++
Purines comprise bases, nucleosides in association with ribose
or deoxyribose, and nucleotides with one or more added phosphate groups.
Purine nucleotides are essential cellular constituents. They are
the building blocks of the polynucleotides, DNA and RNA, and, under
the form of mononucleotides or of nucleosides, also intervene in
numerous cellular functions. Among these are energy transfer (eg, by
adenosine triphosphate [ATP]), metabolic regulation
(eg, by guanosine triphosphate [GTP]), and signaling
(eg, by adenosine). Purine metabolism can be divided into three
pathways (Fig. 168-1):
++
++
1. The biosynthetic pathway, starts
with the formation, often termed de novo, of the
high-energy compound phosphoribosyl pyrophosphate (PRPP) and leads
in 10 steps to the synthesis of the nucleoside monophosphate inosine
monophosphate (IMP). From IMP, two reactions lead to the formation
of adenosine monophosphate (AMP). Subsequently, the nucleoside di-
and triphosphates ADP and ATP and their deoxy counterparts are synthesized.
Two other reactions convert IMP into GMP, from which GDP, GTP, and
their deoxy counterparts are formed.
2. The purine catabolic pathway starts from the
nucleoside monophosphates GMP, IMP, and AMP and produces uric acid,
a poorly soluble compound that tends to crystallize once its plasma
concentration increases above 6.5 to 7.0 mg/dl (0.38–0.47
mmol/l).
3. The purine salvage pathway utilizes the purine
bases guanine, hypoxanthine, and adenine, which are provided by
food intake or the catabolic pathway, and reconverts them by phosphoribosylation
into GMP, IMP, and AMP, respectively. It also utilizes the purine
nucleosides adenosine, guanosine, and their deoxy counterparts by
phosphorylation into the corresponding monophosphates, catalyzed
by kinases.
++
Inborn errors of purine metabolism comprise errors of purine
nucleotide synthesis, of purine catabolism, and of purine salvage. They
should be considered in patients with hyper- or hypouricemia, kidney
stones, and a variety of muscle, neurological, and other symptoms
(Table 168-1). The deficiencies of adenosine
deaminase and purine nucleoside phosphorylase, two
purine catabolic enzymes, cause severe combined immunodeficiency
(SCID; see Chapter 188). Adenosine deaminase
superactivity causes hemolytic anemia. The deficiency of deoxyguanosine
kinase causes a mitochondrial disease. The
deficiency of thiopurine methyltransferase is of importance
in pharmacogenetics.
++
+++
Phosphoribosylpyrophosphate
Synthetase Superactivity
++
Phosphoribosylpyrophosphate (PRPP) synthetase superactivity (OMIM
311850), one of the few known examples of a hereditary anomaly that
enhances the activity of an enzyme, was initially reported by Sperling
and colleagues1 in a patient with early adult-onset
gout. Since then, about two dozen families have been identified
in which excessive production of uric acid could be traced to this
disorder.2,3
+++
Clinical Presentation
++
The disorder presents mostly in young adult males with uric acid
lithiasis or gouty arthritis. The high incidence of renal calculi
composed of uric acid potentially leads to renal insufficiency.
A few patients present in infancy with clinical signs of uric acid
overproduction accompanied by neurological abnormalities, mainly
sensorineural deafness (particularly for high tones) but also hypotonia,
locomotor delay, ataxia, and autistic features. The disorder is
X-linked, but hemizygous females with gout are also found.4 Uricemia
can be very high, reaching 10 to 15 mg/dl (0.60–0.90 mmol/l;
normal is 2.9–5.5 mg/dl, or 0.17–0.32 mmol/l).
Affected individuals synthesize purines at an excessive rate, manifested
by uric acid excretion ranging from 1.0 to 2.4 g per 24 hours (normal
0.5–0.8 g) while maintained on a purine-free diet.
+++
Metabolic Derangement
++
PRPP synthetase catalyzes the formation (from ribose-5-phosphate
and ATP) of PRPP, the first intermediate in the de novo synthesis of
purine nucleotides (Fig. 168-1). Three distinct
isoforms of PRPP synthetase—PRS 1, 2, and 3—have
been identified. They encode highly homologous polypeptides, which
nevertheless differ by several physical and kinetic properties.
PRPP synthetases are subjected to complex regulation, including
stimulation by Pi and feedback inhibition by the nucleotides produced
by PRPP utilizing pathways such as ADP and GDP. PRPP synthetase
superactivity involves the PRS1 isoform. It can be caused by various
kinetic anomalies2,3: (1) regulatory defects, such
as increased sensitivity to the stimulator of the enzyme Pi or decreased
sensitivity to its inhibitors ADP and GDP; (2) catalytic defects
with increased Vmax owing to overproduction of the enzyme
protein; (3) combined regulatory and catalytic defects; and (4)
increased affinity for the substrate ribose-5-phosphate. Since physiological
concentrations of PRPP are not saturating for PRPP amidotransferase,
(which is the first rate-limiting enzyme of the de novo pathway), the
first rate-limiting enzyme of the de novo pathway increased synthesis
of PRPP will result in increased production of uric acid.
++
The three isoforms of PRPP synthetase are encoded by separate
genes, PRPS1, PRPS2, and PRPS1L1. PRPS1 and PRPS2 map to the long and
short arms of the X chromosome (Xq22q-q24 and Xp22.2-p22.3, respectively)
and are expressed in all tissues. The expression of PRPS1L1, an
intronless gene located on chromosome 7, is restricted to testes.
PRPP synthetase superactivity is caused by gain-of-function mutations
of the PRPS1 gene. In six families with purine nucleotide feedback
resistant PRPP synthetase, distinctive point mutations were found
that, when expressed, displayed kinetic properties comparable to those
found in cells from the respective affected individual.5 In
contrast, in patients with catalytically overactive PRPP synthetase,
a two- to fourfold selective acceleration of transcription of a
normal PRPS1 isoform was identified.6 The genetic
basis of this acceleration remains unresolved, since normal- and
patient-derived genes have identical nucleotide sequences up to
850 base pairs 5' to the consensus transcription initiation site.
++
Diagnosis of PRPP synthetase superactivity requires extensive
kinetic studies of the enzyme that can be performed on erythrocytes
or on cultured skin fibroblasts in a few specialized laboratories.
The disorder should be differentiated from partial hypoxanthine-guanine phosphoribosyltransferase
(HPRT) deficiency, which gives similar clinical signs (see below).
Familial juvenile hyperuricemia7 should also be
considered.
+++
Treatment and
Prognosis
++
Patients should be treated with the xanthine oxidase inhibitorallopurinol. This compound is oxidized by xanthine oxidase into
oxypurinol, which has a very high affinity for the reduced molybdenum
site of this enzyme, resulting in essentially irreversible inhibition of
its activity. Inhibition of xanthine oxidase, the last enzyme of
purine catabolism (Fig. 168-1), decreases
the production of uric acid and its replacement by hypoxanthine,
which is about ten times old more soluble, and by xanthine, which
is slightly more soluble than uric acid. Initial allopurinol dosage
is 10 to 20 mg/kg per day in children and 2 to 10 mg/kg
per day in adults. It should be adjusted to the minimum required
to maintain normal uric acid levels in plasma and should be reduced
in subjects with renal insufficiency. In rare patients with a considerable
increase in de novo synthesis, xanthine calculi can be formed during allopurinol
therapy.8 Additional measures to prevent crystallization
are thus recommended. These include a low purine diet (free of organ meats,
sardines, dried beans and peas); high fluid intake; and, since uric
acid and xanthine are more soluble at alkaline than at acid pH, administration
of sodium bicarbonate, potassium citrate, or citrate mixtures to bring urinary pH to 6.0
to 6.5. Adequate control of the uricemia prevents gouty arthritis
and urate nephropathy but does not correct the neurological symptoms.
