (1.6 kb in rat
and 1.3 kb in human) is regulated by the P1 promoter, which is located
between S-COMT and MB-COMT ATG start codons and partly overlaps the
MB-COMT coding sequence. MB-COMT AUG translation initiation codon is
not included in these transcripts, which therefore can code only for S-COMT polypeptide (Tenhunen et al., 1993, 1994; Tenhunen and Ulmanen,
1993).
In most human tissues, there are both transcripts, but in the human
brain, only the longer transcript was found in 16 regions studied (Hong
et al., 1998). When S-COMT and MB-COMT polypeptides of various rat and
human tissues were quantified through Western blot analysis, S-COMT was
usually dominant by a factor of 3 or higher. The only exception was the
human brain, where 70% of the total COMT polypeptides was MB-COMT and
30% was S-COMT, demonstrating the bifunctionality of the 1.5-kb
transcript (Tenhunen et al., 1993, 1994). The varying expression levels
of the human COMT promoters suggest regulation by some tissue-specific
transcription factors. In fact, human COMT promoters contain several
putative binding sites for such factors that may cause the variability
of COMT gene expression in different tissues (Tenhunen et al., 1994).
Both rat and human S-COMTs contain 221 amino acids, and the molecular
masses are 24.8 and 24.4 kDa, respectively. The human S-COMT is
81% identical with the respective rat enzyme (Lotta et al., 1995;
Lundström et al., 1995). Rat MB-COMT contains 43 additional amino
acids, and human MB-COMT contains 50 additional amino acids. The
corresponding molecular masses of MB-COMT are 29.6 and 30.0 kDa. Of
these extra amino acids, 17 (rat) and 20 (human) function as
hydrophobic membrane anchors (Salminen et al., 1990; Tilgmann and
Kalkkinen, 1990, 1991; Bertocci et al., 1991; Lundström et al.,
1991). The remainder of the MB-COMT molecule is suspended in the
cytoplasmic side of the intracellular membranes (Bertocci et al., 1991;
Lundström et al., 1991; Ulmanen and Lundström, 1991).
B. Three-Dimensional Structure of COMT
Rat S-COMT has been recently crystallized at 1.7- to 2.0-Å
resolution (Vidgren et al., 1991, 1994), and the critical atomic structures have been described in detail (Vidgren and Ovaska, 1997).
Therefore, only some of the most important aspects are repeated here.
COMT has as a single domain 
/
-folded structure in which eight
-helices are arranged around the central mixed -sheet. The active
site of COMT consists of the
S-adenosyl-L-methionine-
(AdoMet)-binding domain and the actual catalytic site. The binding
motif of the AdoMet site is similar to the Rossman fold, which is a
common feature of many nucleotide binding proteins. The crystal
structures of several of the methyl transferases that have been
characterized are strikingly similar in the AdoMet-binding regions. The
catalytic site is formed by a few amino acids that are important for
the binding of the substrate, water, and Mg2+ and
for the catalysis of O-methylation (Fig.
1). The Mg2+, which
is bound to COMT after AdoMet binding, converts the hydroxyl groups of
the catechol substrate to be more easily ionizable. Near one of the
hydroxyl groups of the substrate, there is a lysine residue (Lys144) in
COMT that accepts the proton from that hydroxyl, and subsequently the
methyl group from the AdoMet is transferred to the hydroxyl group.
Lysine acts as a general catalytic base in this base-catalyzed
nucleophilic reaction. Mg2+ has an octahedral
coordination to two aspartic acid residues (Asp141 and Asp169), to one
asparagine (Asn170), to both catechol hydroxyls, and to a water
molecule. Hence, Mg2+ ions control the
orientation of the catechol moiety. In addition, the "gatekeeper"
residues Trp38, Trp143, and Pro174 that form the hydrophobic
"walls" and that define the selectivity of COMT toward different
side chains of the substrate participate directly in the methylation
reaction by keeping the planar catechol ring in the correct position
(Fig. 1). They contribute significantly to the binding of the
substrates (and inhibitors) of COMT (Vidgren et al., 1991, 1994, 1999;
Vidgren and Ovaska, 1997). Although Mg2+ ions are
crucial for COMT, most other small molecule methyltransferases are not
Mg2+ dependent (Fujioka, 1992).
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C. Kinetic Reaction Mechanism of COMT
COMT catalyzes the transfer of the methyl group of AdoMet to one
of the hydroxyl groups of the catechol substrate in the presence of
Mg2+ (Guldberg and Marsden, 1975). Methylation of
the 3'-hydroxyl is much more common than that of the 4'-hydroxyl, for
reasons that are discussed later. The mechanism and kinetics of the
O-methylation reaction have been studied using partially
purified enzyme preparations from various sources, most commonly from
rat liver and rat or human brain, and recently also with the use of
recombinant enzymes. The stereochemical course of the reaction has
shown that the methyl transfer proceeds through a direct nucleophilic
attack by one of the hydroxyl groups of the catechol substrate on the
methyl carbon of AdoMet in a tight SN2-like
transition state (Woodard et al., 1980).
A sequential ordered mechanism has been proposed based on product
inhibition studies (Jeffery and Roth, 1987), but there is some doubt as
to their accuracy. AdoMet does seem to be the first substrate to bind,
and S-adenosyl homocysteine is the last product to
dissociate from the enzyme (Rivett and Roth, 1982; Tunnicliff and Ngo,
1983). However, now that the crystal structure of S-COMT has been
resolved, it is possible to extend the kinetic studies to include
recombinant S-COMT and MB-COMT with many different substrates. Thus,
Lotta et al. (1995) proposed a reformulation of the kinetic behavior
and the ordered mechanism of O-methylation. The active site
of COMT, which is located in the outer surface of the enzyme, is a
shallow groove on the surface of COMT. This is the same in both S-COMT
and MB-COMT (Vidgren et al., 1994; Vidgren and Ovaska, 1997). AdoMet is
able to bind even without Mg2+. The binding
pocket of the methionine portion of AdoMet is deeper within the protein
than the Mg2+ site, and it would be impossible
for AdoMet to bind after Mg2+. Moreover, the
catechol substrate cannot bind before AdoMet because then it would also
be impossible for AdoMet to reach its binding site in the narrow groove
located deep in the COMT molecule (Vidgren et al., 1994; Vidgren and
Ovaska, 1997). Therefore, the order in which the compounds bind is as
follows: AdoMet binds first, followed by Mg2+
and, finally, the catechol substrate. This reaction cycle differs from
the previous suggestion that Mg2+ binds to COMT
in a rapid equilibrium before the addition of AdoMet (Jeffery and Roth,
1987).
Although S-COMT and MB-COMT have identical kinetic mechanisms (Ca2+ inhibition, Mg2+ requirement, pH optimum, a similar Km value for AdoMet, recognition by S-COMT antiserum), S-COMT and MB-COMT are certainly different enzymes, and MB-COMT is not a precursor of S-COMT (see above).
Based on early studies with crudely purified enzymes, S-COMT has a high
Km value for dopamine but a very high
capacity (Vmax from 50 pmol/min × mg protein in skeletal muscle to values as high as 14,690 pmol/min × mg protein in the liver). MB-COMT has a much lower
Km value but a low capacity (2-40
pmol/min × mg protein; Guldberg and Marsden, 1975; Roth, 1992).
It is noteworthy that the Vmax values
are strongly dependent on the enzyme activities in various tissues
rather than on the basic kinetic constants of these enzymes.
Physiological substrate concentrations and possible differences in
substrate selectivity have to be considered when the relative
importance of either enzyme subtype is assessed. Dopamine levels in
striatum and hypothalamus of brain homogenates are about 65 and 3 µM,
respectively. The striatal and hypothalamic noradrenaline
concentrations are 0.8 and 12 µM, respectively. It seems that at the
concentrations of catecholamines naturally present, MB-COMT may be more
important in their metabolism (Roth, 1992). According to Roth and
associates (Rivett et al., 1982; Rivett and Roth, 1982; Roth, 1992),
MB-COMT is the predominant enzyme at dopamine concentrations of <10
µM and at noradrenaline concentrations of <300 µM.
In addition, the recombinant human MB-COMT
(Km = 10 µM) has a higher affinity
for catechol substrates than S-COMT
(Km = 108 µM; Malherbe et al.,
1992). Lotta et al. (1995) also reported that the catalytic number
(Vmax) of rat recombinant S-COMT was slightly higher than that of MB-COMT with all four substrates studied.
It is notable that in the study of Lotta et al. (1995), Vmax values were calculated taking
into account the actual enzyme concentration. Thus,
Vmax can be given as U/min, and it
represents the catalytic number
(kcat). At saturating substrate
concentrations, S-COMT functioned two times more efficiently, but the
catalytic number for both COMT isoforms was similar for all substrates, nor was there any substrate selectivity. The
Km and
kenzyme values varied greatly between
the two isoforms and were strongly dependent on the substrate. Malherbe
et al. (1992) reported that S-COMT has a 15-fold higher
Km value and a 5-fold higher
kenzyme value for catecholamines than
MB-COMT. However, this is not the case for dihydroxybenzoic acid (DBA)
and L-3,4-dihydroxyphenylalanine (L-dopa; Table 1).
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The catalytic sites of S-COMT and MB-COMT have identical amino acid sequences, but the membrane-bound portion of MB-COMT or the charged membrane itself causes more favorable, although poorly characterized, binding interactions even though there is no conformational change in the basic enzyme structure of MB-COMT.
New data generated using recombinant enzyme isoforms have confirmed
earlier results but also added further detail to the picture, particularly regarding the binding differences of the various COMT
substrates (Lotta et al., 1995; Tables 1 and
2). The atomic structure and the sequence
comparison reveal that all residues that are important for the binding
of the substrates and for the catalytic activity are similarly
conserved in human and rat COMT. Only two amino acids are different in
the active site. When the three substrates (L-dopa,
dopamine, and DBA), which have different affinities for the active site
of COMT, are compared, it is apparent that the kinetic differences are
due to interactions of the substrate side chains with COMT residues.
Generally, the binding of the catechol ring is similar to binding of
enzyme to nitrocatechol-type inhibitors (e.g., OR-486 or OR-1840) when
this is viewed with crystallized rat S-COMT (Vidgren et al., 1994). DBA
has a charged carboxyl moiety, but the molecule is planar and fits well
between the gatekeepers Trp38 and Pro174 (Fig. 1), and thus it has high affinity. Dopamine has a positively charged amino group that, despite
its rotational freedom, still makes a repulsive contact with one of the
gatekeepers. L-Dopa has the largest, double-charged side
chain, and therefore the propulsions are strong and its affinity to
COMT is lower.
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COMT is able to methylate only one of the two catechol hydroxyls. Both
COMT isoforms favor 3-O-methylation, and MB-COMT is even
more regioselective than S-COMT (Table 2). The
meta/para ratio is higher, 22 to 88 (depending on
the substrate) in MB-COMT than in S-COMT, 4 to 15 (depending on the
substrate; Lotta et al., 1995). The reason for favoring
3-O-methylation over 4-O-methylation may be that
as the p-hydroxyl group (i.e., 4-O-hydroxyl)
approaches the AdoMet; this forces the side chain to become orientated
in an unfavorable position with the hydrophobic protein residues of the
catalytic site. As stated, L-dopa has the
strongest repulsive interactions, which are reflected not only in its
high Km value but also as in the high
3-O-methylation/4-O-methylation ratio (Table 2).
Recently, molecular dynamic simulation studies have been used to
explain the preference of meta-O-methylation over
para-O-methylation. The catechol ring has a tilt
of about 30 degrees compared with that of the X-ray structure of the
active site. This directs any substituent at the
para-position of the catechol ring into a hydrophobic pocket
formed by Trp38 and Tyr200. Hydrophobic substituents are accommodated
in this pocket so that para-O-methylation is
favored, whereas polar substituents are repelled, making
meta-O-methylation favorable (Lan and Bruice,
1998). In addition, the distances of the hydroxyls to the active methyl
group of AdoMet are very different (2.6 versus 4.8 Å), so no
competition for the methylation site is possible (Vidgren et al.,
1999). Different meta/para ratios of substituted
catechols are solely a consequence of their relative ability to bind in
two dissimilar orientations to the active site of COMT (Vidgren et al.,
1999).
Those catechols that contain electronegative substituents (like
NO2, CN, and F) are potent inhibitors, but poor
substrates, of COMT (Bäckström et al., 1989; Borgulya et
al., 1989). This point has been recently clarified in a semiempirical
study using dinitrocatechol (OR-486) as a model inhibitor (Ovaska and
Yliniemelä, 1998). As mentioned, Lys144 acts as a general base in
the methylation and can activate one of the catechol hydroxyls before
the nucleophilic attack of the methyl group of AdoMet. The
electronegative nitro groups reduce the nucleophilicity of the ionized
catechol hydroxyls (Fig. 1). Therefore, dinitrocatechol itself is not
methylated at all but instead is a potent COMT inhibitor. Among the
mononitro catechols, the decrease in nucleophilicity is evidently less
than that in dinitrocatechol, and the ultimate effect depends on the side chain. For instance, entacapone is to all intents not
O-methylated at all, whereas tolcapone is
O-methylated by about 3% (Dingemanse, 1997).
D. Other Enzymological Aspects
COMT is not easily induced or suppressed. The capacity of COMT in
the peripheral tissues is probably so high that only a minor fraction
of the protein is ever needed. Therefore, general peripheral COMT
inhibition may be difficult to achieve. This may be crucial for the
safety of COMT inhibitors. Locally, such as in the gastrointestinal mucosa (Schultz and Nissinen, 1989), the new powerful COMT inhibitors can suppress COMT activity sufficiently to cause significant changes in
the metabolism of L-dopa.
Some special treatments or situations may increase COMT activity, but
most of them cause at best only a doubling of activity. Very early, it
was found that long-term treatment with pyrogallol could elevate the
COMT activity in the liver (Guldberg and Marsden, 1975). Dipyridamole
(Li et al., 1991), exogenous AdoMet (Baldessarini, 1987), and butylated
hydroxyanisole (Lam, 1988) can slightly increase COMT activity.
Pregnancy and progesterone treatment seem to induce COMT activity in
the uterus. In the rat, estrogen exposure results in decreased hepatic
COMT activity (Cohn and Axelrod, 1971; Parvez et al., 1975). On the
other hand, subchronic estrogen treatment increases COMT immunostaining
in hamster kidney (Weisz et al., 1998b). There also is a gender
difference. The COMT activity in the liver of male subjects is about
30% higher than that in females (Boudikova et al., 1990). During
aging, COMT activity in the liver increases by about 10-fold from birth
to adulthood (Guldberg and Marsden, 1975). A similar trend occurs in
the kidney, where Km values increase
about 5-fold during aging (Vieira-Coelho and Soares-da-Silva, 1996).
This could be explained by assuming that the predominant form of COMT
is not the same in newborn rats and adult animals. Some diseases seem
to be associated with altered COMT activities. COMT activity in the
spinal cord appears to be decreased in Huntington's disease (McGeer et
al., 1993), but it is slightly increased in amyotrophic lateral
sclerosis where monoamine oxidase (MAO)-B activity was
substantially elevated (Ekblom et al., 1993). Chromosomal microdeletion
at 22q11-13 (where the human COMT gene is also located) has
been associated with a number of defects or syndromes (Lachman et al.,
1996a). This phenomenon and genetic COMT polymorphism in various
diseases are discussed later (see Genetic Polymorphism: Variants
with Different Thermostability).
We also made an anecdotal finding that high concentrations of ethanol
can reduce MB-COMT activity but increase S-COMT activity in recombinant
enzyme forms. However, the concentrations needed are from 50 to 1000 mM, which could scarcely be attained even in cases of ethanol
intoxication (Reenilä et al., 1995).
E. Genetic Polymorphism: Variants with Different Thermostability
The level of COMT enzyme activity is genetically polymorphic in
human tissues with a trimodal distribution of low
(COMTLL), intermediate
(COMTLH), and high
(COMTHH) activities (Weinshilboum and
Raymond, 1977; Boudikova et al., 1990; Jeanjean et al., 1997). This
polymorphism, which according to segregation analysis of family studies
is caused by autosomal codominant alleles, leads to 3- to 4-fold
differences in COMT activity in human erythrocytes and liver
(Weinshilboum and Raymond, 1977; Boudikova et al., 1990; Jeanjean et
al., 1997). Low COMT activity is associated with enzyme thermolability,
even at 37°C (Scanlon et al., 1979; Spielman and Weinshilboum, 1981;
Boudikova et al., 1990). Recently, the molecular basis of the
thermolability was revealed with the baculovirus expression system;
namely, substitution of Val108 by Met108 in the S-COMT (or the
corresponding amino acids 158 in the MB-COMT) is caused by transition
of guanine to adenine at codon 158 of the COMT gene
(Grossman et al., 1992b; Lotta et al., 1995). Although there are some
other mutations in the COMT gene (Lachman et al., 1996b), it seems
quite established that the low thermal stability and the low COMT
activity go together. Because there is only one COMT gene without any
known tissue-specific splice variants (Tenhunen et al., 1994;
Lundström et al., 1995), it is likely that the codon
108/158 polymorphism, causing the change in thermostability, also
leads to functional alterations of COMT in all tissues (Lachman et al.,
1996b). The catalytic activity per se is the same with the two
variants, and the labile enzyme variant is fully stabilized by AdoMet
binding. In the following discussion, we do not necessarily go to the
methodological details when describing the association of COMT activity
with a number of diseases. In publications printed after 1996, the
molecular difference may be demonstrated. Earlier, either the
thermostability or only the COMT activity was reported.
Polymorphism of COMT activity could have clinical implications, but the
true relationships in about 30 genetic mapping studies with a number of
diseases have not been very impressive. There are, however, three or
four disorders in which some relationship has been observed. First,
there is an obsessive-compulsive disorder in which the affected persons
follow various anxiety-reducing rituals, which seems to be correlated
to low COMT activity allele (Karayiorgou et al., 1997). Second, low
COMT activity allele appears to have some association with aggressive
and highly antisocial impulsive schizophrenia (Strous et al., 1997a,b;
Lachman et al., 1998). A special velo-cardio-facial syndrome in which
the chromosome 22q11 is deleted (including the COMT gene)
also carries with it similarly bizarre behavior, but several types of
other symptoms, including schizophrenia, may appear (Lachman et al.,
1996a,b; Papolos et al., 1996). However, generally there is only a weak (Ohmori et al., 1998) or no association between COMT activity alleles
and schizophrenia (Chen et al., 1996; Daniels et al., 1996; Riley et
al., 1996; Wei et al., 1996; Karayiorgou et al., 1998). Paradoxically,
in one study, the high activity allele was preferentially transmitted
from healthy parents to their schizophrenic children (Li et al., 1996).
Recently, Tiihonen et al. (1999) reported a clear indication of the
association of the late-onset alcoholism (type 1) and the low-affinity
allele of COMT in a Finnish population. When 123 alcoholics were
compared with 246 race- and gender-matched controls, the odds ratio for
alcoholism of subjects with COMTLL was 2.51 over those with COMTHH. The frequency of
low allele was also significantly higher in the alcoholics than in 3140 Finnish blood donors representing a general population. The estimate
for population etiological percentage of the
COMTLL in type 1 alcoholism was 13.3%.
With respect to depression, the results are variable. Several studies
do not show any relationship between depression and COMT (BIOMED
European Bipolar Collaborative Group, 1997; Gutíerrez et al.,
1997; Kunugi et al., 1997b), although some show a low COMT activity
allele or a low COMT activity in the erythrocytes in patients with
major depression but not in those with the bipolar form (Karege et al.,
1987; Ohara et al., 1998b). In some patients, the link seems to be with
bipolar manic-depressive illness (Li et al., 1997). Recently it has
been shown that the low COMT activity allele is dominant in
rapid-cycling (cycle 1-2 days) bipolar manic-depressive disorder
(Kirov et al., 1998; Papolos et al., 1998). These findings confirm the
results of an earlier report that patients with velo-cardio-facial syndrome, which includes a rapid-cycling bipolar disorder, have low
COMT activity (Lachman et al., 1996a; Papolos et al., 1996). Interestingly, polysubstance abusers have been reported to more commonly have the high COMT activity allele than the controls (Vandenbergh et al., 1997). No association has been found between anxiety disorders and COMT polymorphism (Ohara et al., 1998a).
Some ethnic differences have been recognized. For instance, the
frequency of the low COMT activity allele is lower in Kenyan than in
Caucasian or South-West Asian individuals (McLeod et al., 1998).
However, black Americans have higher COMT activity than white Americans
(McLeod et al., 1994). Low COMT activity is found in a Saami population
(Klemetsdal et al., 1994).
The association of the COMT alleles with Parkinson's disease (PD) has
been extensively studied, but usually no association has been found
(Hoda et al., 1996; Syvänen et al., 1997; Xie et al., 1997).
However, some Japanese individuals with the low COMT activity allele
may have an increased risk for PD (Kunugi et al., 1997a; Yoritaka et
al., 1997). As cited above, different populations have different
frequencies of COMTLL and
COMTHH alleles and therefore may have
variation in individual responses to therapy with a drug preparation
containing L-dopa (levodopa; Reilly et al., 1980;
Rivera-Calimlim and Reilly, 1984; Klemetsdal et al., 1994). It would be
interesting to know whether the confirmed COMTLL PD patients really can benefit more
from the levodopa therapy than COMTHH
patients, as has been suggested based on COMT activity analysis from
erythrocytes (Reilly et al., 1980). It is also known that patients with
high COMT activity in their erythrocytes have encountered more adverse
effects during levodopa therapy and have had more frequent on-off
effects than patients with low COMT activity (Reilly et al., 1983;
Rivera-Calimlim and Reilly, 1984).
Finally, low COMT activity allele is found more frequently in
patients with breast cancer than in healthy controls (Lavigne et al.,
1997), particularly in women with menopausal symptoms (Thompson et al.,
1998). This phenomenon may be related to the decreased metabolism of
catecholestrogens because COMT also metabolizes these compounds (see
Estrogen Metabolism and Role of COMT and COMT Inhibitors).
However, a recent extensive case-control study on patients with
invasive breast cancer in North Carolina (654 cancer patients and 652 controls) did not show any relation of the COMT genotype to the breast
carcinoma (Millikan et al., 1998).
In addition to humans, other mammals can have different genetically
determined COMT activity. There are two rat strains with different
activity levels (Weinshilboum et al., 1979; Roth et al., 1990);
however, the rat polymorphism has some other structural basis from the
human polymorphism because both rat and pig have Leu108 (not Met108 or
Val108; Salminen et al., 1990) in the critical site of the COMT
polypeptide, which determines the level of activity and thermal
stability (Salminen et al., 1990; Malherbe et al., 1992).
F. Distribution of COMT
In mammals, COMT is widely distributed throughout the organs of
the body. COMT is an intracellular enzyme. As discussed (see One
COMT Gene and Two Proteins), the COMT protein in vertebrates appears mostly in a soluble form (as S-COMT), and only a minor fraction
is in the particular form (as MB-COMT; Guldberg and Marsden, 1975;
Roth, 1992).
1. Brain COMT.
The cellular localization of COMT has been
studied in several ways. A number of lesion studies (Rivett et al.,
1983; Kaakkola et al., 1987) and immunohistochemical studies (Karhunen
et al., 1995a; Lundström et al., 1995) have demonstrated that
there is no significant COMT activity in presynaptic dopaminergic
neurons, but some activity is present in postsynaptic neurons and
substantial activity is located in glial cells.
). Because there are no
S-COMT- and MB-COMT-specific antisera available, it has not been
possible to separately analyze the tissue distribution of the two
enzymes forms. Immunoblotting analysis of the primary cell cultures of
the rat brain cells shows that both isoforms of COMT occur in
astrocytes, oligodendrocytes, and neurons (Karhunen et al., 1995b).
Some indication of subcellular distribution can be obtained through
differential centrifugation. The mammalian S-COMT activity resides in
the nonsedimenting, cytoplasmic fractions, and MB-COMT activity resides
in the sedimenting fractions, the equivalent of the microsomal fraction
(Jeffery and Roth, 1984; Roth, 1992).
With immunoblotting and COMT activity determinations for the analysis
of the COMT isoforms, it was shown that both in rat brain and in
baculovirus-infected insect cells, the MB-COMT polypeptide resides in
the subcellular fractions containing endoplasmic reticulum and cell
membranes. These two components could not be separated in this study
(Tilgmann et al., 1992). In mammalian cell preparations overexpressing
either S-COMT or MB-COMT, the subcellular localization has been studied
in detail (Ulmanen et al., 1997). Overexpressed S-COMT was localized in
cytosol and in the nucleus, whereas MB-COMT was a microsomal protein.
This was determined through double immunostaining against an
established marker protein of the rough endoplasmic reticulum. MB-COMT
was located in the rough endoplasmic reticulum, facing the cytoplasm.
It is worth noting that no MB-COMT has been found in the cell membrane
(Ulmanen et al., 1997).
When the cellular distribution of COMT in the rat striatum was studied
in an attempt to selectively destroy glial cells with fluorocitrate, we
found that initially there was a decrease in COMT activity at 1 and 2 days after lesioning, followed by a marked increase at 3 days. At the
same time, a microglial marker (alkaline phosphodiesterase) was
increased. Immunohistochemical analysis also revealed a major increase
in microglia (OX-42-immunoreactive cells) but not of astroglia (glial
fibrillary acidic protein immunoreactive cells). No neuronal COMT
immunoreactivity was detectable in these studies (Reenilä et al.,
1997).
2. COMT in Other Tissues.
COMT has been found in practically
all mammalian tissues investigated. The highest COMT activity in both
rat and humans is in the liver, followed by the kidneys and
gastrointestinal tract (both stomach and intestine; Nissinen et al.,
1988b; Schultz and Nissinen, 1989; Männistö et al., 1992b).
These findings have been confirmed and expanded to spleen and
submaxillary glands through the use of immunohistochemistry (Karhunen
et al., 1994; Lundström et al., 1995). In pancreas, COMT
immunoreactivity was found in and 
cells but not in cells
(Karhunen et al., 1994). The importance of pancreatic COMT activity has
also been demonstrated through other means. After the pretreatment of
conscious rats with OR-486 (dinitrocatechol), a potent
nitrocatechol-type COMT inhibitor, the uptake of radioactivity from
[13C]L-dopa into the pancreas was
increased by 4-fold. Most of the radioactivity was derived from
[13C]dihydroxyphenylalanine
([13C]DOPAC; Bergström et al., 1997).
Positron emission tomography (PET) studies have demonstrated high COMT
activity in kidney, liver, intestine, stomach, spleen, lungs, and heart
of mice (Ding et al., 1996); they also confirmed high COMT activity in
baboon liver and kidney.
). COMT mRNA (mostly that of S-COMT) has also been visualized
in epithelial cells of proximal tubules, the thick ascending limb of
loop of Henle, and the ureter (Meister et al., 1993), where the enzyme
is thought to regulate the metabolism of dopamine and other
catecholamines. COMT mRNA has been detected in the prenatal kidney at
gestational day 18 (Meister et al., 1993). After birth, the
Km values of rat kidney COMT activity appear to increase as a function of age from 3.3 µM at age 3 days to
16.9 µM at age 30 days (see Other Enzymological Aspects).
Vmax values (which are dependent on
the total amount of enzyme in the tissue, which was not
measured) were not much altered (Vieira-Coelho and Soares-da-Silva,
1996). The same authors also claim that the inhibitory activity of
tolcapone on kidney COMT would be radically altered as a function of
the age. However, because the changes in the COMT protein content were
not measured, this conclusion remains uncertain. The same is true for
the comparison between kidney and liver COMT inhibition with tolcapone.
Human kidney COMT activity was also assayed by De Santi et al. (1998b),
who found it to be 159 pmol/min × mg protein when DBA was
used as the substrate. COMT activity in the liver was three times
higher than kidney activity, and in the duodenum, activity about 30%
less than that in kidney was noted. In that study, entacapone was found
to be severalfold more active than tolcapone as a COMT inhibitor in all
tissues studied. This is an unexpected result that has not been confirmed.
Although it has been demonstrated that the metabolism of the newly
formed dopamine in the rat kidney is primarily mediated through MAO,
with only a minor involvement of COMT (Fernandes and Soares-da-Silva,
1994), both nitecapone (Eklöf et al., 1997) and entacapone
(Hansell et al., 1998) induced a copious diuresis and natriuresis,
which in both cases was inhibited by a dopamine D1 antagonist. However, the amounts of dopamine
excreted were only marginally increased by entacapone but much more by
N-(2-pyridone-6-yl)-N'N'-di-n-propylformamidine (CGP 28014), which is not at all a COMT inhibitor (see below; Hansell
et al., 1998). Nitecapone was shown to enhance
Na+,K+-ATPase inhibition in
the proximal tubular cells (Eklöf et al., 1997). Finally, there
seems to be a weak but significant correlation of
histamine-N-methyltransferase and COMT activities in the
human renal cortex (De Santi et al., 1998a).
Opossum kidney cells in culture can synthesize and metabolize dopamine
(by MAO and COMT) in a manner similar to rat renal tubular cells.
Therefore, these cultured cells can be used as an easily available and
standardized model system to explore the role of dopamine in kidney
function (Guimarães et al., 1997).
In hamster kidney, a 2- or 4-week estrogen treatment induces COMT
immunoreactivity, as well as in the nucleus, where only S-COMT was seen
(Weisz et al., 1998b), supporting the nuclear localization of S-COMT as
discussed. This probably is a compensatory mechanism inducing a
metabolizing enzyme to oppose the formation of electrophilic quinones
and semiquinones from catecholestrogens, particularly in this animal
species (see Estrogen Metabolism and Role of COMT and COMT
Inhibitors).
Rather high COMT activities have been described in human lung by De
Santi et al. (1998b) and in rat lung by Bryan-Lluka (1995). There also
is a substantial amount of COMT in the eye, in both the ciliary body
and the retinal ganglion cell layer (Karhunen et al., 1994). COMT
activity has also been demonstrated in spinal membranes of monkeys and
pigs (Kern et al., 1995).
COMT activity is detectable in the skin, with the activity being higher
in epidermis than in dermis (Bamsdah, 1969). COMT is found in
keratocytes, where it may metabolize epinephrine, but also in
melanocytes, which make up only 3 to 5% of the epidermal cell
population (Smit and Pavel, 1995). The epidermis of patients with
vitiligo with contains more COMT than does the epidermis from healthy
controls (Lepoole et al., 1994). Intermediate indolic compounds, like
5,6-dihydroxyindole-2-carboxylic acid, are formed in the synthesis of
eumelanin from tyrosine. These intermediates can be readily oxidized to
toxic quinone derivatives. The polymerization of these potentially
toxic intermediates to melanin can be considered a detoxification
process. COMT activity may further reduce the amount of
dihydroxyindoles and therefore protect normal melanocytes against their
own reactive compounds generated during melanogenesis. If the
metabolism of dihydroxyindoles by COMT is inhibited, these metabolites
can accumulate and become harmful to normal skin (Smit and Pavel,
1995). However, there is a possibility for a novel therapeutic role for
COMT inhibition in melanoma treatment (Shibata et al., 1993; Smit et
al., 1994). It has been shown that malignant melanocytes contain both
S-COMT and MB-COMT. At physiological concentrations of
dihydroxyindoles, MB-COMT may be more relevant and functionally more
significant than S-COMT (Shibata et al., 1993; Smit and Pavel, 1995).
In melanoma cells, the accumulation of reactive dihydroxyindoles after
effective COMT inhibition could selectively damage melanoma cells (Smit
et al., 1994; Smit and Pavel, 1995).