+++
Phosphoribosylpyrophosphate
Synthetase Deficiency
++
A severe deficiency of PRPP synthetase has been reported by Wada
and colleagues9 in a Japanese boy with hypouricemia,
mental retardation, convulsions, increased urinary excretion of
orotic acid, and megaloblastic bone-marrow changes. He showed a
remarkable and unexplained recovery of enzyme activity after treatment
with adrenocorticotrophic hormone. His disease has, however, not
been proven genetic.
++
Recently, loss-of-function mutations of the PRPS1 gene have been
identified in several patients with X-linked disorders characterized by
peripheral neuropathy, hearing impairment, and optic atrophy; these
disorders mainly included Arts syndrome (OMIM 301835),10 Rosenberg-Chutorian
syndrome, and Charcot-Marie Tooth disease CMTX5 (OMIM 311070).11 PRPP
synthetase activity was markedly decreased, although not completely deficient.
There were no reductions of serum and urinary uric acid. It is speculated
that the defect could be compensated by synthesis of ATP via the
S-adenosylmethionine pathway and that administration of the latter
compound could be beneficial.
+++
Adenylosuccinate Lyase
Deficiency
++
The deficiency of adenylosuccinate lyase (ADSL, also known as
adenylosuccinase; OMIM 103050) is the first enzyme defect reported
in humans to affect the de novo pathway of purine synthesis.12 Over
60 patients have been identified so far.13,14
+++
Clinical Presentation
++
Patients present with widely variable associations of psychomotor
retardation, epilepsy, autistic features (failure to make eye-to-eye contact,
repetitive behavior, temper tantrums, autoagressivity), hypotonia,
and secondary feeding problems. Several clinical pictures can be
distinguished. In the first reported presentation, often referred
to as type I, patients display moderate to severe
psychomotor retardation, frequently accompanied by epilepsy after
the first years and various behavior disturbances. In some patients,
intractable convulsions start within the first days to weeks of life15-17 and
are sometimes associated with severe muscular hypotonia necessitating
mechanical ventilation.18 Impaired intrauterine growth,
microcephaly, fetal hypokinesia, and a lack of fetal heart rate
variability have also been observed.18
++
Rare patients, referred to as type II, are strikingly
less retarded as compared with the other cases.19 One
patient suffered from profound muscle hypotonia accompanied by slightly
delayed motor development.20 Finally, a few patients
display mainly autistic features.14,21,22 Dysmorphic
features are usually absent, but a suggestive facial dysmorphism, including
brachycephaly, anteverted nostrils, long smooth philtrum, and thin
upper lip, has been reported.23 Fundoscopy; auditory,
somatosensory, and visual evoked responses; nerve conduction velocities;
and electromyography are normal, the latter also being true for
patients with muscular hypotonia and wasting. Computed tomography
and brain MRI often show hypotrophy or hypoplasia of the cerebellum,
particularly of the vermis.
+++
Metabolic Derangement
++
ADSL catalyzes two steps in the synthesis of purine nucleotides
(Fig. 168-1): (1) the conversion of succinylaminoimidazolecarboxamide
ribotide (SAICAR) into aminoimidazolecarboxamide ribotide (AICAR),
the eighth step of the de novo pathway, and (2)
the formation of AMP from adenylosuccinate (S-AMP), the second step
in the conversion of IMP into AMP. The reaction is a nonhydrolytic
cleavage of the C-N bond linking the succinate moiety of the two
substrates to their nucleotide part, similar to that catalyzed by
the urea cycle enzyme argininosuccinate lyase. ADSL is composed
of four subunits of about 50 kDa. The enzyme of Thermotoga
maritima has been crystallized,24 and,
based on its structure, homology models of the human and Bacillus
subtilis enzyme have been constructed.25,26
++
ADSL deficiency results in the accumulation of succinylaminoimidazole
carboxamide (SAICA) riboside and succinyladenosine (S-Ado) in urine,
cerebrospinal fluid (CSF), and to a minor extent plasma. These succinylpurines,
the hallmarks of the disorder, are the products of the dephosphorylation
by 5'-nucleotidase(s) of SAICAR and adenylosuccinate (S-AMP). In
profoundly retarded type I patients, CSF concentrations of both
succinylpurines are 100 to 200 μmol/l,
and the S-Ado/SAICA-riboside ratios are between 1 and 2.
In ADSL-deficient patients with milder clinical pictures, the CSF
concentrations of SAICA-riboside are in the same range, but those
of S-Ado tend to be higher. This results in S-Ado/SAICA-riboside
ratios around or above 2, even reaching 4 to 5 in the strikingly less
retarded type II patients. In urine, the concentrations of the succinylpurines
can reach up to 5 μmol per mg of creatinine, and
their ratio reflects that in CSF. Comparison of the activities of
ADSL with both S-AMP and SAICAR as substrates showed that both are lost
in parallel, to about 30% of normal in fibroblasts of profoundly
retarded type I patients. In contrast, in fibroblasts of a mildly retarded
type II patient, activity with S-AMP was reduced to 3% of
normal, whereas that with SAICAR was 30% of control. These findings
provide an explanation for the markedly higher S-Ado/SAICA-riboside
ratio in the body fluids of the latter patient.
++
Two main hypotheses can be put forward to explain the symptoms
of ADSL deficiency: impaired synthesis of purine nucleotides and toxic
effects of the accumulating succinylpurines. Although ADSL deficiency
might be expected to lead to decreased synthesis of purine nucleotides,
normal levels of the latter were measured in freeze-clamped liver,
kidney, and muscle of patients. This suggests that residual ADSL
activity remains sufficient to allow flux through the pathway of
purine synthesis, as evidenced by the finding that in fibroblasts
of the mildly retarded patient, in whom ADSL activity with S-AMP
was only 3% of normal, conversion of S-AMP into IMP remained
possible. In addition, the ADSL defect could be circumvented by
supply of purines from nonaffected cell types via purine salvage.
It is therefore more likely that the symptoms of ADSL deficiency
are due to toxic effects of the accumulating succinylpurines. Moreover,
the observation of generally less severe mental retardation of patients
with similar SAICA-riboside levels, but with S-Ado/SAICA-riboside
ratios above 2, suggests that SAICA-riboside is the offending compound and
that S-Ado could protect against its toxic effects.
++
Positron emission tomography revealed a marked decrease in the
uptake of [18F]-labeled 2-fluoro-2-deoxyglucose
in the cortical areas of ADSL-deficient patients, suggesting interference
with glucose metabolism.27 However, the mechanism
of this effect remains unexplained. Owing to the structural analogy
of the succinylpurines with adenosine and N-methyl-D-aspartate
(NMDA), the possibility that they might interfere with the receptors
of these compounds was investigated. Interferences could not be
evidenced, but SAICA-riboside induced a significant degeneration
of the pyramidal neurons of rat hippocampus28 and
was toxic to undifferentiated neurons.29
++
The adenylosuccinate lyase (ADSL) gene has been mapped to chromosome
region 22q13.1-13.2 in humans. The gene is 23 kb in length and comprises
13 exons.30 Correction of a first 1337 nucleotide
sequence31 has shown that the open reading frame
of the human cDNA comprises 1452 nucleotides and encodes a protein that
contains 484 amino acids (E.A. Fon, EMBL database, accession number X65867).30,32 Analysis
of the first family with ADSL deficiency had revealed a T-to-C substitution,
resulting in an S413P change,31 now labeled S438P.30,32 To
date, in accordance with the variability of the clinical symptoms, about
50 missense mutations have been reported in apparently unrelated
sibships13,14 (Adenylosuccinate Lyase Mutation
Database, http://www.icp.ucl.ac.be/adsldb).
In about half of the families, the patients are compound heterozygotes.
The most frequently encountered mutation is R426H, previously identified as
R401H; this accounts for about one third of the alleles investigated.