Some tumors, notably pheochromocytomas, contain high amounts of COMT,
particularly MB-COMT. High metanephrine levels in patients with
pheochromocytoma are derived from catecholamines produced and
metabolized within the tumors. Hence, measurement of plasma-free metanephrine concentrations may be important for the rapid diagnosis of
these patients (Eisenhofer et al., 1998).
Human (Keränen et al., 1994), hamster, guinea pig, rabbit, hen,
dog and monkey (Zürcher et al., 1996), and rat (Nissinen et al.,
1992) erythrocytes contain some COMT activity. The activity varies from
species to species, but it is high in rat and quite low in humans.
Erythrocyte COMT offers a convenient way of monitoring the COMT
inhibition in the body during COMT inhibitor therapy. Several human
studies have shown an excellent correlation between the inhibition of
erythrocyte COMT activity and the concentration of a COMT inhibitor in
plasma or the decrease of 3-O-methyl-dopa (3-OMD) levels in
plasma (Keränen et al., 1994; Dingemanse et al., 1995b). Even
human lymphocytes contain a significant amount of COMT, almost half of
which appears to be MB-COMT (Sladek-Chelgren and Weinshilboum, 1981).
G. General Importance of COMT
1. Substrates of COMT.
The physiological substrates of COMT
include L-dopa, catecholamines (dopamine, norepinephrine,
epinephrine), their hydroxylated metabolites, catecholestrogens (Ball
and Knuppen, 1980 2. Quantitative Role of COMT in Metabolism of Catecholamines.
The relative importance of enzymes in metabolizing catecholamines and
the uptake processes of catecholamines have been clarified. Without
exogenous levodopa loading, the high-affinity neuronal reuptake
(uptake1) is an efficient elimination system for
the released catecholamines, being responsible for most of their
elimination both in peripheral tissues and the brain (Kopin, 1985), ascorbic acid, and dihydroxyindolic intermediates
of melanin. Several dietary and medicinal products are also COMT
substrates, such as triphenols and substituted catechols,
dobutamine, isoprenaline, rimiterol, -methyldopa,
benserazide, carbidopa, dihydroxyphenyl serine (Maruyama et al., 1996),
flavonoids, and dihydroxy derivatives of tetrahydroxyisoquinolones. The
general function of COMT is the elimination of biologically active or
toxic catechols and some other hydroxylated metabolites. During the
first trimester of pregnancy, COMT protects the placenta and the
developing embryo from activated hydroxylated compounds formed from
aryl hydrocarbons by hydroxylases (Barnea and Avigdor, 1990). COMT also
acts as an enzymatic detoxicating barrier between the blood and other
tissues shielding against the detrimental effects of xenobiotics (e.g.,
in the intestinal mucosa and the brain). COMT may also serve some
unique or indirect functions in the kidney and intestine tract by
modulating the dopaminergic tone; the same may be true in the brain:
COMT activity may regulate the amounts of active dopamine and
norepinephrine in various parts of the brain and therefore be
associated with mood and other mental processes.;
Männistö et al., 1992b; Cass et al., 1993). The role of the
extraneuronal transport is less clear (Friedgen et al., 1996b). The
contribution of metabolism, including COMT, is unimportant, and
therefore COMT inhibition does not affect dopamine levels to a
detectable degree. It is equally clear that inhibition of MAO (by
pargyline) and COMT (by tolcapone), each separately, has little, if
any, effect on the removal of norepinephrine, epinephrine, and dopamine
on passage through the systemic and pulmonary circulation. The
pulmonary, but not the systemic, clearance of catecholamine can be
reduced by a combined blockade of MAO and COMT (Friedgen et al.,
1996a). Thus, inhibition of COMT does not greatly alter the elimination of infused catecholamines or exercise-induced elevation of
catecholamines, which is an important safety-increasing factor in the
use of potent COMT inhibitors (see later). However, the elimination
pathways are adaptable. When MAO is blocked, both COMT and
phenolsulfotransferase activities are increased, but these two pathways
do not compete with each other (Buu, 1985).; Messiha et al., 1972; Fahn, 1974; Da Prada
et al., 1984; Männistö et al., 1992b). During the
combination therapy, 3-OMD is the major metabolite (Fahn, 1974;
Rivera-Calimlim et al., 1977; Reilly et al., 1980; Da Prada et al.,
1984; Hardie et al., 1986), and the role of COMT inhibitors becomes
extremely meaningful.
). COMT inhibition by tropolone or a nitrocatechol-type
inhibitor, Ro 41-0960, augmented by 4-fold the transport of ligands
that were COMT substrates but did not affect substrates that were not metabolized by COMT. The uptake of serotonin was not altered by COMT
inhibitors; however, no such potentiation was seen with dopamine uptake
into mouse neostriatal synaptosomes. That may be due to the fact that
dopamine is protected by intracellular sequestration into synaptic
vesicles, organelles that were lacking from the cell lines used in cell culture.
3. COMT Knockout Mice.
The ultimate importance of COMT will
probably be clarified in a new strain of animals lacking the COMT gene.
Such mice were recently produced by Gogos et al. (1998). The mice
appeared to be normal, they had only minor changes in their behavior,
and the brain neurochemistry of catecholamines was virtually unaltered. The mice were also able to breed normally. Surprisingly, all of the
changes were sex and region specific. Mutant male mice, totally lacking
COMT activity, had an almost 3-fold increase of dopamine levels in the
frontal cortex but not in the striatum or hypothalamus, but females
showed no changes. It is noteworthy that despite the complete lack of
COMT gene and protein, residual homovanillic acid (HVA) levels were
detectable in several brain areas. This points to the possibility that
there exists a still-unidentified methylation pathway in the brain.
Female knockout mice had impaired emotional reactivity in the
black-white box. On the other hand, heterozygous males (but not
homozygous males or any females) were more aggressive than their
wild-type counterparts. These findings suggest that 1) the importance
of COMT in the behavior has probably been underestimated and 2) a
complete lack of COMT can be effectively compensated, at least in mice.
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III. COMT Inhibitors |
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A. First-Generation COMT Inhibitors
In 1975, Guldberg and Marsden reviewed early COMT inhibitors.
Several of these compounds (e.g., gallates, tropolone, U-0521, 3',4'-dihydroxy-2-methyl-propiophenone) have been used as in vitro tools; however, their efficacy in vivo is low, and they are short acting. Moreover, they lack selectivity and are rather toxic. For
instance, U-0521 depressed the function of the rat vas deferens at 1 µM and above, which apparently invalidates its use as a COMT inhibitor in smooth muscle preparations (Rice et al., 1997). Also, the
limited clinical experiences with gallates, tropolone, ascorbic acid,
and U-0521 were disappointing (Ericsson, 1971; Reilly et al., 1983;
Reches and Fahn, 1984). It is worth noting that millimolar concentrations of ascorbic acid may well reduce COMT activity significantly, as was recently shown in primate spinal meninges (Kern
and Bernards, 1997).
B. Second-Generation COMT Inhibitors
Three laboratories independently developed very potent, highly
selective, and orally active COMT inhibitors. Nitrocatechol is the key
structure in most of these molecules (Fig.
2). Only CGP 28014 is a chemically
distinct compound (Bäckström et al., 1989; Borgulya et al.,
1989; Waldmeier et al., 1990a).
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In principle, the new COMT inhibitors can be divided into three groups:
1) mainly peripherally acting compounds, 2) broad-spectrum compounds
working in both the periphery and the brain, and 3) atypical compounds,
probably acting preferably in the brain. This classification was
derived from our comparative studies (Männistö et al.,
1992a; Törnwall and Männistö, 1993), where the
decrease in the plasma and brain 3-OMD was used as a marker for
peripheral COMT inhibition. The decline of the brain HVA [and
3-methoxytyramine (3-MT) after pargyline treatment] was used as a
signal of COMT inhibition in the brain.
Because the amount of COMT is high in the liver, kidney, and intestinal
mucosa (Sharpless et al., 1973; Nissinen et al., 1988b), peripherally
acting COMT inhibitors would be primarily interesting. The clearest
clinical application of these COMT inhibitors would be as adjuncts to
levodopa in PD (Männistö and Kaakkola, 1989, 1990).
L-Dopa is a catechol, and it is predominantly
O-methylated to 3-OMD, especially when the peripheral
decarboxylation of L-dopa is inhibited by
benserazide or carbidopa (Nutt and Fellman, 1984). Although the
elevation in 3-OMD levels was not harmful as such, peripheral COMT
inhibition should enhance the brain penetration of
L-dopa. Those COMT inhibitors that gain access to
the brain would provide additional information about the role of
metabolism to potentiate brain dopaminergic and adrenergic functions.
C. Properties of New Compounds
1. COMT Inhibition.
Nitrocatechols are so-called tight-binding
inhibitors of COMT, although their binding to COMT is fully reversible
(Schultz and Nissinen, 1989; Lotta et al., 1995; Borges et al., 1997). Prolongation of the preincubation time with an inhibitor markedly reduces the IC50 values of nitrocatechol-type
inhibitors (Schultz and Nissinen, 1989). This particular property has
been the source of many problems when the kinetic parameters have been
characterized. It is necessary to perform inhibition kinetic studies in
a special way to avoid invalid conclusions. In the presence of varying
inhibitor concentrations, the reaction velocity of COMT increases
progressively with increasing enzyme concentrations and parallels the
velocity curve without an inhibitor if sufficiently high amounts of
enzyme are used. Also, the effect of the nitrocatechol-type inhibitors can be gradually abolished by dialysis (about 50% in 5 h, fully reversed within 24 h; Schultz and Nissinen, 1989).; Nissinen et al., 1992). When analyzed with pure recombinant COMT enzyme forms, Ki
values of nitecapone and entacapone were around 1 nM or even lower
(Lotta et al., 1995). They are also selective because their in vitro
IC50 values for tyrosine hydroxylase,
dopamine--hydroxylase, DDC, and MAO-A and -B are in the micromolar
range (Männistö et al., 1988; Nissinen et al., 1988a,
1992). They strongly inhibit the COMT activity in a variety of tissues.
An oral dose of 10 mg/kg nitecapone or entacapone causes nearly
complete inhibition of duodenal COMT activity for 1 to 3 h and
highly significant inhibition in erythrocyte and liver COMT activity
for several hours. Full recovery of duodenal COMT activity is achieved
at 8 to 12 h after drug administration. The striatal COMT activity
is slightly and transiently suppressed by entacapone administration but
not at all by nitecapone administration; full recovery is attained at
3 h. Oral ID50 values of nitecapone and
entacapone have been in a range of about 1 to 5 mg/kg in the liver and
duodenum but at least 25 mg/kg or higher in the brain (Schultz and
Nissinen, 1989; Zürcher et al., 1990a; Nissinen et al., 1992).
). It seems to be slightly more potent at inhibition of
liver COMT activity than entacapone and nitecapone, with
IC50 and Ki
values of 36 nM in the rat liver (Zürcher et al., 1990a, 1991;
Borgulya et al., 1991). In both recombinant COMT forms, the
Ki value of tolcapone was about 0.3 nM
(Lotta et al., 1995). Tolcapone can penetrate into the brain and
inhibits the brain COMT activity in vivo with an
ID50 value of 26 to 28 mg/kg (Da Prada et al.,
1991; Zürcher et al., 1991). A high oral dose of tolcapone (100 mg/kg) causes virtually complete inhibition of rat heart and kidney
COMT activity at 15 to 30 min. Liver and brain COMT activity is less
effectively suppressed (maximally by 70% at 30 min). About 50%
inhibition is measured at 11 h (heart and kidney), at 8 h
(liver), and at 6 h (brain) after dosing. Full recovery of COMT
activity occurs by 16 h. The inhibition of COMT activity by
tolcapone is fully reversible, as it is with entacapone and nitecapone.
Tolcapone does not affect MAO,
hydroxyindole-O-methyltransferase, histamine-N-methyltransferase, or
phenyl-ethanolamine-N-methyltransferase activities or
adrenergic (, ), serotonergic, or cholinergic receptors
(Zürcher et al., 1990a,b).
Nitrocatechols with NO2 group in positions other
than that in the "classic" nitrocatechols have also been described
(Fig. 2; Perez et al., 1992, 1993, 1994). These dihydroxyvinyl-type compounds are able to bind to the active site of COMT (Vidgren and
Ovaska, 1997), but their efficacy against both human and pig enzymes is
generally lower than that of compounds with the
NO2 in the classic site of the catechol ring. The
situation is complicated by the fact that the in vitro comparisons have
usually been made using pig liver COMT, which differs from rat and
human enzymes by having a polar arginine in position 38 instead of
hydrophobic Trp38. This difference leads to reduced affinity (higher
Km value for catechol substrates) and
higher Ki values for inhibitors. However, even in human COMT, 2-(3,4-dihydroxy-2-nitrophenyl)vinyl phenylketone (vinylphenylketone, or QO IIR) has 4- to 13-fold higher Ki values than nitecapone,
entacapone, and tolcapone (Vidgren and Ovaska, 1997). Vinylphenylketone
is a tight-binding, reversible COMT inhibitor that resembles the
classic nitrocatechols (Perez et al., 1993, 1994).
Also an endogenous COMT inhibitor, 6-nitronorepinephrine (Fig. 2) has
been described and quantified in pig and rat brain, where its
concentration is around 75 pg/g (Shintani et al., 1996). This compound
is generated from the reaction between nitric oxide and norepinephrine.
If the synthesis of nitric oxide is inhibited, the amount of
6-nitronorepinephrine is decreased. Perfusion of 6-nitronorepinephrine
into the paraventricular nucleus elevated norepinephrine levels and
decreased 3-methoxy-4-hydroxyphenyl glycol (MHPG) levels. In vitro,
COMT inhibition is modest because the IC50 value
is as high as 7.5 µM. However, 6-nitronorepinephrine inhibits also
the reuptake of norepinephrine into the synaptosomes, with an
IC50 value of 31 µM. In summary,
6-nitronorepinephrine is a potential, although fairly weak, endogenous
signal molecule that may link the actions of catecholamines and nitric
oxide (Shintani et al., 1996). However, 6-nitronorepinephrine cannot be
used as a drug.
CGP 28014, a hydroxypyridine compound (Fig. 2), was first described by
Waldmeier et al. (1990a,b). CGP 28014 and its major metabolite,
2-amino-6-hydroxypyridine, are not COMT inhibitors in vitro until
millimolar concentrations are reached. CGP 28014 does not affect
receptors of the various neurotransmitters and other endogenous
substances (Waldmeier et al., 1990a).
An active iron chelator, 1,2-dimethyl-3-hydroxypyridine-4-one (L1,
CP20), is a further example of another type of COMT inhibitor that,
however, also inhibits tyrosine and tryptophan hydroxylases (Waldmeier
et al., 1993).
2. Effects on L-Dopa and Catecholamine Metabolism.
In rats, oral administration of entacapone and nitecapone (3-30 mg/kg)
in combination with levodopa and carbidopa effectively reduce 3-OMD
formation and elevate serum and brain L-dopa, dopamine, DOPAC, and HVA levels (Männistö et al., 1988, 1992a;
Nissinen et al., 1988a, 1992). Similar findings have been reported in
Cynomolgus monkeys in whom the i.v. efficacy of entacapone
and nitecapone was equipotent (Cedarbaum et al., 1991). Part of the
levodopa saved from COMT is metabolized through alternative pathways
because the increase of serum L-dopa is smaller
than the decrease in 3-OMD would lead one to expect.
)]. It was
found that carbidopa had an L-dopa-sparing effect in both plasma and muscle, and this effect was further enhanced with
entacapone. The T1/2
of
L-dopa was prolonged from 0.76 to 2.66 h by
entacapone, whereas it was only 0.4 h without carbidopa and entacapone. A similar shift occurred in the muscle although to a lesser
extent. The T1/2
of
L-dopa in plasma was not altered by either
treatment. Formation of 3-OMD was greatly reduced in both plasma (by
98%) and muscle (by 85%) with entacapone. Skeletal muscle may be an
important L-dopa storage site (Deleu et al., 1995) whose capacity can be significantly increased with entacapone (Ordonez et al., 1974).
The effective decrease in 3-OMD, HVA, and 3-MT levels, as well as in
COMT activity in the brain, can be used as criteria for the
classification of tolcapone as a brain-penetrating COMT inhibitor (Zürcher et al., 1990a,b, 1991; Männistö et al.,
1992a). In the time course studies, 13 mg/kg tolcapone p.o. decreased
several 3-O-methylated metabolites of catecholamines (3-MT,
HVA, and MHPG). The formation of 3-MT was suppressed by 90% as early
as 1 h after dosing, and that of HVA and MHPG was suppressed more
slowly and modestly (80 and 60%, respectively) at about 4 h after
tolcapone. Whole brain dopamine was not changed, whereas DOPAC levels
were doubled for 6 to 8 h (Zürcher et al., 1991).
After the oral administration of 30 mg/kg tolcapone with 10 mg/kg
levodopa and 15 mg/kg benserazide, plasma and whole brain L-dopa levels increased by 4-fold and 3-OMD decreased to
very low levels in rats. Dopamine formation in the whole brain
increased for at least 6 h (Zürcher et al., 1990a,b).
In a further study on rats, 20 mg/kg levodopa (combined with 15 mg/kg
benserazide) has been compared with 10 mg/kg levodopa (and 15 mg/kg
benserazide) plus 30 mg/kg tolcapone. The triple treatment
approximately doubled the area under the curve (AUC) of plasma
L-dopa compared with the conventional double treatment with
twice as high a dose of levodopa. Thus, the bioavailability of levodopa
was increased 3.5-fold by the addition of tolcapone (Zürcher et
al., 1991). However, it is worth noting that the T1/2 of levodopa was not markedly
prolonged. The plasma levels of 3-OMD, which normally exceed the plasma
L-dopa levels by a factor of 2, remained low and
near the detection limit for at least 8 h (Zürcher et al.,
1991).
In rats, tolcapone (30 mg/kg i.p.) was also able to prolong the
elimination half-life (+116%) and area under the plasma apomorphine concentration-time curve (+31%), and apomorphine was present in striatum for a 85% longer time period (Coudoré et al., 1997). Even 2-(3,4-dihydroxy-2-nitrophenyl)vinyl phenylketone (QO IIR, or
vinylphenylketone) has recently been proved to act as a peripherally acting COMT inhibitor in in vivo studies in rats (Rivas et al., 1999).
The in vivo effects of CGP 28014 in rats mimicked those of the other
COMT inhibitors. The ED50 values are 2 to 8 mg/kg
p.o. when the endpoints were the decrease in striatal HVA, the decline of striatal 3-MT after clorgyline treatment (both without exogenous levodopa), or the formation of 3-OMD from the exogenous levodopa (without a DDC inhibitor). It was long acting when administered at high
doses of 100 mg/kg (>12 h) or 300 mg/kg (>24 but <36 h). CGP 28014 increased striatal AdoMet levels. However, it unexpectedly increased
striatal 5-hydroxyindoleacetic acid and tryptophan levels (Waldmeier et
al., 1990a,b).
In our studies in rats, CGP 28014 proved to be a poor COMT inhibitor in
the periphery, or at least its inhibitory effect on 3-OMD formation was
slow, becoming significant only by 3 h. However, it behaved as an
efficient COMT inhibitor-like compound in the brain, preventing both
HVA and 3-MT formation (Männistö et al., 1992a). This kind
of brain priority has not been described previously for the COMT
inhibitors. In fact, in earlier studies, CGP 28014 inhibited both 3-OMD
and HVA formation to an equal degree after both p.o. and i.p.
administration (Waldmeier et al., 1990a,b). However, in these studies,
either CGP 28014 was administered without levodopa or levodopa was
administered without the inhibition of the peripheral DDC. Therefore,
only moderate amounts of 3-OMD were produced in the periphery because
most of the L-dopa was decarboxylated to dopamine. This
unusual behavior of CGP 28014 was also seen in rat kidney studies,
where it enhanced dopamine and DOPAC secretion but did not affect
sodium secretion. This is in contrast to the effect of entacapone,
whose action on dopamine secretion was much less but whose action on
sodium secretion was much stronger (Hansell et al., 1998).
3. Microdialysis Studies.
In vivo microdialysis technique is a
widely used method for the estimation of extracellular levels of
various substances, such as neurotransmitters and their metabolites.
The effects of entacapone, tolcapone, and CGP 28014 alone or in
combination with levodopa and DDC inhibitors on brain
L-dopa and dopamine metabolism have been investigated in
rats. The aims have been, on one hand, to clarify the mechanisms of
brain dopamine metabolism and, on the other hand, to confirm the
beneficial effect of combination of a COMT inhibitor with levodopa on
brain dopamine formation. Only a few studies have dealt with
norepinephrine levels and metabolism (Li et al., 1998).
). Contrary to several other findings in
rats, Brannan et al. (1997) reported that 30 mg/kg entacapone had clear
central effects, reducing the brain COMT activity at 2 h. Even at
a dose as low as 10 mg/kg, brain COMT activity was reduced by 80%, and
the only "peripherally selective" doses were 2.5 and 5 mg/kg. The
brain-penetrating reference compound OR-486 (dinitrocatechol; Nissinen
et al., 1988a) had very similar effects. Central effects of entacapone
were seen with levodopa- and carbidopa-treated rats as well (Brannan et al., 1997). We cannot explain these results, which are in conflict with
the general view of other groups (Männistö et al., 1992a; Zürcher et al., 1993; Törnwall et al., 1994). Entacapone
did not modify the effects of clorgyline, selegiline, and nomifensine on striatal dopamine metabolism in the rat brain (Kaakkola and Wurtman,
1992).
Tolcapone, at doses of 3 to 30 mg/kg i.p. or p.o., dose-dependently
decreased the efflux of HVA and increased levels of L-dopa and DOPAC but did not affect the output of dopamine in the rat striatum
(Acquas et al., 1992; Kaakkola and Wurtman, 1993; Napolitano et al.,
1995b). In addition, the efflux of 3-MT declined rapidly after
tolcapone (40 mg/kg s.c.; Cumming et al., 1992). Tolcapone potentiated
the effect of nomifensine on rat striatal dopamine efflux, whereas it
did not significantly modify the effects of MAO inhibitors (clorgyline,
pargyline, and selegiline; Cumming et al., 1992; Kaakkola and Wurtman,
1993). We did not find in the rat striatum any potentiation by
tolcapone on dopamine levels that had been elevated by pargyline or
amphetamine (Tuomainen et al., 1996). Neither entacapone nor tolcapone
altered the extracellular levels of 5-hydroxyindoleacetic acid in any
of the above studies.
Li et al. (1998) thoroughly studied the effect of tolcapone on the
extracellular levels of dopamine and norepinephrine and their
metabolites in a number of brain areas in anaesthetized rats. Although
HVA and MHPG levels were clearly reduced, no increases were seen in
dopamine in striatum or nucleus accumbens or norepinephrine in nucleus
accumbens, frontal cortex, or two parts of the hippocampus.
Steulet et al. (1993) reported that CGP 28014 (30 mg/kg i.p.) did not
significantly change the output of dopamine and DOPAC from rat
striatum, whereas Törnwall et al. (1994) found a slight increase
in the efflux of dopamine and DOPAC. Both groups reported a significant
reduction in the output of HVA. However, when CGP 28014 was
administered intrastriatally, it induced dose-dependent and severalfold
elevation of extracellular dopamine levels but unexpectedly did not
alter HVA levels. This tyramine-like action was abolished by
nomifensine, indicating that CGP 28014 must be transported into nerve
terminals via the dopamine uptake system (Steulet et al., 1993). These
findings suggest that CGP 28014 must undergo peripheral metabolism
before it can inhibit COMT. No active metabolite has been identified,
and therefore other mechanisms of action should be considered
(Männistö et al., 1992b; Steulet et al., 1993; Waldmeier et
al., 1993).
The microdialysis data show that entacapone is principally a
peripherally active COMT inhibitor, whereas tolcapone and CGP 28014 have major affects on the O-methylation of brain dopamine. These studies have also confirmed that MAO plays a more important role
than COMT in the metabolism of extracellular dopamine. Reuptake (uptake1) is even more important than either of
the metabolizing enzymes in termination of the actions of
catecholamines in the synapse (see Quantitative Role of COMT in
Metabolism of Catecholamines).
b. Drugs Given with Levodopa. When combined with levodopa
and DDC inhibitors, both entacapone and tolcapone caused a significant supplementation of the extracellular levels of L-dopa,
dopamine, and DOPAC in the rat striatum (Acquas et al., 1992; Brannan
et al., 1992; Kaakkola and Wurtman, 1993; Napolitano et al., 1995a,b). The levodopa-induced increase in HVA output was attenuated by tolcapone
but not by entacapone (Törnwall et al., 1994). Both entacapone
and tolcapone reduced the levels of 3-OMD (Törnwall et al., 1994;
Napolitano et al., 1995a,b).
As described, the effect of entacapone has also been studied in rat
skeletal muscle where both carbidopa (100 mg/kg/day p.o.) and
entacapone (15 mg/kg i.v.) had distinct levodopa-sparing effects, and
when given together, this effect was further enhanced (Deleu et al.,
1995).
In conclusion, combination studies with levodopa- and
nitrocatechol-type COMT inhibitors have demonstrated in rats that the inhibition of COMT activity results in increased dopamine formation and
release in brain after the administration of levodopa and DDC
inhibitors. A significant effect can be achieved even if the COMT
inhibition is restricted to peripheral tissues.
When combined with levodopa and carbidopa, CGP 28014 did not elevate
dopamine levels in the microdialysis fluid of the rats. It transiently
decreased HVA formation but did not alter 3-OMD levels (Törnwall
et al., 1994), confirming our results with brain homogenates
(Männistö et al., 1992a).
4. Voltammetric Studies.
Garris and Wightman (1995) studied
the effect of various drugs, including tolcapone (40 mg/kg), on the
dopamine efflux from the rat caudate-putamen and basolateral amygdaloid
nucleus, elicited by electrical stimulation of ascending dopamine
fibers at various frequencies. Dopamine efflux was monitored by
fast-scan cyclic voltammetry. Tolcapone caused negligible effects in
both regions at frequencies of 30 Hz or higher. However, at 20 Hz,
these workers did detect a nearly 50% increase in dopamine release. In
contrast, the effects of two uptake blockers, nomifensine and cocaine,
to stimulate dopamine efflux were robust in both brain regions. Similar results were recently obtained by Budygin et al. (1999) with 30 mg/kg
tolcapone and 10 mg/kg GBR 12909, a new dopamine uptake blocker. These
results suggest that under normal conditions,
uptake1, rather than transmitter metabolism,
regulates extracellular levels of dopamine.
5. Estrogen Metabolism and Role of COMT and COMT
Inhibitors.
Estrogen metabolism, including the crucial role of
COMT in the metabolism of 2- and 4-hydroxylated estrogens
(catecholestrogens), was reviewed by Ball and Knuppen (1980) and
recently by Zhu and Conney (1998). Some of the highlights and safety
aspects are described here. A schematic summary of the main pathways,
and of the role of COMT in particular, is given in Figs.
3 and
4.
|
|
-estradiol), which is reversibly oxidized to estrone,
can undergo numerous metabolic routes. Among these, the NADPH-dependent
hydroxylation reactions, catalyzed mainly by multiple forms of
cytochrome P-450 enzymes, are relevant to this review. There are many
hydroxylation products that are formed, but only the catecholestrogens
are substrates of COMT, and at high concentrations they may act as
competitive COMT inhibitors. Catecholestrogens and their
O-methylated products, 2- and 4-methoxy estradiols or 2- and
4-methoxy estrones, are not inert metabolites but may possess unique
activities that are not necessarily directly associated with the
actions of their parent hormones (Zhu and Conney, 1998).
It is difficult to estimate the relative amounts of various
hydroxylated estrogens, but in women, 2-OH-estradiol, which is formed
mainly in the liver, is the most abundant catecholestrogen detected in
urine (50% or more; Dannan et al., 1986; Kerlan et al., 1992; Zhu et
al., 1993; Suchar et al., 1995). The other hydroxylated estrogens are
much less abundant; 4-OH-estradiol represents about 5% (<15% of that
of 2-OH-estradiol) and the 16 -hydroxylated product represents more
than 10% of the total. It is noteworthy that in extrahepatic
"target" tissues, 4-hydroxylation is much more common (see below).
There are very low levels of 2-OH-estrogens in the systemic
circulation, mainly because they are rapidly further metabolized by
COMT and conjugation enzymes (sulfation, glucuronidation; Ball et al.,
1978; Fishman and Martucci, 1979; Emons et al., 1983, 1987; Zhu and
Liehr, 1996). In addition to liver, 2-OH-estrone and 2-OH-estradiol are
also formed locally in target tissues like uterus and breast.
2-OH-estrogens have multiple effects in the body, only a few of which
are relevant here. First, they bind to estrogen receptor but only with
low affinity, and their hormonal potency is much less than that of
their parent compounds. This means that their net effect is
antiestrogenic (MacLusky et al., 1983; van Aswegen et al., 1989;
Feigelson and Henderson, 1996). Second, they may generate estrogen
semiquinones and quinones and highly toxic oxygen radicals (Liehr et
al., 1986b; Liehr, 1990; Liehr and Roy, 1990). The latter possibility
seems rather theoretical because instead of having tumorigenic activity
in routine tests, they tend to inhibit spontaneous tumorigenesis in
estrogen-sensitive tissues (Liehr et al., 1986a; Li and Li, 1987; see
below). Even a subsequent O-methylation product of
2-OH-estradiol, 2-methoxy estradiol, inhibits angiogenesis, and is a
strong inhibitor of tumor cell proliferation (Seegers et al., 1989;
Lottering et al., 1992; D`Amato et al., 1994; Fotsis et al., 1994;
Hamel et al., 1996; Klauber et al., 1997). Interestingly, in patients
with depression, the formation of 2-methoxyestrogens is increased,
whereas that of 4-hydroxylation is reduced (Banger et al., 1990). In
summary, 2-OH-estrogens and their O-methylation products are
evidently beneficial and anticarcinogenic (Fig. 3).
4-Hydroxylation of estrogens is not extensive in the liver, but this
reaction is much more common in the extrahepatic target tissues of
estrogens, such as pituitary (Bui and Weisz, 1988), myometrium and
myomal tissues (Lier et al., 1995), and breast (Lier et al., 1995;
Hayes et al., 1996). This point has been thoroughly discussed by Weisz
(1991, 1994) and recently by Weisz et al. (1998a,b) and Cavalieri's
group (Cavalieri et al., 1997; Cavalieri and Rogan, 1998; Stack et al.,
1998). The actions of 4-OH-estrogens are completely different from
those of 2-OH-derivatives. First, 4-OH-estradiol binds to estrogen
receptors as effectively as estradiol itself, and its binding is long
lasting (Martucci and Fishman, 1976; MacLusky et al., 1983; van Aswegen
et al., 1989). Consequently, 4-OH-estrogens are hormonally very active
(Martucci and Fishman, 1976; Franks et al., 1982). Second,
4-OH-estradiol undergoes a metabolic redox cycling to generate free
radicals and chemically very reactive semiquinone and quinone
intermediates, which can damage DNA and other cellular components,
induce cell transformation, and initiate tumorigenesis (Liehr et al.,
1986b; Liehr, 1990; Liehr and Roy, 1990; Fig. 4). Third, 4-OH-estradiol
is a strong carcinogen in the hamster kidney. Injections of either
4-OH-estradiol or estrone-3,4-quinone cause kidney and liver tumors but
not mammary tumors (Liehr et al., 1986a; Li and Li, 1987).