A splicing error that leads to deletion of amino acids 206-218,32 two
nonsense mutations (R337X30 and R190X), and a mutation
in the 5'UTR of the ADSL gene (-49T/C33),
have also been identified. The latter was found in three unrelated patients,
suggesting that mutation in a regulatory region of the gene might
be an unusually frequent cause of ADSL deficiency.
++
Nearly all mutations identified hitherto are distant from the
fumarate lyase signature of the enzyme (amino acids 288–298,
encoded by nucleotides 862–894) and do not affect the conserved
histidyl 159 residue believed to function in catalysis. Expression
studies of these mutations show that they provoke thermolability
and decrease Vmax of ADSL with its two substrates in parallel.34 Studies
of mutant Bacillus subtilis enzymes corresponding to
human ADSL deficiencies are consistent with these observations.22,25 A
single mutation, R303C, located five amino acids distal from the
fumarate lyase signature and found in two independent, mildly retarded
type II patients, does not affect enzyme stability; however, when
expressed, it displays markedly less activity with S-AMP than with
SAICAR, in accordance with the findings in fibroblasts.
++
The marked clinical heterogeneity of ADSL deficiency justifies
systematic screening for the disorder in unexplained psychomotor
retardation (profound or mild), accompanied or not with autistic
features, and in neurological disease with convulsions or hypotonia.
Diagnosis is based on the presence of SAICA-riboside and succinyl
adenosine (S-Ado) in urine and CSF, which are normally nearly undetectable.
For systematic screening, a modified Bratton-Marshall test, performed
on urine, appears most practical.35 However, false-positive
results may be observed in patients who receive antiepileptic medications
or antibiotics, particularly sulfonamides, because the test was
originally devised for measurement of sufonamides. The succinylpurines
can also be detected by other techniques, including thin-layer chromatography.36 Final
diagnosis requires HPLC with UV spectral analysis12 or proton
NMR spectroscopy.37
++
Although not required for diagnosis, measurements of ADSL’s
activity with S-AMP as substrate show markedly decreased (even undetectable)
activities in liver and kidney of all patients investigated. In
muscle, activity is within normal range in some patients but is
reduced in those presenting with growth retardation and muscular
wasting. In peripheral blood lymphocytes, ADSL activity is about
40% of normal. The activity of erythrocyte ADSL is normal
in some patients but is decreased in others. As a rule, a partial
deficiency of ADSL is measured in cultured fibroblasts.
++
Prenatal diagnosis of ADSL deficiency has been performed twice
on chorion villi of the mother of a patient who had a C5T mutation on
the maternal allele and a C1185A mutation on the paternal allele.38
+++
Treatment and
Prognosis
++
With the aim to replenish hypothetically decreased concentrations
of adenine nucleotides in adenylosuccinate lyase (ADSL)-deficient tissues,
some patients have been treated for several months with oral supplements
of adenine (10 mg/kg per day) and allopurinol (5–10 mg/kg
per day). Adenine can be incorporated into the adenine nucleotides
via adenine phosphoribosyltransferase (APRT), whereas allopurinol
is required to prevent xanthine oxidase from converting adenine
into minimally soluble 2,8-dihydroxyadenine, which forms kidney
stones (see below). No clinical or biochemical improvement was recorded, with
the exception of some acceleration of growth. More recently, oral
administration of ribose (10 mmol/kg per day) has been
reported to reduce seizure frequency in an ADSL-deficient girl,39 although
not in a sustained way. Uridine (2 mmol/kg per day) also
had a slight beneficial effect.40
++
The prognosis for survival of ADSL-deficient patients varies
greatly. Several of those presenting with early epilepsy have died
within the first months of life. In markedly retarded patients,
further evolution is characterized by absent or minimal progression
of psychomotor development and persistence of autistic behavior,
except for occasional improvement of eye contact. Mildly retarded
patients have reached adult age and are able to work in a protected
environment.
+++
Aica-Ribosiduria
(Atic Deficiency)
++
This disorder41 (OMIM 608688), identified on
the de novo pathway of purine synthesis, was diagnosed in a female
infant presenting with profound mental retardation, marked dysmorphic
features (prominent forehead and metopic suture, brachycephaly,
wide mouth with thin upper lip, low-set ears, and prominent clitoris
due to fused labia majora), and congenital blindness. Urine analysis
showed a positive Bratton-Marshall test. This led to the identification
of a massive excretion of 5-amino-4-imidazolecarboxamide (AICA)
riboside, the dephosphorylated counterpart of AICAR (Fig.
168-1), also termed ZMP. Analysis of the patient’s
erythrocytes revealed accumulation of ZMP and of the di- and triphosphates,
ZDP and ZTP, respectively. Incubation of her fibroblasts with AICA-riboside
led to accumulation of AICAR (not observed in control cells), indicating
impairment of the final steps of de novo purine biosynthesis. Assay
of AICAR transformylase/IMP cyclohydrolase (ATIC), the
bifunctional enzyme catalyzing the two last steps of the pathway,
revealed a profound deficiency of the transformylase and a partial deficiency
of the cyclohydrolase. Sequencing of the ATIC gene (gene map locus
2q35) showed a K426R change in the transformylase region in one
allele and a frameshift in the other. The recombinant enzyme carrying
mutation K426R completely lacks AICAR transformylase activity. The
discovery of this novel inborn error of purine synthesis reinforces
the necessity to perform a Bratton-Marshall test in all cases of
unexplained mental retardation or neurological symptoms.
+++
Myoadenylate Deaminase
Deficiency
++
The deficiency of myoadenylate deaminase (OMIM 102770), the muscle-specific
isoenzyme of AMP deaminase (AMPD), was first reported by Fishbein
and colleagues.42 Since then, the defect has been
found in 1% to 2% of the Caucasian population,
but most deficient individuals are asymptomatic.43
+++
Clinical Presentation
++
Two distinct forms of myoadenylate deaminase deficiency are known.
The first form, primary myoadenylate deficiency, was initially detected
in young, otherwise healthy adults; however, later on, a wide variability
was observed with respect to the age (1.5–70 years) of
patient at the onset of symptoms.44,45 The defect
is associated with isolated muscular weakness, fatigue, cramps,
or myalgia following moderate to vigorous exercise, sometimes accompanied
by an increase in serum creatine kinase and minor electromyographic
abnormalities. Rhabdomyolysis and myoglobinuria may also occur.
Muscular wasting or histological abnormalities are usually absent.
The defect was also detected in patients with hypotonia or cardiomyopathy.
Later on, identifying the deficiency in asymptomatic family members
of affected subjects, finding that approximately 2% of
muscle biopsies taken for diagnostic purposes are AMPD deficient,
and studies of the AMPD gene led to the conclusion that the defect
is present in 1% to 2% of the Caucasian population,
most being asymptomatic.
++
The second form of the disorder, secondary myoadenylate deaminase
deficiency, is found in several neuromuscular disorders, including amyotrophic
lateral sclerosis, fascioscapulohumeral myopathy, Kugelberg-Welander
syndrome, polyneuropathies, Werdnig-Hoffmann disease, and myophosphorylase
deficiency. The symptoms of these disorders dominate those of myoadenylate
deaminase deficiency, although some synergism may be observed.46
+++
Metabolic Derangement
++
AMP deaminase catalyzes the conversion of AMP into IMP with liberation
of ammonia. Four isoenzymes of human AMPD are known. Isoenzyme M,
or myoadenylate deaminase, is found only in skeletal muscle; isoenzyme
L predominates in liver and brain; and isoenzymes E1 and E2 are
found in erythrocytes. The muscle enzyme is closely associated with the
contractile apparatus and is highly regulated. Kinetic studies suggest
that myoadenylate deaminase is nearly inactive in resting muscle but
becomes highly active during contraction due to modifications of
the ATP/ADP and GTP/GDP ratios; of the concentrations
of K+, H+, Pi, and creatine phosphate; and of
myofibrillar binding. This increased activity accounts for the production
of ammonia during muscle contraction. AMPD, together with adenylosuccinate
synthetase and adenylosuccinate lyase, forms the purine nucleotide
cycle (eFig. 168.1). Several functions have
been proposed for the purine nucleotide cycle in muscle: (1) removing
AMP formed during exercise to favor the regeneration of ATP from ADP
by myokinase (adenylate kinase); (2) releasing IMP and ammonia,
both stimulators of glycolysis and hence of energy production; (3) producing
fumarate, an intermediate of the citric acid cycle, and hence also
a provider of energy. It has therefore been suggested that the muscle
dysfunction observed in primary myoadenylate deaminase deficiency
is caused by impairment of energy production required for muscle
contraction. Paradoxically, complete and partial myoadenylate deficiency
did not affect intermediates of the citric acid cycle or the cellular
energy charge during exhaustive exercise.47 Studies
in adenylate kinase knockout mice have corroborated that it is not
essential that AMP deaminase operates during intense muscle contraction.48 Together
with the identification of numerous asymptomatic myoadenylate deaminase-deficient
individuals, these findings suggest that myoadenylate deaminase
deficiency is not a primary cause of muscle dysfunction but might
have a triggering or synergistic effect in association with other,
hitherto unidentified disorder(s).
++
++
Three AMPD genes exist: AMPD1, which produces isoenzyme M (myoadenylate
deaminase); AMPD2 isoenzyme L; and AMPD3 isoenzyme E. The AMPD1
gene is located on chromosome 1 in the region p13-p21. Most Caucasian
and African American subjects with documented primary myoadenylate deaminase
deficiency have been found homozygotes for a single mutant AMPD1
allele with C-to-T transitions at nucleotide 34 in exon 2 and at
nucleotide 143 in exon 3. The mutation at nucleotide 34 produces
a nonsense mutation, Q12X, which severely truncates myoadenylate
deaminase and accounts for its lack of activity. The mutation at
nucleotide 143 results in a missense mutation, P48L, which does
not influence enzyme activity. Population studies show that this
mutant allele is found in 10% to 14% of Caucasians.
This accords with the finding that about 2% of diagnostic
muscle biopsies are AMPD deficient and suggests that the mutation
arose in a remote western European ancestor.
++
More recently, other, more rare mutations of the AMPD1 gene have
been identified in myoadenylate deaminase deficient individuals.
A Japanese patient with myopathy was identified as a compound heterozygote
for two missense mutations, producing R388W and R425H changes.49 Ten
Caucasian patients with exercise-induced myalgia were found compound
heterozygotes for the C34-T/C143T mutation and a G468T
mutation, the latter resulting in a Q156H substitution enzyme with
labile catalytic activity.50
++
Interestingly, a protection against heart failure has been repeatedly
documented in heterozygous subjects for mutant AMPD1.51-53 This
could be explained by increased catabolism of AMP to cardioprotective
adenosine rather than to IMP. Nevertheless, a systemic increase
in adenosine following exhaustive exercise could be observed in
homozygous but not in heterozygous subjects for mutant AMPD1.54 Individuals
with a complete but totally asymptomatic deficiency of erythrocyte AMPD
have been identified in Japan, Korea, and Taiwan.55
++
Screening for myoadenylate deaminase deficiency can be performed
by a semi-ischemic exercise test. A sphygmomanometer cuff is inflated
around the upper arm to the mean arterial pressure. Upon squeezing
a hand manometer for 2 minutes, the several-fold elevation of venous
plasma ammonia, recorded in normal subjects, is absent in myoadenylate deaminase
deficiency. Final diagnosis is established by histochemical or biochemical
assay in a muscle biopsy. In the primary form, the activity of AMPD
is mostly below 2% of normal, and as a rule, little or
no immunoprecipitable enzyme is found. In the secondary form, the
residual activity is 2% to 15% of control, appreciable
immunoreactivity is usually present, and progressive declines of AMPD
activity have been documented. AMPD activity is deficient in approximately 2% of
all muscle biopsies taken for a wide variety of indications, making
this defect the most common metabolic disorder of skeletal muscle.
+++
Treatment and
Prognosis
++
Symptomatic myoadenylate deaminase deficient patients may display
a gradual worsening of their symptoms, which may lead to marked
handicap. They should be advised to exercise with caution to prevent
rhabdomyolysis and myoglobinuria. Administration of ribose (2–60
g per day orally in divided doses) has been reported to improve
muscular strength and endurance in some patients56but not
in others. After ribose is converted into PRPP, it could increase
the synthesis of purine nucleotides. In one patient, xylitol, which
is converted into ribose, proved beneficial at the dose of 15 to
20 g/day orally.57
+++
Adenosine Deaminase Deficiency
++
Adenosine deaminase (ADA) catalyzes the deamination of adenosine
and its deoxy counterpart deoxyadenosine. A gross deficiency of ADA
was first reported by Giblett and colleagues58 in
two unrelated children with profound impairment of both cellular
(T-cell) and humoral (B-cell) immunity, known as severe combined
immunodeficiency disease (SCID). Since that time, several
hundred patients with ADA deficiency have been diagnosed. The disorder
is discussed in Chapter 188.
+++
Adenosine Deaminase Superactivity
++
A hereditary, approximately fiftyfold elevation of erythrocyte
ADA has been shown to cause nonspherocytic hemolytic anemia.59 It is
explained by an enhanced catabolism of the adenine nucleotides,
including ATP, owing to the increased activity of ADA.
+++
Purine Nucleoside Phosphorylase Deficiency
++
Their finding of ADA deficiency prompted Giblett and colleagues
to search for other defects of purine and pyrimidine metabolism
in patients with immune disorders. This resulted in the discovery
of purine nucleoside phosphorylase (PNP) deficiency in a child with
an isolated defect of T-cell function.60 The disorder is
much less frequent than ADA deficiency, with about 50 patients reported.
It is discussed in Chapter 188.
+++
Xanthine Oxidase Deficiency
++
The deficiency of xanthine oxidase (also termed xanthine
dehydrogenase and xanthine oxidoreductase,
or XOR), characterized by xanthinuria, was the
first enzyme defect reported in human purine metabolism.61 Originally
described as a benign disorder, it was subsequently found that it
could also occur in combination with a devastating disorder similar
to isolated sulfite oxidase deficiency.62 The combination is
caused by inability to synthesize a molybdenum cofactor common to
three oxidases: XOR, aldehyde oxidase, and sulfite oxidase. Accordingly,
three types of XOR deficiency are now recognized: (1) type
I, isolated XOR deficiency (OMIM 278300); (2) type
II, combined XOR and aldehyde oxidase deficiency (OMIM 603592),
which causes equally benign xanthinuria; and (3) combined
deficiency of XOR, aldehyde, and sulfite oxidase (OMIM 252150),
provoking a lethal disease.
+++
Clinical Presentation
++
Type I and type II XOR deficiency can be completely asymptomatic,
although in about one third of the cases, kidney stones are formed.
Most often, they are not visible on X-ray, and they may appear at
any age and cause a variety of symptoms, including hematuria, renal
colic, and even acute renal failure. Myopathy may be present, precipitated
by strenuous exercise and associated with crystalline xanthine deposits
in muscle. In combined XOR and sulfite oxidase deficiency, the clinical
picture of sulfite oxidase deficiency (which is also found as an
isolated defect63 overrides that of XOR deficiency.
The symptoms include neonatal feeding difficulties and intractable
seizures, myoclonia, increased or decreased muscle tone, eye lens
dislocation, and severe mental retardation.
+++
Metabolic Derangement
++
XOR catalyzes the conversion of hypoxanthine into xanthine and
that of xanthine into uric acid (Fig. 168-1).