The above quinone hypothesis, bypassing the importance of pure estrogen
receptor stimulation as a key factor in estrogen-dependent cancer, has
obtained strong support from recent studies on the abundant appearance
of cytochrome P-4501B1, converting estrogens to 4-OH-estrogens in the
breast tissue, and apparently also in uterus and ovaries, more
effectively than in tissues not prone to estrogen-linked cancer (Hayes
et al., 1996; Spink et al., 1998). Catecholestrogens can be metabolized
further by COMT, conjugated to glucuronide or sulfate but also oxidated
to semiquinones and quinones. Particularly reactive is
estrogen-3,4-quinone, which has been shown to bind to DNA, creating
so-called depurinating adducts both in in vitro studies and in animals.
These adducts are quickly falling off, taking with them two bases of
DNA: adenine and guanine. The gaps formed in the DNA have a strong
potential to create gene mutations and eventually cause cancer
(Cavalieri et al., 1997). These events are schematically summarized in
Fig. 4. Because the formation of 4-OH-estrogens is very low, at least in humans, it is quite unclear how large a risk this metabolic route
may have. However, any procedure inhibiting the normal metabolism of
estrogens may increase formation of 4-OH-estrogens and can be regarded
as potentially dangerous.
Interestingly, after 2 or 4 weeks of estrogen treatment, COMT
immunoreactivity was increased in hamster kidneys, and it was seen even
in the nucleus (Weisz et al., 1998b). This is unexpected because in
contrast to human COMT, hamster kidney COMT lacks nuclear localization
signal sequence. The nuclear entry of COMT was also selective because
another second phase II enzyme, CuZn-superoxide dismutase, remained
extranuclear. The authors suggest that the nuclear translocation of
COMT after estrogen treatment is a protective response to the threat
of the genome by electrophilic products of catechols, quinones
and semiquinones (Weisz et al., 1998b). It can be concluded that
4-OH-estradiol is a potentially harmful compound, a strong tumorigenic
substance that may be associated with the genesis of estrogen-dependent
cancers (Figs. 3 and 4).
The role of COMT in the O-methylation of estrogens and how
this is altered by potent COMT inhibitors are areas of concern. Catecholestrogens are generally good substrates of COMT. 2-OH-estrone in particular is rapidly O-methylated, and 2-methoxy estrone
is one of the most abundant estrogen metabolites in human plasma and
urine (Kraychy and Gallagher, 1957; Ball et al., 1979). Both 2-methoxy
estradiol and 2-methoxy estrone have very high affinity for the sex
hormone-binding protein (Dunn, 1983). Little is known about the role of
4-methoxy derivatives, but their activity is inferior to that of the
corresponding 2-methoxy derivatives.
Mono-O-methylated estrogens have little or no affinity for
estrogen receptors, and they do not exert any estrogenic effect on
target tissues (Merriam et al., 1980). It seems, therefore, that
O-methylation of catecholestrogens is primarily a
detoxification pathway. However, 2-methoxy estradiol has a number of
effects on its own, not related to estradiol, 2-OH-estradiol or other methoxy derivatives. It inhibits the proliferation of several cancer
cell lines in vitro (Seegers et al., 1989; Lottering et al., 1992;
Fotsis et al., 1994; Cushman et al., 1995; Klauber et al., 1997). For
example, human breast cancer cell lines were particularly sensitive to
2-OH-estradiol, and it depressed the growth of some implanted cancers
(Fotsis et al., 1994; Klauber et al., 1997). 2-Methoxy estradiol also
disturbs the function of microtubules (D'Amato et al., 1994; Hamel et
al., 1996), and it is one of the most potent endogenous inhibitors of
angiogenesis known (Fotsis et al., 1994; Klauber et al., 1997). Even
the inhibitory effects of some enzyme inducers, like phenobarbital and
indole-3-carbinol, on spontaneous breast carcinogenesis in mice may be
mediated by enhanced formation of 2-OH-estradiol and its rapid
metabolism to 2-methoxy estradiol (Osborne et al., 1990; Bradlow et
al., 1991; Jellinck et al., 1991, 1993).
The 16 -hydroxylation of estrogens does not have any direct
connection to COMT. However, this route may be activated if the COMT
pathway is inhibited, and therefore the properties of 16 -OH-estrogens are important (Fig. 3). Both 16 -OH-estrone and 16 -OH-estradiol retain most of the hormonal activity of their parent
compounds (Fishman and Martucci, 1980). 16 -OH-estrone seems to bind
covalently to the estrogen receptor and may activate the classic
estrogen-mediated oncogene expression and evoke long-term growth
stimulation (Hsu et al., 1991). It is fairly well established, based on
a number of different experiments, that increased formation of 16 -OH-estrogens is associated with an increased risk of mammary cancer
in mice and humans (Bradlow et al., 1986, 1995; Fishman et al.,
1995).
We want to stress that the above summations have not been confirmed in
other types of experiments and that they are not universally accepted.
Also, other groups have demonstrated that increased urinary excretion
of catecholestrogens, but not 16 -OH-estrogens, can be correlated
with an increased risk for nonfamilial breast cancer (Lemon et
al., 1992). The same outcome was reached in a Finnish study where
catecholestrogens dominated compared with 16 -OH-estrogens in
the population at high risk to develop breast cancer (Adlercreutz et
al., 1994a,b). Finally, 16 -OH-estrone and 16 -OH-estradiol are
quite weak carcinogens in the hamster kidney tumor model (Li and Li,
1987; Seegers et al., 1989; Li et al., 1995). Furthermore, during
pregnancy, women excrete large amounts of 16 -OH-estrogens, but
full-term pregnancy does not increase the risk of breast cancer
(Merrill, 1958; MacMahon et al., 1970; Fotsis, 1987; Yuan et al., 1988;
Henderson et al., 1996). Our conclusion is that there is not
sufficiently strong evidence to conclude that 16 -OH-estrogens are
carcinogenic but also that their harmful effects cannot be excluded. It
seems that they do not possess any beneficial effects on
estrogen-dependent cancers.
The intention of this discussion is to demonstrate that there is a
possibility that inhibition of COMT by new drugs may seriously interfere with the metabolism of catecholestrogens. However, to the
best of our knowledge, no studies have been conducted on this matter.
6. Behavior.
Generally, when administered alone, COMT
inhibitors have virtually no effect on the motor behavior of rodents
(Maj et al., 1990; Männistö et al., 1992b;
Männistö, 1998). However, a compound that interferes with
adrenergic transmission would be predicted to have important
cognition-enhancing effects via improved attention and motivation or
secondarily via enhanced cholinergic functions (Levin et al., 1990;
Kelland et al., 1993). In fact, we have found that COMT inhibitors
affect several phases of learning in a simple passive avoidance
paradigm (Khromova et al., 1997). In more sophisticated studies,
spatial working memory (radial-arm maze) of intact rats was facilitated
after the pretraining i.p. administration of tolcapone (10 mg/kg).
Similarly, tolcapone improved the performance of senescent poor
performers in a spatial memory task (linear arm maze). However,
tolcapone was not able to counteract the performance deficits in rats
whose memory had been impaired by scopolamine or bilateral lesions in
the nucleus basalis magnocellularis (Liljequist et al., 1997).
). In our
studies, however, levodopa also was needed to reveal the
antidepressant-like effect of tolcapone in two other rat models of
depression (Männistö et al., 1995). In addition, it is
quite interesting that entacapone (30 mg/kg i.p., a dose that
apparently penetrates the blood-brain barrier to some extent),
administered with levodopa (100 mg/kg i.p.) induced a place preference
in rats (Katajamäki et al., 1998).
Maj et al. (1990) described behavioral effects of tolcapone. It
increased exploratory activity of rats at doses of 10 and 30 mg/kg
p.o., and the hyperactivity and stereotypy induced by amphetamine and
nomifensine were potentiated by tolcapone. Tolcapone slightly
antagonized fluphenazine-induced catalepsy but did not protect against
pimozide- or haloperidol-induced catalepsy. Also, Himori and Mishima
(1994) found that pretreatment with tolcapone (30 mg/kg) slightly
potentiated the antagonist effect of levodopa and benserazide on
haloperidol-induced catalepsy in the mice administered 1-methyl-4-phenylpyridium intracerebroventricularly. Very high doses of
3-OMD (800 mg/kg) attenuated this effect of tolcapone.
The antiakinetic action of a subthreshold dose of levodopa (5 mg/kg) in
the MPTP-treated mice was potentiated by small doses of tolcapone (1 and 3 mg/kg). The same was true with two MAO inhibitors: pargyline (5 mg/kg) and selegiline (L-deprenyl; 1, 5 and 10 mg/kg). When
given together, tolcapone and pargyline increased both the peak effect
and the duration of the antiakinetic action. In the case of tolcapone
plus L-deprenyl, only the duration of action was prolonged
(Fredriksson and Archer, 1995).
Entacapone potentiated the turning behavior induced by levodopa and
carbidopa in rats with unilateral nigral lesions (Etemadzadeh et al.,
1989). Nitecapone resembled entacapone in this respect (Männistö et al., 1988). Comparative studies have confirmed that inhibition of COMT outside the brain (e.g., after entacapone and
nitecapone) contributed nearly equally to the levodopa-induced turning
behavior as that occurring also within the brain (i.e., after
tolcapone; Törnwall and Männistö, 1993). We were not able to show any positive interaction between entacapone (3 mg/kg) and
clorgyline (3 mg/kg) on the levodopa/carbidopa-induced turning behavior
(Törnwall and Männistö, 1993). Entacapone potentiated the behavioral effects of levodopa also in common marmosets (Smith et
al., 1997).
When given with levodopa and benserazide, tolcapone potentiated the
increase in motor activity plus the antihypothermic effect and
anticataleptic activity of levodopa or levodopa plus pargyline (Maj et
al., 1990). Tolcapone potentiated levodopa/carbidopa-induced turning
behavior at doses of 3 to 30 mg/kg (Törnwall and
Männistö, 1993).
The effects of tolcapone and MAO inhibitors, separately and together,
on levodopa-induced turning were thoroughly studied by Heeringa et al.
(1997). There was a significant potentiation of levodopa-induced (10 mg/kg levodopa, 15 mg/kg benserazide) contralateral turning by
tolcapone (30 mg/kg) or an MAO-A inhibitor, Ro 41-1049, but not with an
MAO-B inhibitor, Ro 19-6327. Pretreatment with tolcapone and either of
the MAO inhibitors further potentiated the levodopa-induced turning.
The blockade of three dopamine-metabolizing enzymes (COMT, MAO-A,
MAO-B) at the same time, leaving only conjugation reactions intact,
most effectively prolonged the turnings.
In addition, a vinylphenylketone-type COMT inhibitor (QO IIR) was able
to potentiate the reversal of reserpine-induced akinesia by
levodopa/carbidopa in rats at a dose range of 7.5 to 30 mg/kg i.p. and
catalepsy and hypothermia in mice at 30 mg/kg i.p., as did the
reference compounds nitecapone and Ro 41-0960 at 30 mg/kg (Rivas et
al., 1999). CGP 28104 was as effective as entacapone and tolcapone in
potentiating the turning behavior induced by levodopa and carbidopa
(Törnwall and Männistö, 1993).
7. S-Adenosyl-L-Methionine-Saving Effect
of COMT Inhibitors.
It is well known that the levodopa therapy in
PD patients depletes their body levels of AdoMet levels, and this
depletion also occurs in the brain (Benson et al., 1993). This can be
explained by enhanced O-methylation of the large doses of
L-dopa present, which consumes the methyl groups
of AdoMet. Long-term levodopa treatment leads to compensatory
enhancement of the synthesis of AdoMet by increasing the activity of
methionine adenosyl transferase (Benson et al., 1993). In turn, when
high doses of exogenous AdoMet were administered, they depleted
nigrostriatal and forebrain tyrosine hydroxylase and brain dopamine
(Charlton, 1997). When the COMT-induced O-methylation was
effectively inhibited by first-generation COMT inhibitors (like
tropolone but not pyrogallol; Waldmeier and Feldtrauer, 1987) and
second-generation compounds (like tolcapone; Da Prada et al., 1994;
Miller et al., 1997; Yassin et al., 1998), AdoMet levels in the brain
were increased. Homocysteine levels were restored by tolcapone (Da
Prada et al., 1994; Miller et al., 1997). Tolcapone enhanced the
activity of methionine adenosyl transferase (Yassin et al., 1998). Even
CGP 28014, an atypical inhibitor of O-methylation that does
not inhibit the COMT enzyme itself, elevated AdoMet levels in the brain
(Waldmeier et al., 1990a). Interestingly, MAO inhibition appeared to
have just the opposite effect as that observed after COMT inhibition
(Yassin et al., 1998).
; Männistö et al., 1995) and cognition-improving (Khromova et al., 1995, 1997)
effects of COMT inhibitors. Some studies have shown that the quality of
life is increased in PD patients receiving COMT inhibitor therapy
(Welsh et al., 1995; Baas et al., 1997; Waters et al., 1997). One
report has also described improved cognition during tolcapone therapy
(Gasparini et al., 1997). In fact, these findings fit well to the
animal data demonstrating both antidepressive and cognition-improving
properties of COMT inhibitors (Moreau et al., 1994;
Männistö et al., 1995; Khromova et al., 1997; Liljequist et
al., 1997).
8. Other Effects of COMT Inhibitors.
The nitrocatechols, of
which nitecapone and entacapone have been studied in detail, are quite
effective antioxidants (Suzuki et al., 1992), nitric oxide radical
scavengers (Marcocci et al., 1994), and iron chelators (Haramaki et
al., 1995; Orama et al., 1997), and they can protect cells from lipid
peroxidation (Haramaki et al., 1995). Nitecapone has also been shown to
prevent ischemia-reperfusion injury in experimental heart surgery in
rats (Haramaki et al., 1995). In all these studies, the
concentrations used (high micromolar or even millimolar) were
very much higher than those needed to inhibit COMT. These actions do
not appear to be related to COMT inhibition.
). It is interesting that tolcapone (at 1 or 100 nM), like
selegiline (at 1 nM to 10 µM), can protect aggregation cultures of
rat brain exposed to a combination of anoxia and hypoglycemia for 30 min. Tolcapone (10 µM) was itself toxic. The beneficial effect was
seen after only 1 day in culture, not at 2- or 4-day after ischemia
(Ekblom et al., 1998).
Nitecapone increased bicarbonate secretion from rat and human duodenum
after both i.v. and intraluminal administration (Flemström et
al., 1993; Knutson et al., 1993; Flemström and Säfsten,
1994). High concentrations of nitecapone (maximum effect at 225 µM)
increased synthesis and secretion of gastric sulfomucin (Slomiany et
al., 1993). Although these findings point to some gastroprotective properties, nitecapone has not been found to be of any use as an
antiulcer therapy in humans.
The mechanism of tolcapone-induced dose-dependent diarrhea has been
studied in conscious dogs bearing 40-cm jejunal segment loops. Buffered
physiological solution was perfused, and the secretions were analyzed
at 15-min intervals. Acute oral administration of tolcapone (10 or 30 mg/kg) was well tolerated but induced a dose-dependent efflux of
intestinal fluid, sodium, potassium, chloride, and bicarbonate (Larsen
et al., 1998). A similar enhancement of bicarbonate excretion has been
seen in gastric mucosa studies (Flemström et al., 1993; Knutson
et al., 1993; Flemström and Säfsten, 1994). The
coadministration of levodopa and benserazide accelerated the onset of
tolcapone-induced secretion. No diarrhea developed in animals in these
studies, nor did the treatments cause any signs of mucosal damage. The administration of tolcapone or the drug combinations for 7 days reduced
secretory responses, demonstrating that tolerance did develop (Larsen
et al., 1998). The importance of these findings to the pathogenesis of
diarrhea remained open, but it cannot be excluded that the increased
secretion may induce diarrhea in some susceptible patients. It is
somewhat puzzling that dopamine itself has been reported to have an
antisecretory effect in the rat jejunum preparation (stripped
epithelial sheets). This action was mediated through
2-adrenergic receptors (Vieira-Coelho and
Soares-da-Silva, 1998).