Under physiological conditions, the enzyme is a dehydrogenase that
utilizes NAD+ as the electron acceptor and yields NADH.
In several in vitro and in vivo situations, particularly glutathione
depletion and proteolysis, the enzyme converts into an oxidase that
utilizes molecular oxygen as the electron acceptor and produces
superoxide and hydrogen peroxide. This conversion plays a role in
ischemia-reperfusion damage.64 XOR is composed
of two subunits, each of which has a molybdenum center that contains
molybdenum under the form of a molybdenum cofactor.
++
The deficiency of XOR results in the near total replacement of
uric acid by hypoxanthine and xanthine as the end products of purine
catabolism (Fig. 168-1). This alteration
is clearly manifested in urine, in which over 90% of oxypurine
excretion is normally uric acid. Generally, plasma hypoxanthine
is not elevated (or is minimally elevated) in the various forms of
XOR deficiency, owing to efficient reutilization by hypoxanthine-guanine
phosphoribosyltransferase. Plasma xanthine, normally below 1 μM,
may rise to 10 to 40 μM, which is way below the
normal uric acid level because of the markedly higher renal clearance of
xanthine. Concentrations of hypoxanthine and xanthine are approximately
10 times higher than normal in muscle of xanthinuric subjects. The
limited solubility of xanthine may explain the numerous birefringent
crystals observed in the muscles of patients complaining of pain
and stiffness. The role of hypoxanthine, which increases during
vigorous exercise in both normal and xanthinuric subjects, is unclear,
since it is much more soluble than xanthine.
++
The inheritance of type I and type II XOR deficiency and of combined
XOR and sulfite oxidase deficiency is autosomal recessive. Studies
of the XOR gene, localized on chromosome 2p23-p22, have led to the
identification in hereditary xanthinuria type I of two mutations,
resulting respectively, in a nonsense substitution and a termination
codon.65 Xanthinuria type II is caused by mutations
of a molybdenum cofactor sulfurase gene.66,67More
than 30 different mutations in three different molybdenum cofactor
biosynthetic genes (MOCS1, MOCS2, and GEPH) have been identified
in combined XOR and sulfite oxidase deficiency.68
++
Both in isolated and combined XOR deficiency, plasma concentrations
of uric acid below 1 mg/dl are measured; they may decrease
to virtually undetectable values on a low-purine diet. Urinary uric
acid is reduced to a few percent of normal and is replaced by hypoxanthine
and xanthine. In combined XOR and aldehyde oxidase deficiency (xanthinuria
type II), patients are unable to oxidize allopurinol. In combined
XOR, aldehyde, and sulfite oxidase deficiency, the urinary changes
of xanthinuria are accompanied by an excessive excretion of sulfite
and other sulfur-containing metabolites, such as S-sulfocysteine,
thiosulfate, and taurine. The enzymatic diagnosis requires liver
or small intestine mucosa, the only human tissues that normally
contain substantial amounts of xanthine oxidase. Sulfite oxidase
and the molybdenum cofactor can be assayed in liver and fibroblasts.
Mutant XOR has been investigated in a few patients. Despite immeasurably
low activity, immunoreactive protein remains detectable in the duodenal
mucosa of some patients but not of others.6
+++
Treatment and
Prognosis
++
Isolated XOR deficiency is mostly benign. A low purine diet should
be prescribed and fluid intake increased. The prognosis of combined XOR
and sulfite oxidase deficiency is very poor. So far, all therapeutic
attempts, including low-sulfur diets, the administration of sulfate
and molybdenum, and trials to bind sulfite with thiol-containing
drugs, have been unsuccessful.
+++
Hypoxanthine-Guanine Phosphoribosyltransferase
Deficiency
++
In 1967, Seegmiller and coworkers discovered a complete deficiency
of hypoxanthine-guanine phosphoribosyltransferase (HPRT or HGPRT)
in children with the Lesch-Nyhan syndrome (OMIM 300322), an incapacitating neurological
disorder that, with few exceptions, is limited to males.69 It
is characterized by choreoathetosis, spasticity, compulsive self-mutilation
manifested by biting away lips and tongue and ends of the fingers,
and three- to fourfold normal uric acid levels in plasma and urine.70 That
same year, a partial deficiency of HPRT was described in patients
with milder presentations, sometimes termed Kelley-Seegmiller
syndrome or now more often called Lesch-Nyhan variants (OMIM 300323);
this disorder involves overproduction of uric acid that leads to
severe gouty arthritis in early adult life and involves minor or no
neurological dysfunction.71 This led to the identification
of a continuous spectrum of HPRT deficiencies, ranging from gout
to a devastating neurological syndrome.72
+++
Clinical Presentation
++
At birth, children with complete HPRT deficiency (Lesch-Nyhan
disease) appear entirely normal. The first indication of the disease
may be the presence of brownish to red-orange sand in the diapers,
particularly when the baby becomes dehydrated. Some of the infants
are very irritable, with episodes of screaming, suggesting the possibility
of renal colic. Most infants show normal development during the first
4 to 6 months of life. During the second part of the first year,
impaired motor development becomes evident by the inability to support
the head, generalized muscle hypotonia, and voluntary movements
of both athetoid and choreiform types with spasticity. Recently,
the motor syndrome of complete HPRT deficiency has been reclassified
as a severe action dystonia, superimposed to a baseline hypotonia.73 By
1 year of age, pyramidal symptoms consist of an increase in muscle
tone, an increase in deep tendon reflexes, a sustained ankle clonus, and
a scissoring of the lower extremities, and extensor plantar responses
are usually present. An increased incidence of dislocation of the hips
and club feet may be related to the early hypotonia. Severely affected
children are never able to walk.
++
Over the years, the patients develop a compulsive aggressiveness
and self-mutilation that involves biting away lips, tongue, and ends
of the fingers if given the opportunity. Because it is painful,
children are fearful of the biting. Biting can, in some cases, begin
with the eruption of incisors. In other patients, it may be delayed
until early adolescence. In any given patient, biting can be highly
variable, with patients going through periods when they show extensive
self-mutilation and other periods when this is no longer a problem.
Self-mutilation tends to be correlated, at least in some cases,
with emotional stress. In addition, some older children develop
opisthotonic spasms, which appear to be at least semivoluntary.
An accompanying laryngeal spasm and stridor sometimes produces a
temporary cyanosis. If the patient’s head is in range of
a hard object at the time of a spasm, he or she may injure it. Children
also sometimes throw themselves from the bed if left unattended
or may injure themselves on sharp edges of wheelchairs that are
left unpadded.
++
Aggressive acts against others are also included in the bizarre
behavior of these children. This can take the form of biting, hitting, spitting,
or kicking. Eyeglasses are common targets for their aggression.
Patients often pinch or strike attendants in areas of sexual significance,
and become verbally aggressive, often with a remarkably shocking
vocabulary. Projectile vomiting is also used by older children as
a weapon, especially when the child becomes upset emotionally. Although
this peculiar behavior would seem to alienate them from others,
these children are often favorite patients of ward personnel, because
they are charming and smile and laugh easily. They are very responsive
to their environment and show a remarkably good sense of humor.
They have a characteristic dysarthric speech, although they can
usually make themselves understood to those who are caring for them.
++
The first descriptions of Lesch-Nyhan syndrome included mental
retardation. However, reevaluation with specific tests for their
motor difficulties has shown attention deficits but preservation
of nonverbal intelligence, and some patients display normal intelligence.74-76
++
Partial HPRT deficiency is found in rare patients with gout.
Most of them or normal on neurological examination, but occasionally spasticity,
dysarthria, and a spinocerebellar syndrome are found.77 Some
but not all of the patients with less severe deficiencies of the HPRT
enzyme with resulting minimal neurological dysfunction also show
a compulsive behavior. Whereas most subjects with Lesch-Nyhan syndrome
do not develop gouty arthritis, this finding is common in partial
HPRT deficiency.
+++
Metabolic Derangement
++
HPRT is normally expressed in all cells of the body. Early studies
reported that its specific activity is highest in the basal ganglia,
which correlates with the brain area that can cause analogous movement
disorders. However, later investigations have not confirmed these findings.
HPRT activity is also high in testes, which accords with failure
of sexual maturation and atrophic testes found in some patients with
the most severe enzyme deficit. Alterations in the kinetic properties
of the enzymes in patients comprise, besides reduction of the maximal
velocity up to its complete loss, reduced affinities for phosphoribosyl
pyrophosphate (PRPP) and the purine bases, which have been shown
to correlate with the gene lesions (see below).
++
The considerably excessive production of uric acid in HPRT deficiency
is explained by an enhancement of the purine synthesis rate, which
is caused by the increased amount of PRPP that accumulates from
its underutilization (caused by the HPRT mutation). Fibroblasts
cultured from affected patients show a two- to threefold accumulation
of PRPP, and their erythrocytes show a tenfold accumulation over
normal values. Since PRPP is a rate-limiting, normally not saturating
substrate for the first, presumably rate-determining reaction of
purine biosynthesis de novo catalyzed by the enzyme PRPP glutamine
amidotransferase, its elevation will increase uric acid production.
Further support for this concept comes from the correlation of increased
intracellular PRPP in fibroblasts and excessive rates of purine
synthesis in PRPP synthetase superactivity (see above).
++
Theoretically, the deficiency of HPRT should provoke accumulation
of its substrates (hypoxanthine, guanine, and its deamination product
xanthine) and of PRPP and depletion of its products (GMP and the
other guanine nucleotides and IMP). Owing to the conversion of IMP
into AMP (Fig. 168-1), the synthesis of the
latter and the other adenine nucleotides could also be decreased.
Hypoxanthine and xanthine are elevated in body fluids of HPRT-deficient
patients, but their toxicity is ruled out by the absence of neurological
symptoms in isolated xanthine oxidase deficiency (see above) and
the absence of worsening neurological symptoms upon allopurinol
treatment. Because allopurinol lowers uric acid but does not influence
the neurobehavioral abnormalities, uric acid toxicity is ruled out.
PRPP toxicity is also unlikely, since its elevation in PRPP synthetase superactivity
(see above) is not associated with the neurological symptoms of
HPRT deficiency. Similarly, accumulation of AICAR, an intermediate
of de novo purine biosynthesis, and its further phosphorylation
products (collectively termed Z-nucleotides) is
unlikely to be involved, since it has also been recorded in disorders
without neurobehavioral abnormalities.
++
Depletion of guanine or adenine nucleotides is also proposed
as an explanation for the neurological symptoms of HPRT deficiency.
ATP depletion might result in impairment of energy production, DNA
repair,78 and metabolic defense against oxidant
stress. GTP depletion, which can be shown in HPRT-deficient cells
under certain conditions,79 has been proposed to
interfere with the function of G proteins and the formation of tetrahydrobiopterin
(BH4), a cofactor in dopamine synthesis. However, other studies80 have invalidated
the latter hypothesis, which might have provided an explanation
for the abnormalities of the dopaminergic system observed in Lesch-Nyhan
patients.
++
Multiple studies have shown abnormalities of the dopaminergic
neurotransmitter system in Lesch-Nyhan disease. Autopsy studies81,82 showed
a reduction in the basal ganglia (to 10–30% of
control) of dopamine; its metabolite homovanillic acid; and tyrosine
hydroxylase, the dopamine synthetizing, BH4-dependent enzyme. In
vivo PET studies of the brains of Lesch-Nyhan subjects have revealed large
reductions in dopaminergic terminals not only in the basal ganglia,
but also in other brain areas.83,84 Dopamine deficiency
is also found in HPRT knockout mice.85 How HPRT deficiency
impairs the dopaminergic system remains an open question. One possibility could
be an increased sensitivity to dopamine-associated oxidant stress,
as thought to play a role in Parkinson disease. Dopamine and other catecholamines
are toxic to many cell types, probably through various mechanisms,
including spontaneous and metal-catalyzed oxidation that produces
reactive oxidant species such as superoxide radicals.86 Studies
in HPRT knockout mice provide evidence for an increased oxidant
stress, which is not affected by overexpression of superoxide dismutase
indicating that other mechanisms intervene.85
++
The human hypoxanthine-guanine phosphoribosyltransferase (HPRT)
gene spans approximately 45 kb on the long arm of chromosome X at
Xq26-q27.2 and contains nine exons.87 It encodes
a tetrameric protein composed of four identical subunits of 219
amino acids with a mass of 25 kDa. The three-dimensional structure
of the human enzyme has been resolved by X-ray crystallography.88 A
single restriction fragment length polymorphism with three alleles
is known. A rare histidine-to-arginine mutation at codon 60 reduces
HPRT activity by approximately 40% but does not provoke symptoms89 and
might thus be a rare polymorphism. The HPRT gene on the inactivated
X-chromosome is rendered transcriptionally silent by methylation
of the majority of its CpG clusters.90
++
Over 250 mutations causing HPRT deficiency have been reported,91 and
new mutations are regularly identified.92,93 Although with
some overlap, they can be divided into three categories, based on
clinical phenotype: (1) full-fledged Lesch-Nyhan disease; (2) intermediate
with some neurological dysfunction but no self-injurious behavior;
and (3) mild, with uric acid overproduction without neurological
symptoms. As reviewed by Jinnah and colleagues,91 single
base substitutions account for approximately 65% of all
mutations; deletions account for 25%; and insertions, duplications,
and complex mutations account for the remaining 10%. About
60% of the point mutations causing amino acid substitutions,
nearly all those causing a premature stop, and 80% of those
leading to splicing errors provoke Lesch-Nyhan disease. As expected,
90% of intermediate and mild cases have point mutations
leading to amino acid substitutions. Mutations are found throughout
the HPRT gene, but several hot spots seem to exist. Although some
mutations that alter HPRT kinetics implicate the binding site of
its substrates, most are located outside, which suggests that they
provoke conformational changes in the active site. Overall, location
of the HPRT mutations seems to have a less significant influence
on phenotype than the different amino acid changes that can be found
at the same codon. Structural and functional analysis with modeling
methods has also shown that most mutations can be explained by the
predicted effect on protein structure and function.94 With
rare exceptions,95 genotype/phenotype
correlations are thus quite apparent in HPRT deficiency.
++
Although Lesch-Nyhan disease is an X-linked disorder, a small
number of females with the disorder have been identified.96–99 This
can be explained by the coexistence of a nonsense mutation or deletion
on one of the parents’ alleles, with a nonrandom inactivation
of the other parent’s allele.
++
The excessive excretion of uric acid forms the basis for a relatively
simple screening test in which the ratio of uric acid to creatinine
(both expressed as mg/100 ml in the morning urine sample)
is measured.100 Lesch-Nyhan patients with the full
syndrome show ratios between 2.0 to 5.3, as opposed to 0.2 to 0.6
for normal individuals. However, some overlap with the normal may
be found in the first few months or years of life because of high
ratios observed at birth in normal infants, followed by a marked
age-related decline. Although most patients also show an elevation
of serum urate—up to 18 mg/dl (1 mmol/l),
particularly in later stages of the disease—this measurement
cannot be used to rule out Lesch-Nyhan disease, as around 5% to
10 % may show a normal serum urate, particularly in early
life.
++
Measuring HPRT is most easily performed in red blood cells or
skin fibroblasts. In patients with the most severe enzyme defect, HPRT
activity is virtually absent in erythrocyte lysates. In patients
with less severe deficiencies, HPRT activity may range from less than
0.1 to 10% to 20% of normal. In rare kinetic mutations
(see below), near-normal activities can be measured when saturating substrate
concentrations are used.