Nitrocatechols may inhibit oxidative phosphorylation and uncouple
mitochondrial energy production. One study compared entacapone and
tolcapone; it was reported that tolcapone was severalfold more
effective as an uncoupler than entacapone. The effective concentrations
of tolcapone were in the same range as those achieved in plasma during
tolcapone therapy (Nissinen et al., 1997). The relevance of this
finding to the adverse effects of tolcapone (e.g., to the hepatic
failure, rhabdomyolysis, and diarrhea) remains to be clarified.
9. Physicochemical Properties and Animal
Pharmacokinetics.
Although the COMT-inhibiting
Ki and IC50
values of nitrocatechols are in the low nanomolar range in vitro, doses
as high as 10,000 nM/kg are needed to achieve adequate COMT inhibition
in vivo. Evidently, these compounds have poor general intracellular availability. There are very little published data on the
physicochemical properties of the new COMT inhibitors. Nitrocatechols
are weak acids, and the pKa values of
both entacapone and tolcapone are 4.5. The water solubility of both
compounds is low, less than 0.005% (<0.05 µg/ml); it is
particularly poor in acidic milieu and improved considerably at pH 7.4 (entacapone, 7 mg/ml). The solubility in organic solutions and alcohols
is substantially better. The log P (octanol/0.1 N HCl) of
entacapone is 2.01; that of tolcapone is 2.6, indicating the better
lipid solubility of the latter compound (Roche, 1997; P. T. Männistö and S. Kaakkola, unpublished data).
mentions unpublished observations that the brain extraction ratio of
entacapone is 3% and is not dependent on the concentration in plasma.
In contrast, he claims that tolcapone has a concentration-dependent (5-100 µg/l) brain penetration of 24 to 46% but does not provide any experimental details to amplify these statements. These values are
extremely high compared with our unpublished data generated after the
i.v. injection of varying doses of entacapone and tolcapone. The brains
were made blood free by saline perfusion before collection. Our
brain/plasma ratios were from less than 0.1 to 0.27% for entacapone and from 0.6 to 1% for tolcapone during the first 60 min after i.v.
injection of the drugs (P. T. Männistö and S. Kaakkola, unpublished data, 1995). Both experiments agree on one point: the brain penetration of tolcapone is severalfold higher than that of entacapone.
Studies into the pharmacokinetics of tolcapone in rats have been
published (Funaki et al., 1994, 1995). After 10 mg/kg i.v., the
T1/2 is less than 60 min, the AUC is 21.8 µg × h/ml, the total clearance is 7.84 ml/min × kg, and
the VD is 0.291 liters/kg. Oral doses
of 20 and 40 mg/kg led to a slightly longer
T1/2. The maximum drug concentration in
plasma (Cmax) values were 15.9 and 20.2 µg/ml and the AUC values were 21.1 and 49.0 µg × h/ml
after 20 and 40 mg/kg, respectively. It can be calculated that the oral bioavailability of tolcapone in the rat is 48 to 56%.
Data on entacapone rat kinetics in rats have not been published, but in
general, the kinetic values after i.v. administration are close to
those of tolcapone. However, after oral administration, the plasma
entacapone levels are lower than those obtained with tolcapone. The
oral bioavailability appears to be less than 20% in the rat (P. T. Männistö and S. Kaakkola, unpublished data, 1995).
Nitrocatechols are extensively metabolized, and only a small fraction
is recovered intact in the urine. The metabolic profile is described in
more detail in relation to the human pharmacokinetics.
The reason for the low oral bioavailability of nitrocatechols is still
unknown. Their poor water solubility could indicate poor absorption,
but also their abundant metabolism may start during the first pass. It
seems likely that both factors contribute to their incomplete oral bioavailability.
10. Toxicity.
A full preclinical toxicology program has been
conducted with entacapone and tolcapone; both drugs have recently been
marketed throughout the world. A discussion on human toxicology appears in the clinical section of this review (see Safety). The
acute toxicity of nitrocatechols is generally low (Borgulya et al., 1989). The acute i.p. LD50 value of Ro 41-0960, a
structural analog of tolcapone, in mice is about 100 mg/kg, whereas
that of nitecapone and entacapone is about 500 mg/kg (Törnwall
and Männistö, 1991). Levodopa and carbidopa treatments do
not significantly enhance the acute toxicity of COMT inhibitors. Oral
LD50 values for tolcapone are 1600 to 1800 mg/kg
in mice and greater than 2 g/kg in rats (Borgulya et al., 1991). The
same is true for entacapone and nitecapone. Because their effective
doses are in the range of 3 to 30 mg/kg, the therapeutic index for oral
administration in rodents is more than 50. The lethal threshold for
tolcapone in rats and dogs was more than 100 µg/ml (Roche, 1997).
These values can be compared with the approximate therapeutic
plasma levels of 3 to 6 µg/ml attained after 100 and 200 mg of
tolcapone delivered in three doses each day.
). Moderate epithelial
hyperplasia of the nonglandular part of the rodent stomach (not present
in humans) was seen in rats and mice. In the same studies, 35% of rats
developed renal epithelial tumors, depending on the dose. The safety
margin of this phenomenon was about 10-fold above the therapeutic
plasma concentrations of tolcapone. Finally, an increased incidence of
uterine adenocarcinomas was seen at the 250 mg/kg/day dosage level.
This finding may be related to the fact that dopamine agonists produced
uterine tumors that are specific to rats. In addition, the
dopamine-prolactin-estrogen axis may be disturbed. Catecholestrogens
are also very good substrates of COMT, and a blockade of their
metabolism will lead to accumulation (Ball et al., 1972; Ball and
Knuppen, 1980; see Estrogen Metabolism and Role of COMT and COMT
Inhibitors). It is worth noting that the liver was not a target of
tolcapone toxicity in rodents.
We are not aware of the detailed results of the long-term toxicity
studies of entacapone. However, genotoxicity and carcinogenicity studies have not revealed any serious toxicity. Some instances of
anemia has been noted during repeated administration, probably due to
iron chelation by entacapone. In reproduction toxicity studies in
rabbits, there was some retardation of fetal growth and bone
development (Orion, 1998).
11. Conclusions from Animal Studies. The nitrocatechol-type COMT inhibitors alter L-dopa metabolism and potentiate the action of levodopa plus DDC inhibitors much more effectively than the first-generation COMT inhibitors. Some of the new compounds (entacapone, nitecapone) hardly penetrate the blood-brain barrier but still significantly increase striatal dopamine levels and potentiate the behavioral effects of levodopa in the same way as the brain-penetrating compounds (tolcapone, Ro 41-0960).
The mechanism of action of CGP 28014 is not known. Because it or its known metabolite do not directly inhibit COMT, it might inhibit the transfer of the COMT substrates to the enzyme (i.e., uptake2). However, in glioma cells, no such inhibition could be seen (M. Törnwall and P. T. M., unpublished results, 1993). A further possibility would be that CGP 28014 forms an unknown metabolite, and this is not formed in vitro, only in vivo. Nevertheless, CGP 28014 mimics the other COMT inhibitors in being an inhibitor of O-methylation, preferably that acting in the brain. It is also positive in alleviating the symptoms in animal model of PD.| |
IV. Positron Emission Tomography Studies |
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6-[18F]Fluoro-L-dopa (6-FD) is
an analog of L-dopa that is used in PET studies as a tracer
of the presynaptic dopaminergic system. Like L-dopa, it is
decarboxylated by DDC to [18F]dopamine and
O-methylated by COMT to
3-O-methyl-[18F]dopa (3-OMFD). In
routine PET studies with 6-FD in combination with DDC inhibitors,
3-OMFD represents a considerable proportion of the radioactivity in
both plasma and brain (Firnau et al., 1987, 1988). Various kinetic
models have been used to differentiate the specific from nonspecific
activity. Selective COMT inhibition would reduce the formation of
3-OMFD, simplify PET modeling, and improve the quality of PET images.
When given in combination with 6-FD and DDC inhibitors, nitecapone,
entacapone, tolcapone, and CGP 28014 substantially reduce the plasma
levels of 3-OMFD in monkeys (Guttman et al., 1993; Miletich et al.,
1993; Günther et al., 1995; Doudet et al., 1997; Holden et al.,
1997; Psylla et al., 1997). The decreased peripheral metabolism of 6-FD
after COMT inhibition is reflected as an increased striatal uptake of
6-FD and significantly better PET image contrast. They increased the
influx constant Ki(o) (using occipital
counts as input function) by 45% but did not affect the
decarboxylation of 6-FD (Guttman et al., 1993). In comparative studies,
tolcapone was most active in improving the availability of 6-FD,
followed by CGP 28014 and entacapone. CGP 28014 was more active p.o.
than i.v. (Günther et al., 1995; Psylla et al., 1997). Some
studies have also been done in rats (Pauwels et al., 1994).
Hartvig et al. (1992) reported that tolcapone, administered in
combination with [11C]L-dopa or
6-FD but without a DDC inhibitor to monkeys, did not affect the brain
uptake for any of the ligands. Tolcapone did not change the
decarboxylation rate of L-dopa in striatum but increased
that of 6-FD, probably indicating a significant contribution of 3-OMFD
to background activity. Interestingly, the combination of tolcapone
plus benserazide significantly inhibited the central decarboxylation of
[11C]L-dopa, possibly due to an
elevated brain entry of benserazide and its active metabolite (Tedroff
et al., 1991). No such effect was observed with tolcapone plus carbidopa.
Quite a few PET studies with COMT inhibitors have been performed in
humans, all of them with entacapone or nitecapone. There is no doubt
that the quality of PET scans, expressed as a ratio of the activity in
the striatum to that in the occipital lobe or cerebellum, is improved.
This improvement is seen both in normal volunteers and in patients with
PD. With nitecapone, the increase was 22% in both cases (Laihinen et
al., 1992). In entacapone-treated volunteers and PD patients, the
improvements have varied from 2 to 41% (Sawle et al., 1994; Ruottinen
et al., 1995, 1997; Ishikawa et al., 1996). The worst rates were seen
in the most advanced cases of PD, for whom the patients had been
treated with levodopa for years, and the best results occurred in
normal volunteers and in de novo PD patients (Ruottinen et al., 1995,
1997).
To conclude, PET studies with 6-FD have shown that the peripheral COMT inhibitors substantially improve the brain entry of 6-FD; it is likely that such an effect is also obtained in clinical practice with the use of levodopa and DDC inhibitor and with COMT inhibitors. Thus, the peripheral COMT inhibitors can improve the quality of PET imaging with 6-FD.
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V. Practical and Theoretical Clinical Uses of COMT Inhibitors |
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The main clinical application of COMT inhibitors would be as
adjunct (or additional adjunct) in the levodopa therapy of PD (Männistö and Kaakkola, 1989, 1990; Männistö et
al., 1992b, 1994). As discussed, COMT inhibitors can reduce the
formation of 3-OMD from L-dopa. Therefore, the
bioavailability of levodopa would be improved, its entry to the brain
increased, and possibly the half-life of L-dopa prolonged.
Peripherally active inhibitors would also be anticipated to achieve
these effects on L-dopa metabolism. A brain-entering
compound (e.g., tolcapone) might further potentiate the effect of
L-dopa by slowing down the metabolism of dopamine formed
from L-dopa in the brain. Thus, a triple therapy (levodopa plus DDC inhibitor plus COMT-inhibitor) would replace the present double therapy in PD.
It is conceivable that a COMT inhibitor could increase the formation of dopamine from L-dopa, even when administered without a DDC inhibitor. This combination may have some beneficial cardiovascular effects, such as increased renal blood flow and improvement in heart function.
COMT inhibitors could also potentiate or prolong the action of
compounds with a catechol structure (in addition to levodopa). Such
drugs include bronchodilating compounds (epinephrine, isoprenaline, rimiterol), dopamine agonists (dobutamine, fenoldopam, apomorphine), and some antihypertensive drugs (-methyldopa). There also is the
theoretical possibility of increasing the effect of endogenous catecholamines.
Inhibition of COMT in the brain would be beneficial in restoring norepinephrine levels and improving the symptoms caused by the deficit of this transmitter. Norepinephrine, or rather its deficiency, has been thought to be one of the main neurotransmitters implicitly involved in depression. Correction of this deficit with COMT inhibitors, in analogy to tricyclic uptake inhibitors and MAO inhibitors, could represent a novel way to treat depression.
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VI. Human Studies with COMT Inhibitors |
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Because CGP 28014 was not further developed for clinical purposes,
there is extremely limited human data on its properties (Bieck et al.,
1990, 1993); therefore, the clinical part of this review concentrates
on nitrocatechol-type COMT inhibitors.
A. Human Pharmacokinetics of COMT Inhibitors
The COMT inhibitors with nitrocatechol structure are rapidly
absorbed after oral administration, and
Cmax is usually reached in 0.5 to
2 h (see Fig. 5). Oral
bioavailability of tolcapone (60%) is about double that of entacapone
(32-36%), but the quantitative roles of poor absorption and the
first-pass metabolism in the incomplete bioavailability are still
unknown. Entacapone and nitecapone appear to be absorbed slightly more
rapidly than tolcapone. Clearly, higher
Cmax and AUC values are obtained with
tolcapone than with entacapone, in part due to better bioavailability
and lower clearance of tolcapone (Tables
3 and 4).
AUC and Cmax values are dose proportional after entacapone (Keränen et al., 1994) and
tolcapone (Dingemanse et al., 1995a). The volume of distribution at
steady state is small for all COMT inhibitors (Table 4). Nitrocatechols are abundantly bound to plasma proteins: tolcapone, about 99.9%, and
others, 97 to 98%. At the therapeutic doses, all COMT inhibitors are
rapidly eliminated, with an apparent T1/2
of 1.5 to 3 h after oral administration. After i.v.
administration, entacapone has the shortest
T1/2, about 0.5 h, and
tolcapone has the longest, about 1.2 h (Heikkinen et al., 1994;
Keränen et al., 1994; Dingemanse et al., 1995a).
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All COMT inhibitors are abundantly metabolized, mainly in the liver.
The main part of absorbed entacapone is eliminated via the biliary
route to feces (Wikberg et al., 1993; Heikkinen et al., 1994), whereas
about 40% of tolcapone dose is excreted to feces (Roche, 1997). Only
0.5% or less of an oral dose of entacapone and tolcapone is excreted
unchanged in the urine (Wikberg et al., 1993; Heikkinen et al., 1994).
The main urine metabolite is the glucuronide of the parent compound,
representing 70, 60, and 30% of the metabolites of entacapone,
nitecapone, and tolcapone, respectively (Taskinen et al., 1991; Wikberg
and Taskinen, 1993; Wikberg et al., 1993; Da Prada et al., 1994).
Entacapone [the (E)-isomer of the molecule] has one active
metabolite, its (Z)-isomer. Its AUC accounts for only about
5% of the total plasma AUC of both isomers (Wikberg et al., 1993;
Keränen et al., 1994). Entacapone and nitecapone are not
O-methylated in humans (Taskinen et al., 1991; Wikberg et
al., 1993), whereas about 3% tolcapone is converted to
3-O-methyltolcapone (Da Prada et al., 1994; Dingemanse,
1997; Roche, 1997). This metabolite has a long elimination
T1/2 of about 35 h (Dingemanse et al.,
1995a), which may suggest that accumulation could occur. However,
during the long-term administration of tolcapone, only minor
accumulation of 3-O-methyltolcapone was detected due to
suppression of its formation by tolcapone itself (Dingemanse et al.,
1996). In contrast to entacapone and nitecapone, about 3% of tolcapone
is oxidized by cytochrome P-450 isoenzymes to active alcohol and
carboxyl acid metabolites (Da Prada et al., 1994; Roche, 1997).