++
Carrier detection of Lesch-Nyhan disease has long been accomplished,
most practically by assaying individual hair follicles, which are advantageous
because they are largely clonal in females. Prenatal diagnosis of
Lesch-Nyhan disease can be made by measuring HPRT activity in cultured
amniocytes and chorionic villi. These techniques are now largely
supplanted by molecular diagnosis in families in which the mutation
is known.101
+++
Treatment and
Prognosis
++
Treatment comprises controlling the uric acid overproduction,
the motor syndrome, and the behavioral manifestations. Allopurinol,
at doses up to 20 mg/kg of body weight per day, prevents
the damage to kidneys caused by the excessive amounts of uric acid
excreted in the urine. Although it produces a marked decrease in
both urine and serum uric acid, it fails to decrease de novo purine
synthesis: Hypoxanthine and xanthine merely replace uric acid in urine.
Established renal calculi can thereby be substantially reduced in
size. Further general measures for treatment are those of gout,
including high fluid intake (at least 50 ml/kg per day).
This will diminish the occurrence of urinary concretions composed
of xanthine that have been noted occasionally in children treated
with allopurinol.8 Adequate nutrition should also
be prescribed, since many of these children take a very long time
to eat and may actually be malnourished in an understaffed institution.
++
Although children with the Lesch-Nyhan syndrome seem to be incapable
of learning from punishment, they do respond to positive experiences,
and some success in behavior modification has been achieved by simply turning
away from the child when they display aberrant behavior. Partial
improvement in self-mutilation has also been reported from using
both positive and negative conditioning programs. Patients should
also be made more comfortable by appropriate restraints. Hands can
be kept away from the mouth and yet be left free for use by constructing
loose-fitting wraparound fabric splints for the elbows. Chronic
irritation in the mouth from sharp teeth edges can be eliminated
by lip guards and even tooth extraction.
++
Despite trials with a variety of agents, no successful pharmacological
treatment of the motor syndrome and the behavioral manifestations
has yet been found. Children treated with diazepam (Valium) are
less spastic. Levodopa with or without a decarboxylase inhibitor
has been reported to improve, worsen, or not influence the dystonia.
Hydroxytryptophan has been reported to reduce self-mutilation but
only transiently and is hardly used anymore. Risperdal and carbamazepine
may be useful. Gabapentin has been suggested to improve self-injurious
behavior.102 Disappearance of self-mutilating behavior
with bilateral chronic stimulation of the globus pallidus internus
has been reported.103,104 Recently, local injection
of botulinum toxin has also been advocated.105 Lesch-Nyhan
disease was one of the first disorders proposed for gene therapy
and continues to serve as a model for its development. Bone marrow
transplantation has been performed in a small number of Lesch-Nyhan
patients without obvious benefit and with significant morbidity,
since the majority of the patients died shortly thereafter.
+++
Adenine Phosphoribosyltransferase
Deficiency
++
The deficiency of adenine phosphoribosyltransferase (APRT; OMIM
102600) was first identified in heterozygotes with a partial enzyme defect.106 Later,
Cartier and Hamet107 reported a homozygous 4-year-old
with urinary crystals and stones. Approximately 300 patients have been
diagnosed worldwide, but up to 50% of APRT-deficient subjects
may be asymptomatic.
+++
Clinical Presentation
++
The deficiency may clinically manifest in childhood, even from
birth, but may also remain silent for several decades.108 Symptoms include
urinary passage of gravel, small stones, and crystals, frequently
accompanied by abdominal colic, dysuria, hematuria, and urinary
tract infection. Some patients may present with acute anuric renal
failure, while others have developed chronic renal failure requiring
dialysis and transplantation. The urinary precipitates are composed
of 2,8-dihydroxyadenine (2,8-DHA) and radiotranslucent.
+++
Metabolic Derangement
++
The APRT deficiency results in the loss of adenine salvage (Fig. 168-1), provided by food and by the
polyamine pathway. Consequently, adenine is oxidized by xanthine
oxidase into 2,8-DHA. Symptoms are caused by the very poor solubility
of 2,8-DHA. Its solubility in urine, at pH 5 and 37°C, is about
0.3 mg/dl, considerably less than that of 15 mg/dl
for uric acid.
++
Two types of APRT deficiency are known. Patients with type I
deficiency have no detectable activity in erythrocyte lysates. In
patients with type II deficiency, significant residual activity
is found, reaching 5% to 25% of normal when measurements
are made at supraphysiological, saturating concentrations of PRPP. However,
kinetic studies reveal that the S0.5 value for PRPP, which
is about 3 μM for the normal enzyme, is increased
to 50 to 80 μM. This decreased affinity for PRPP
results in near inactivity under physiological conditions. Consequently,
APRT activity is not detectable in intact cells such as erythrocytes
or fibroblasts. Confirmatory diagnosis of type II deficiency is
performed on viable T cells.109 To date, type II
patients have been found only in Japan, where they account for approximately 80% of
the affected subjects.110 Determining the three-dimensional
structure of human APRT111 has provided insights
in the mechanisms by which many mutations hamper normal enzyme activity.
++
The human adenine phosphoribosyltransferase gene has been mapped
to chromosome 16q24.3. Both in Caucasian and Japanese subjects,
numerous polymorphisms are observed. In type I deficiency, more
than 30 mutations have been identified.108 About
half of these are single base changes. T insertion mutations, resulting
in frameshift, and splicing errors are also frequent. The observation
of several more common mutations suggests founder effects. All the
type II Japanese patients carry the same c.2069T- > C substitution
in exon 5, resulting in an M136T change. Approximately 80% are
homozygous. Another W98X nonsense mutation, and a CCGA insertion
resulting in a frameshift, account for nearly all the other cases.112
++
The presence of brownish spots on diapers or the finding of round,
brownish crystals in urine under a light microscope suggest the presence
of 2,8-DHA. Notwithstanding its very low solubility, 2,8-DHA can
also be identified by HPLC. Confirming its presence requires complex
analyses, including UV and infrared spectrography, mass spectrometry,
X-ray crystallography, and capillary electrophoresis. It is therefore
usually easier to measure adenine phosphoribosyltransferase (APRT)
activity. As said under “Metabolic Derangement,” based
on the level of residual enzyme activity in red blood cell lysates,
two types of APRT deficiency are recognized. In both types, APRT
activity is absent in intact cells.
+++
Treatment and
Prognosis
++
In patients with symptoms, allopurinol should be given (as detailed
under treatment of PRPP synthetase superactivity) to inhibit the
formation of 2,8-DHA. Both in patients with stones and in those
without symptoms, dietary purine restriction and high fluid intake
are recommended. Alkalinization of the urine is, however, not advised:
unlike that of uric acid, the solubility of 2,8-DHA does not increase
up to pH 9. Shock-wave lithotripsy has been beneficial in a small
number of patients.
++
Ultimate prognosis depends on renal function at the time of diagnosis:
late recognition may result in irreversible renal insufficiency requiring
chronic dialysis, and early treatment may result in prevention of
stones. Kidney transplantation has been followed by recurrence of
microcrystalline deposits and subsequent loss of graft function.
+++
Deoxyguanosine Kinase
Deficiency
++
In three large consanguineous kindreds of Druze origin with the
hepatocerebral form of mitochondrial DNA depletion syndrome (characterized
by early progressive liver failure, neurological abnormalities,
hypoglycemia, and increased lactate), homozygosity mapping was used
to search for the causative gene defect.113 It
led to the identification of a mitochondrial deoxyguanosine kinase
deficiency. This enzyme phosphorylates the deoxy counterpart of
guanosine (Fig. 168-1) into deoxy GMP. The
disorder is discussed in Chapter 176.