During long-term administration at therapeutic doses, neither
entacapone nor tolcapone accumulates in plasma (Dingemanse et al.,
1996; Jorga et al., 1997c; Gordin et al., 1998a). The combination of
levodopa and DDC inhibitor with entacapone or tolcapone does not
significantly affect the pharmacokinetics (Dingemanse et al., 1995b;
Gordin et al., 1998a; Jorga et al., 1998b). A dosage reduction for
entacapone and tolcapone is recommended in patients with liver impairment because of their increased bioavailability and reduced clearance (Jorga et al., 1997b; 1998c; Gordin et al., 1998b).
B. COMT Inhibition
All nitrocatechol-type COMT inhibitors dose-dependently and
reversibly inhibit the COMT activity in human erythrocytes (Kaakkola et
al., 1990; Keränen et al., 1994; Dingemanse et al., 1995a). Tolcapone appears to be the most potent of these inhibitors; it also
has the longest inhibitory activity. After a single dose (100-200 mg)
of entacapone and nitecapone, the COMT activity has fully recovered
within 6 to 8 h, whereas the corresponding time is 13 to 15 h
after tolcapone (Fig. 5). During repeated administration of entacapone
or tolcapone, no tolerance developed to the inhibitory activities
(Dingemanse et al., 1996; Gordin et al., 1998a). Nitecapone is better
at inhibiting human gastric and duodenal than erythrocyte COMT
activity, suggesting that it may have greater activity locally in the
intestine compared with other peripheral organs, as has also been
demonstrated in rats (Nissinen et al., 1988a; Schultz et al., 1991).
C. Effect on Levodopa Pharmacokinetics
Entacapone, nitecapone, and tolcapone dose-dependently increase
the AUC values of L-dopa in healthy volunteers, without
significantly changing Cmax values
(Table 5, Fig.
6; Kaakkola et al., 1990; Keränen
et al., 1993; Dingemanse et al., 1995b; Jorga et al., 1997c, 1998b;
Sedek et al., 1997; Jorga, 1998). Some tendency to prolongation of
Tmax values of
L-dopa has been observed with higher doses of
entacapone and tolcapone (Keränen et al., 1993; Dingemanse et
al., 1995b; Sedek et al., 1997). Tolcapone seems to be the most potent
of these COMT inhibitors in increasing the AUC of
L-dopa (Fig. 6). Multiple doses of entacapone or
tolcapone do not change their effects on L-dopa
pharmacokinetics (Dingemanse et al., 1996; Jorga et al., 1997c; Gordin
et al., 1998a). The effects of entacapone and tolcapone are generally
consistent regardless of the decarboxylase inhibitor (carbidopa or
benserazide) used with levodopa (Jorga et al., 1997c, 1998b;
Myllylä et al., 1997b). However, entacapone increases the AUC of
L-dopa 5 to 10% more with levodopa and
benserazide than with levodopa and carbidopa (Myllylä et al.,
1997b; Orion, 1998). Both entacapone and tolcapone are also effective
in combination with controlled-release levodopa preparations (Ahtila et
al., 1995; Jorga et al., 1997a, 1998b). In contrast to standard
levodopa and carbidopa formulation, when controlled-release levodopa
preparations are used, entacapone appears to slightly increase the peak
concentration of L-dopa, whereas tolcapone
increases the Tmax value of
L-dopa (Ahtila et al., 1995; Jorga et al.,
1998b).
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Similar to the situation in healthy volunteers, both entacapone and
tolcapone significantly increase the AUC of L-dopa in patients with PD. A 23 to 48% increase in the AUC of
L-dopa has been observed after a single 200-mg dose of
entacapone (Table 6). The corresponding
figures for tolcapone (200 mg) vary from 51 to 58%. There are no data
on L-dopa pharmacokinetics in PD patients after a single
dose of 100 mg of tolcapone, which is the recommended clinical dose for
tolcapone. After 7 to 8 weeks of treatment with tolcapone up to 200 mg
t.i.d., AUC values of L-dopa have increased by 33%
(Yamamoto et al., 1997). The corresponding value after 8 weeks of
treatment with entacapone (1200 mg/day) was 43% (Nutt et al., 1994).
The latter study is notable because repeated plasma samples were taken
throughout the day. This study demonstrated that during entacapone
treatment, the mean daily L-dopa concentration increased
despite the reduction in levodopa dose (
27%), and the daily
variation in plasma L-dopa levels significantly decreased
(Nutt et al., 1994). This would indicate less fluctuation in clinical
disability in PD patients, as was found by Nutt et al. (1994). The use
of entacapone for 10 days led to a significant increase in AUC of
L-dopa in PD patients receiving either standard or
controlled-release levodopa formulations (Kaakkola et al., 1995). In
this study, entacapone increased L-dopa peak level with a
controlled-release levodopa preparation but not with a standard levodopa preparation.
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COMT inhibitors reduce the elimination of L-dopa, which equates with an increase in its elimination T1/2. This effect has been observed consistently in studies of patients treated with entacapone (see Table 8). The same is true for tolcapone in both volunteer and PD patient studies (Tables 7 and 8).
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D. Effect on 3-OMD Levels
As would be expected, all COMT inhibitors reduce the formation of
3-OMD in both healthy subjects and PD patients. Because 3-OMD has a
long T1/2, about 15 h (Kuruma et al.,
1971), the inhibition of 3-OMD formation in PD patients can be observed
only after long-term treatment with COMT inhibitors. In healthy
subjects, single doses of entacapone, nitecapone, and tolcapone
dose-dependently suppressed the formation of 3-OMD, with tolcapone
being the most potent of the three inhibitors (Table 5, Fig. 6;
Kaakkola et al., 1990; Keränen et al., 1993; Dingemanse et al.,
1995b; Sedek et al., 1997; Jorga et al., 1998b). A similar inhibition
of 3-OMD formation has been observed after entacapone or tolcapone in
combination with controlled-release levodopa formulations (Ahtila et
al., 1995; Jorga et al., 1997a, 1998b). In PD patients, about 50 and 70% inhibition of 3-OMD formation has been demonstrated after entacapone and tolcapone, respectively (Table 7).
E. Effect on Plasma Catecholamine Metabolism
Because COMT is involved in the metabolism of dopamine,
norepinephrine, and epinephrine, it is also necessary to consider the
actions of COMT inhibitors on peripheral catecholamine metabolism. A
single dose (up to 100 mg) or 1-week treatment with nitecapone (100 mg
t.i.d.) has not affected plasma dopamine, norepinephrine, and
epinephrine levels either at rest or during exercise (Sundberg et al.,
1990, 1993b). The same result has been obtained after treatment with a
single dose (up to 200 mg) or repeated doses (400 or 800 mg t.i.d. for
1 week) of entacapone (Sundberg et al., 1993a; Illi et al., 1994,
1996a). However, both entacapone and nitecapone significantly changed
the metabolic profile of plasma catecholamines. Plasma concentrations
of the MAO-dependent metabolites DOPAC and 3,4-dihydroxyphenyl glycol
increased, whereas those of the COMT-dependent metabolite MHPG
decreased after entacapone and nitecapone administration (Sundberg et
al., 1990, 1993a,b; Illi et al., 1994, 1996a). There are no published
data available concerning the effect of tolcapone on plasma
catecholamines or their metabolites.
In healthy subjects treated with levodopa and DDC inhibitor, COMT
inhibitors increased substantially the plasma concentrations of DOPAC
and induced a variable decrease in levels of HVA (Table 5; Kaakkola et
al., 1990; Keränen et al., 1993; Zürcher et al., 1993;
Ahtila et al., 1995; Sedek et al., 1997).
In PD patients, a single dose of entacapone (200 mg) in combination
with levodopa and benserazide did not alter the plasma levels of
dopamine and norepinephrine but decreased those of MHPG (Lyytinen et
al., 1997). In contrast, tolcapone has been reported to elevate
significantly plasma dopamine levels (Oechsner et al., 1998). Similar
to the situation in healthy subjects, treatment with entacapone or
tolcapone decreased the plasma concentrations of HVA and increased
those of DOPAC in PD patients (Myllylä et al., 1993; Davis et
al., 1995a; Ruottinen and Rinne, 1996a,c; Lyytinen et al., 1997;
Oechsner et al., 1998).
These results suggest that supplementation of the present fixed ratio
levodopa and DDC inhibitor therapy with a COMT inhibitor may lead to
the conversion of some peripheral L-dopa to dopamine and
further to DOPAC because the extent of the DDC inhibition may not be
sufficient. Oechsner et al. (1998) suggested that the dose of the DDC
inhibitor should be increased when combined with tolcapone to avoid the
peripheral side effects of dopamine. It would be interesting to
evaluate the different possible ratios of levodopa, DDC inhibitor, and
COMT inhibitor to identify the optimum dosage schedules.
F. Clinical Efficacy
Only entacapone and tolcapone have been studied in true clinical trials. Both have been effective in several open and double-blind clinical studies. It should be noted that due to serious, although rare, adverse reactions, marketing of tolcapone was suspended in the European Union and Canada in late 1998. In the United States, tolcapone should be used as an adjunct only in patients with PD on levodopa and carbidopa who are experiencing symptom fluctuations and who are not responding satisfactorily to or who are not appropriate candidates for other adjunctive therapies. The reasons for tolcapone withdrawal are discussed in Safety. As of early 1999, entacapone is marketed throughout the European Union. It is intended as an adjunct to standard preparations of levodopa and carbidopa or levodopa and benserazide in the treatment of patients with PD and end-of-dose motor fluctuation.
The comparison of the results achieved with these two inhibitors is complicated because of differences in patient materials, study designs, treatment periods, medications, and presentations of the results. With these reservations, some of the clinical results are presented in Tables 8 and 9. All studies with entacapone and most of the studies with tolcapone have been conducted in PD patients who were experiencing clinical fluctuations [i.e., end-of-dose deterioration (wearing off)] and often dyskinesias.
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Both entacapone and tolcapone as single doses with levodopa and DDC
inhibitor show significant clinical benefits (Table 8). Both drugs
prolong the motor response (the "on time") to levodopa. The dose of
200 mg of entacapone with each levodopa and DDC inhibitor dose has
been selected for further clinical use, in part based on the results of
a dose-finding study (Ruottinen and Rinne, 1996a) and in part because
of compatible pharmacokinetic profiles of levodopa and entacapone. The
dose-response relationship for tolcapone has not been as unambiguous in
clinical trials (Tables 8 and 9). At present, the manufacturer
recommends that tolcapone should be initiated with 100 mg t.i.d.
together with the first levodopa dose and then at 6-h interval with a
12-h nighttime break (Roche, 1997, 1998). The dose can be increased to
200 mg t.i.d., but dopaminergic adverse reactions may be a limiting factor.
The efficacy of entacapone has been elucidated in two double-blind
studies of 6 months' duration and that of tolcapone in several
double-blind studies of a maximum of 3 months' duration (Table 9).
Both entacapone and tolcapone generally increase the on time and
correspondingly decrease the "off time" in advanced PD patients.
The actual increase in daily on time has varied from about 1 to 2 h with entacapone and from about 0 to 2.5 h with tolcapone (Table
9). Typically, the clinical efficacy of COMT inhibitors is observed in
the early days of treatment, as would be expected because they increase
the AUC and half-life of L-dopa already on the first day.
In line with their effect on L-dopa pharmacokinetics, COMT
inhibitors permit a reduction in daily levodopa dosage by about 100 to
200 mg. After withdrawal of entacapone or tolcapone, a rapid worsening
of PD symptoms is observed and a levodopa dose adjustment upward is
needed (Parkinson Study Group, 1997; Roche, 1997, 1998; Rinne et al.,
1998).
Entacapone has potentiated the magnitude of the levodopa effect; the
scores of Unified Parkinson's Disease Rating Scale (UPDRS) have
significantly improved (Parkinson Study Group, 1997; Rinne et al.,
1998). A similar tendency, although generally nonsignificant, has been
observed in long-term studies with tolcapone (Baas et al., 1997; Dupont
et al., 1997; Myllylä et al., 1997a; Rajput et al., 1997; Adler
et al., 1998). Furthermore, investigators' global measures of disease
severity indicate that both entacapone and tolcapone have positive
effects on PD symptoms (Baas et al., 1997; Kurth et al., 1997;
Myllylä et al., 1997a; Parkinson Study Group, 1997; Rajput et
al., 1997; Adler et al., 1998; Rinne et al., 1998). The patients'
self-reported global evaluations ("patient's diaries") demonstrate
similar positive results in entacapone studies (Parkinson Study Group,
1997; Rinne et al., 1998).
The clinical efficacy of tolcapone has also been investigated in
nonfluctuating PD patients. One of these studies included PD patients
whose fluctuations were controlled by more frequent levodopa dosing
(Dupont et al., 1997). At 6 weeks, tolcapone groups (200 and 400 mg
t.i.d.) had moderately greater dose reduction in their levodopa dose
than the placebo group (about 180 mg for tolcapone and 110 mg for
placebo). The only statistically significant clinical benefit of
tolcapone was observed in UPDRS subscale II (activities of daily
living) with 200 mg t.i.d. dosing. In another study, patients with
wearing off phenomena were excluded. Thus, this can be considered as
more representative of nonfluctuating PD patients (Waters et al.,
1997). At 6 months, tolcapone (100 or 200 mg t.i.d.) produced a
significant improvement in disability, as assessed by UPDRS and
quality-of-life measures. Levodopa doses were slightly but
significantly decreased in the tolcapone groups (about 30 mg). The
beneficial effects of tolcapone were still maintained at 12 months.
Because there have been no studies carried out directly comparing entacapone and tolcapone, it is not possible to conclude whether there are any significant differences in clinical efficacy.
G. Safety
Overall, the COMT inhibitors have been well tolerated. For instance, the number of premature terminations has not differed significantly between placebo and active treatment groups. In general, the incidence of adverse events appears to be higher in tolcapone-treated PD patients than in entacapone-treated PD patients, although a similar trend is also noticed in the placebo groups (Table 10). The main adverse effects related to entacapone and tolcapone are dopaminergic and gastrointestinal events. In all studies, the most commonly observed dopaminergic adverse event has been a worsening of levodopa-induced dyskinesia. In the majority of cases, this occurs during the first weeks of treatment with the COMT inhibitor. The severity of dyskinesia can be minimized by levodopa dose adjustment. Indeed, this adverse event has rarely lead to treatment withdrawal. Nausea, anorexia, vomiting, orthostatic hypotension, sleep disorders, and hallucinations are other dopaminergic events that may be potentiated by a COMT inhibitor.
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The most common nondopaminergic adverse event is diarrhea. Tolcapone
has been associated with diarrhea in about 16 to 18% of cases, and
entacapone has been associated in less than 10% of cases (Table 10).
It usually occurs within the first 6 to 16 weeks of the treatment. The
diarrhea, dose related in the case of tolcapone, may be severe, whereas
most cases in entacapone-treated patients have been mild to moderate.
Diarrhea has been the most common cause of treatment discontinuation: 5 to 6% of patients treated with tolcapone (Roche, 1997, 1998) and 2.5%
of patients treated with entacapone (Orion, 1998). In another study,
diarrhea was experienced by 24% of patients treated with tolcapone
(200 mg t.i.d.) plus selegiline (5 mg b.i.d.) without levodopa for 1 month (Hauser et al., 1998).
Patients should be warned that their urine may color to dark yellow or orange due to the presence of COMT inhibitors and their metabolites. This harmless event appears to be more common with entacapone than with tolcapone.
Mild decreases in hemoglobin, erythrocyte counts, and hematocrit values
have been reported during entacapone therapy, probably due to its
iron-chelating property. This adverse effect has been clinically
significant in 1.5% of the patients (Orion, 1998).
Elevated liver transaminase levels have been reported in 1 to 3% of
patients treated with tolcapone (Roche, 1997), whereas significant
transaminase changes have been observed very rarely in patients treated
with entacapone (Orion, 1998). Recently, three cases of acute, fatal
fulminant hepatitis were described in association with tolcapone
treatment (Assal et al., 1998; European Medicine Evaluation Agency,
1998). In addition, potentially fatal neurological adverse reactions,
including neuroleptic-like malignant syndrome and rhabdomyolysis, were
described. Due to these serious adverse drug reactions, the regulatory
authorities in the European Union and Canada decided that marketing of
tolcapone should be suspended. At this time, more than 100,000 patients
have been treated with tolcapone. In many other countries, including
the United States, the use of tolcapone has now been restricted to
patients experiencing fluctuations and who were not appropriate
candidates for other adjunctive therapies. In these patients, liver
function tests should be taken at baseline and regularly every 2 weeks
after the initiation of tolcapone treatment. It is not currently
considered essential to monitor liver enzyme levels during entacapone therapy.
H. Drug Interactions
There are some pharmacokinetic interactions between COMT
inhibitors and other drugs. Many of these interactions are based on the
fact that the interacting drugs are also substrates of COMT. Most
published information is available for entacapone and nitecapone, but
the same general principles probably can be extended to tolcapone. Some
data can be derived from the official product monographs (Roche, 1997,
1998; Orion, 1998).
The poor solubility of entacapone at acidic pH of the stomach could be
the reason for its poor bioavailability in rats and humans. A high dose
of entacapone seems to decrease the absorption of carbidopa in studies
on human volunteers (Ahtila et al., 1995). Entacapone may chelate iron
in the gastrointestinal tract, and therefore it is advised to have a 2- to 3-h interval between the administration of iron preparations and
entacapone (Orion, 1998). Tolcapone seems to inhibit the
O-methylation of benserazide, which is an excellent
substrate of COMT, to the extent that the brain entry of this compound
is increased. If this should occur, then benserazide can start to
inhibit dopamine synthesis in the striatum. This is not the case with
carbidopa, which is a much less favorable substrate of COMT (Tedroff et
al., 1991).
The infusion of catecholamines or isoprenaline, all of which are
substrates of COMT, as well as physical exercise, which increases plasma levels of catecholamines, can cause disturbances in heart rhythm. COMT inhibitors have only slightly potentiated this natural activity, but there is some prolongation of the duration of action of
catecholamines on pulse rate and blood pressure. Entacapone or
nitecapone does not alter cardiovascular responses, hemodynamics, or
concentrations of unconjugated catecholamines in plasma of healthy
volunteers, but the metabolism profile was clearly shifted toward the
MAO-dependent pathways (Sundberg et al., 1990, 1993a; Myllylä et
al., 1993; Illi et al., 1994, 1995). Furthermore, the tyramine pressor
response was not enhanced by nitecapone (Sundberg and Gordin, 1991).
Tolcapone does not modify the action of ephedrine, an indirectly acting
sympathomimetic drug (Roche, 1997). Moclobemide, an MAO-A inhibitor
(Illi et al., 1996a), or imipramine (Illi et al., 1996b) did not
significantly interact with entacapone, nor did desipramine interact
with tolcapone (Roche, 1997, 1998).
Because entacapone and tolcapone undergo different routes of
metabolism, there could be differences in their interactions with the
metabolism of other drugs. In the case of tolcapone, there is no sign
of competition for the glucuronidation of desipramine or cytochromal
oxidation of tolbutamide or warfarin (Roche, 1997, 1998). Entacapone is
not eliminated via cytochrome P-450 enzymes (Wikberg et al., 1993;
Wikberg and Vuorela, 1994).
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VII. Summary |
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COMT O-methylates catecholamines and other compounds with a catechol structure. The general function of COMT is the elimination of biologically active or toxic catechols and some other hydroxylated metabolites. During the first trimester of pregnancy, COMT present in the placenta protects the developing embryo from activated hydroxylated compounds. COMT also acts as an enzymatic detoxicating barrier between the blood and other tissues shielding against the detrimental effects of xenobiotics. COMT may serve some unique or indirect functions in the kidney and intestine tract by modulating the dopaminergic tone; the same may be true in the brain: COMT activity may regulate the amounts of active dopamine and norepinephrine in various part of the brain and therefore be associated with the mood and other mental processes.
There is one single gene for COMT, which codes for both S-COMT and MB-COMT using two separate promoters. Both rat and human S-COMTs contain 221 amino acids, and their molecular weights are 24.8 and 24.4 kD, respectively. Rat MB-COMT contains 43 and human MB-COMT contains 50 additional amino acids, of which 17 (rat) and 20 (human) are hydrophobic membrane anchors. The remainder of the MB-COMT molecule is suspended on the cytoplasmic side of the intracellular membranes. Rat S-COMT has been recently crystallized at 1.7- to 20-Å resolution. The active site of COMT consists of the AdoMet-binding domain and the actual catalytic site. The catalytic site is formed by a few amino acids that are important for the binding of the substrate, water, and Mg2+ and for the catalysis of O-methylation. The Mg2+, which is bound to COMT only after AdoMet binding, improves the ionization of the hydroxyl groups. The lysine residue (Lys144), which accepts the proton of one of the hydroxyls, acts as a general catalytic base in the nucleophilic methyl transfer reaction.
A series of new and highly selective COMT inhibitors have been developed. Entacapone, nitecapone, and tolcapone are nitrocatechol-type potent COMT inhibitors in vitro (Ki in nanomolar range), whereas CGP 28014 is a hydroxypyridine derivative and ineffective in vitro. In animal studies, these compounds effectively inhibit the O-methylation of L-dopa, thus improving its bioavailability and brain penetration and potentiating its behavioral effects. Entacapone and nitecapone have mainly a peripheral effect, whereas tolcapone and CGP 28014 also inhibit the O-methylation in the brain. In human volunteers, entacapone, nitecapone, and tolcapone dose-dependently inhibit the COMT activity of erythrocytes, improve the bioavailability of L-dopa, and inhibit the formation of 3-OMD.
In clinical studies in PD patients, both entacapone and tolcapone potentiate the therapeutic effect of L-dopa and prolong the daily on time by 1 to 2 h. The two marketed COMT inhibitors have different treatment strategies, advantages, and disadvantages, which are listed in Table 11.
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In the clinic, COMT inhibitors have been well tolerated, and the number of premature terminations has been low. In general, the incidence of adverse events has been higher in tolcapone-treated patients than in entacapone-treated patients. The main events have consisted of dopaminergic and gastrointestinal problems. Dopaminergic overactivity causes an initial worsening of levodopa-induced dyskinesia, nausea, vomiting, orthostatic hypotension, sleep disorders, and hallucinations. Tolcapone has been associated with diarrhea in about 16 to 18% of cases, and entacapone has been associated in less than 10% of cases. Diarrhea has led to discontinuation in 5 to 6% of patients treated with tolcapone and in 2.5% of those treated with entacapone. Urine discoloration to dark yellow or orange is related to the color of COMT inhibitors and their metabolites. Elevated liver transaminase levels are reported in 1 to 3% of patients treated with tolcapone but very rarely, if at all, in patients treated with entacapone. Three cases of acute, fatal fulminant hepatitis have been described in association of tolcapone when more than 100,000 patients have been treated. In addition, a few potentially fatal neurological adverse reactions, including neuroleptic-like malignant syndrome, have described. Because of these serious adverse drug reactions, tolcapone marketing was suspended in Europe and Canada. In early 1999, no restrictions of the use of entacapone have been proposed.
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VIII. Future Aspects |
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Assuming that the safety of entacapone can be substantiated, the
next development would be to prepare a combination tablet, containing
levodopa, a DDC inhibitor, and entacapone. However, before that is
marketed, it will be necessary to perform extensive pharmacokinetic
trials to determine the optimum doses of each component in the tablet.
We would propose that the amount of the DDC inhibitor must be increased
and the amount of entacapone probably will have to be reduced. Oechsner
et al. (1998) reached the same conclusion in the case of tolcapone. A
fixed combination tablet would add the patients' compliance but may
also complicate the individual dose adjustments.
For safety reasons, it would be necessary to clarify the new directions of the metabolism of L-dopa when its metabolism by both DDC and COMT is inhibited. Also, the consequences of inhibition of the inactivation of catecholestrogens by COMT inhibitors should be studied in detail.
A further point of concern is the action of COMT inhibition in the brain. Because COMT knockout mice exhibited behavioral abnormalities, the central actions of COMT inhibitors require clarification.
With respect to completely new COMT inhibitors, it would be necessary to also develop and study compounds not having a nitrocatechol structure because this may be associated with some risk of side effects. In particular, a compound with good bioavailability would be an improvement on the currently marketed drugs.
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Acknowledgments |
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Studies before 1995 cited in this review were supported by the Academy of Finland, and the newer ones were supported by the University of Kuopio through the Ministry of Education. We are also very grateful for discussions with the research staff of both F. Hoffman-La Roche, particularly Karin Jorga, and Orion-Pharma, particularly Dr. Ariel Gordin.
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Footnotes |
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1 Address for correspondence: Dr. Pekka T. Männistö, Department of Pharmacology and Toxicology, University of Kuopio, P.O. Box 1627, FIN-70 211 Kuopio, Finland. E-mail: Pekka.Mannisto{at}uku.fi
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Abbreviations |
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COMT, catechol-O-methyltransferase; S-COMT, soluble catechol-O-methyltransferase; MB-COMT, membrane-bound catechol-O-methyltransferase; AdoMet, S-adenosyl-L-methionine; AUC, area under the curve; DBA, dihydroxybenzoic acid; Cmax, maximum drug concentration in plasma; DOPAC, 3,4-dihydroxyphenyl acetic acid; 6-FD, 6-[18F]fluorodopa; MAO, monoamine oxidase; 3-OMD, 3-O-methyl-dopa; 3-OMFD, 3-O-methyl-[18F]fluorodopa; HVA, homovanillic acid; MHPG, 3-methoxy-4-hydroxyphenyl glycol; DDC, dopa decarboxylase; 3-MT, 3-methoxytyramine; PET, positron emission tomography; UPDRS, Unified Parkinson's Disease Rating Scale; PD, Parkinson's disease; T1/2, elimination half-life; Tmax, time to reach Cmax; U-0521, 3',4'-dihydroxy-2-methylpropiophenone; MPTP, N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; levodopa, a drug preparation containing L-dopa; L-dopa, L-3,4-dihydroxyphenylalanine.
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H. Lu, X. Meng, and C. S. Yang Enzymology of Methylation of Tea Catechins and Inhibition of Catechol-O-methyltransferase by (-)-Epigallocatechin Gallate Drug Metab. Dispos., May 1, 2003; 31(5): 572 - 579. [Abstract] [Full Text] [PDF]
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H. Lu, X. Meng, C. Li, S. Sang, C. Patten, S. Sheng, J. Hong, N. Bai, B. Winnik, C.-T. Ho, et al. Glucuronides of Tea Catechins: Enzymology of Biosynthesis and Biological Activities Drug Metab. Dispos., April 1, 2003; 31(4): 452 - 461. [Abstract] [Full Text] [PDF]
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J. H. Callicott, M. F. Egan, V. S. Mattay, A. Bertolino, A. D. Bone, B. Verchinksi, and D. R. Weinberger Abnormal fMRI Response of the Dorsolateral Prefrontal Cortex in Cognitively Intact Siblings of Patients With Schizophrenia Am J Psychiatry, April 1, 2003; 160(4): 709 - 719. [Abstract] [Full Text] [PDF]
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M. Akil, B. S. Kolachana, D. A. Rothmond, T. M. Hyde, D. R. Weinberger, and J. E. Kleinman Catechol-O-Methyltransferase Genotype and Dopamine Regulation in the Human Brain J. Neurosci., March 15, 2003; 23(6): 2008 - 2013. [Abstract] [Full Text] [PDF]
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R. K. Dubey, D. G. Gillespie, L. C. Zacharia, F. Barchiesi, B. Imthurn, and E. K. Jackson CYP450- and COMT-Derived Estradiol Metabolites Inhibit Activity of Human Coronary Artery SMCs Hypertension, March 1, 2003; 41(3): 807 - 813. [Abstract] [Full Text] [PDF]
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M. Forsberg, M. Lehtonen, M. Heikkinen, J. Savolainen, T. Jarvinen, and P. T. Mannisto Pharmacokinetics and Pharmacodynamics of Entacapone and Tolcapone after Acute and Repeated Administration: A Comparative Study in the Rat J. Pharmacol. Exp. Ther., February 1, 2003; 304(2): 498 - 506. [Abstract] [Full Text] [PDF]
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 C. Worda, M.O. Sator, C. Schneeberger, T. Jantschev, K. Ferlitsch, and J.C. Huber Influence of the catechol-O-methyltransferase (COMT) codon 158 polymorphism on estrogen levels in women Hum. Reprod., February 1, 2003; 18(2): 262 - 266. [Abstract] [Full Text] [PDF]
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M. Huotari, M. Santha, L. R. Lucas, M. Karayiorgou, J. A. Gogos, and P. T. Mannisto Effect of Dopamine Uptake Inhibition on Brain Catecholamine Levels and Locomotion in Catechol-O-methyltransferase-Disrupted Mice J. Pharmacol. Exp. Ther., December 1, 2002; 303(3): 1309 - 1316. [Abstract] [Full Text] [PDF]
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 M. J. Bonifacio, M. Archer, M. L. Rodrigues, P. M. Matias, D. A. Learmonth, M. A. Carrondo, and P. Soares-da-Silva Kinetics and Crystal Structure of Catechol-O-Methyltransferase Complex with Co-Substrate and a Novel Inhibitor with Potential Therapeutic Application Mol. Pharmacol., October 1, 2002; 62(4): 795 - 805. [Abstract] [Full Text] [PDF]
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M. Yan, L. T. Webster Jr., and J. L. Blumer 3-O-Methyldobutamine, a Major Metabolite of Dobutamine in Humans Drug Metab. Dispos., May 1, 2002; 30(5): 519 - 524. [Abstract] [Full Text] [PDF]
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M. Yan, L. T. Webster Jr., and J. L. Blumer Kinetic Interactions of Dopamine and Dobutamine with Human Catechol-O-methyltransferase and Monoamine Oxidase in Vitro J. Pharmacol. Exp. Ther., April 1, 2002; 301(1): 315 - 321. [Abstract] [Full Text] [PDF]
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L. C. Zacharia, E. K. Jackson, D. G. Gillespie, and R. K. Dubey Catecholamines Block 2-Hydroxyestradiol-Induced Antimitogenesis in Mesangial Cells Hypertension, April 1, 2002; 39(4): 854 - 859. [Abstract] [Full Text] [PDF]
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F. Barchiesi, E. K. Jackson, D. G. Gillespie, L. C. Zacharia, J. Fingerle, and R. K. Dubey Methoxyestradiols Mediate Estradiol-Induced Antimitogenesis in Human Aortic SMCs Hypertension, April 1, 2002; 39(4): 874 - 879. [Abstract] [Full Text] [PDF]
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E. L. Cavalieri, P. Devanesan, M. C. Bosland, A. F. Badawi, and E. G. Rogan Catechol estrogen metabolites and conjugates in different regions of the prostate of Noble rats treated with 4-hydroxyestradiol: implications for estrogen-induced initiation of prostate cancer Carcinogenesis, February 1, 2002; 23(2): 329 - 333. [Abstract] [Full Text] [PDF]
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R. K. Dubey, D. G. Gillespie, L. C. Zacharia, M. Rosselli, B. Imthurn, and E. K. Jackson Methoxyestradiols Mediate the Antimitogenic Effects of Locally Applied Estradiol on Cardiac Fibroblast Growth Hypertension, February 1, 2002; 39(2): 412 - 417. [Abstract] [Full Text] [PDF]
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5% more frequently in either COMT
inhibitor group compared with placebo group are presented.