+++
Thiopurine Methyltransferase Deficiency
++
Thiopurine S-methyltransferase (TPMT) catalyzes the S-methylation
of several synthetic pharmacological purine analogs that contain
a thiol group such as 6-mercaptopurine, 6-thioguanine, and azathioprine
that is converted to 6-mercaptopurine in vivo. These drugs are used
to treat various diseases, including cancers, rheumatoid arthritis,
and other autoimmune disorders, and immunosuppressants after organ
transplantation. They are converted via phosphoribosylation by hypoxanthine-guanine
phosphoribosyltransferase and adenine phosphoribosyltransferase into
active thionucleotides, which exert their therapeutic action by
incorporation into DNA and RNA. Their oxidation by xanthine oxidase
and S-methylation by TPMT results in inactivation.
++
The wide variations in therapeutic response and occurrence of
toxic side effects in some patients receiving thiopurines led to
the identification of TPMT as a determining factor in this variability.114,115Approximately
90% of individuals in various ethnic populations have high TPMT
activity, about 10% have intermediate activity, and 1 in
300 lack activity. Patients with no or less efficient methylation
of thiopurines have more extensive conversion to active thionucleotides
that leads to severe, potentially fatal myelosuppression. Therefore,
determining the TPMT status prior to treatment with thiopurines
is now recommended in predictive pharmacogenetics.116
+++
Dihydropyrimidine Dehydrogenase Deficiency
++
The deficiency of dihydropyrimidine dehydrogenase (DPD; OMIM
274270) was first identified in a patient with developmental problems and
overexcretion of uracil and thymine.118 Later on,
the defect was also found in subjects with very variable clinical
symptoms, including asymptomatic relatives of deficient patients, and
in subjects presenting with toxic reactions to 5-fluorouracil.119 This
imparted some doubt on the causal relationship between the deficiency,
which might be quite common, and the symptoms.
+++
Clinical Presentation
++
Two forms of DPD deficiency occur. The first is found in children.
Approximately half of these display epilepsy, motor retardation,
and mental retardation, often accompanied by generalized hypertonia;
hyperreflexia; growth retardation; dysmorphic features, including microcephaly;
ocular abnormalities; and autistic features.120,121In
these patients, the deficiency of DPD is complete or nearly complete. Nevertheless,
the severity of the disorder is highly variable, and even asymptomatic
cases have been identified. The second clinical picture is found
in adults who receive the pyrimidine analog 5-fluorouracil, a classic
treatment of various cancers, including breast, ovary, and colon.122 It
is characterized by severe life-threatening toxicity, manifested
by profound neutropenia; stomatitis; diarrhea; and neurological
symptoms, including ataxia, paralysis, and stupor. In these patients,
DPD deficiency is as a rule partial and revealed only by 5-fluorouracil
therapy.
+++
Metabolic Derangement
++
DPD catalyzes the first step in the degradation of the pyrimidine
bases uracil and thymine, namely their conversion into dihydrouracil and
dihydrothymine, respectively (Fig. 168-3).
Why a profound DPD deficiency becomes manifest in some pediatric
patients but not in others remains an open question and suggests that
other factors contribute to the clinical manifestations of the disorder.
How the defect leads to neurological symptoms also remains elusive,
but reducing the concentration of β-alanine, a
neuromodulator that can block the reuptake of GABA, may play a role.
A depletion of β-aminoisobutyrate may also play
a role. The marked potentiation of the action and thus the toxicity
of the anticancer drug 5-fluorouracil, is explained by an impairment
of the inactivation of this pyrimidine analog via DPD, which normally
accounts for approximately 80% of its catabolism.
++
The human dihydropyrimidine dehydrogenase (DPD) gene has been
mapped to chromosome 1p22. About 20 mutations have been identified
in DPD deficiency.123–126 Most frequent (approximately
50%) is a G to A mutation in the GT5' splicing recognition
sequence of intron 14, IVS14+1G>A (DPYD*2A), which results
in a 165-bp deletion corresponding to exon 14 in the DPD mRNA. Strikingly,
patients who carry the same mutation may display widely variable
clinical symptoms. About 2% of the Dutch population was
found heterozygous for the IVS14+1G>A mutation,127 and
approximately 25% of the patients with the adult form of
DPD deficiency, characterized by 5'-fluorouracil toxicity, are heterozygotes
for this mutation.122 A threefold higher prevalence
of DPD deficiency has been reported in African Americans as compared with
Caucasians.128 The crystal structure of recombinant
pig DPD has been resolved.129 Analysis of the DPD
crystal structure indicates that several mutations interfere with
cofactor binding or electron transport or destabilize the protein.125
++
The pyrimidine catabolites can be detected by various techniques,
including HPLC, GC-MS, analysis of amino acids in urine before and
after acid hydrolysis, and proton NMR spectroscopy.37 Patients
excrete high amounts of uracil (56–683 mmol/mol
creatinine, as compared to 3–33 in control urine) and thymine
(7–439 mmol/mol creatinine, as compared to 0–4
in control urine). Elevations of uracil and thymine in plasma and
cerebrospinal fluid are also recorded.120 Excretion
of both compounds may be less elevated in patients with high residual
DPD activity.
++
The enzyme defect can be demonstrated in the patient’s
fibroblasts, liver, and blood cells, with the exception of erythrocytes.120,122 In pediatric
patients, DPD deficiency is complete or nearly complete; in the
adult cancer patients experiencing acute 5-fluorouracil toxicity,
it is partial, with residual enzyme activities ranging from 3% to
30%.
+++
Treatment
and Prognosis
++
No treatment is available for pediatric patients. Symptoms usually
remain the same, but death in early infancy of a more severely affected
child has been reported. In the adult cancer patients, discontinuation
of 5-fluorouracil results in slow resolution of the toxic symptoms.119,123 Pharmacogenetic
testing of patients before administration of 5-fluorouracil should
be considered.
+++
Dihydropyrimidinase Deficiency
++
Deficiency of dihydropyrimidinase (DHP; OMIM 222748) was first
reported in a single male baby of consanguineous parents, presenting
with convulsions and metabolic acidosis.130 Several
other patients have been diagnosed since then.120,121,131 As
in dihydropyrimidine dehydrogenase (DPD) deficiency, completely
asymptomatic individuals have been identified,132 including
siblings of deficient patients.131
++
In children, complete or nearly complete DHP deficiency presents
features similar to those of PDP deficiency described above.
+++
Clinical Presentation
++
As with DPD deficiency, the clinical picture is very variable
and may include severe psychomotor retardation with epilepsy, delay
in speech development, growth retardation, dysmorphic features including
microcephaly, and white matter abnormalities. Increased sensitivity
to 5-fluorouracil, leading to severe toxicity, has also been reported.133
+++
Metabolic Derangement
++
DHP catalyzes the second step in the degradation of the pyrimidine
bases uracil and thymine, namely the cleavage of dihydrouracil and dihydrothymine
into β-ureidopropionate and β-ureidoisobutyrate,
respectively (Fig. 168-3). Its deficiency
provokes accumulation of dihydrouracil and dihydrothymine. The reasons
for the appearance and the mechanisms of the symptoms remain unexplained.
As in DPD deficiency, reduced concentrations of the neuromodulator β-alanine
may play a role. Also as in DPD deficiency, the increased sensitivity
to 5-fluorouracil is explained by DHP’s role in the catabolism
of this pyrimidine analog.
++
Studies of the DHP gene, localized on chromosome 8q22, have led
to the identification of various mutations in symptomatic and asymptomatic
individuals.131,134 Enzyme expression showed no
significant difference in residual activity between the mutations
of the symptomatic and the asymptomatic individuals.
++
Urinary dihydrouracil and dihydrothymine, normally found in small
amounts, may reach 400 to 600 mmol/mol creatinine.120 They
can be measured by the techniques listed above for uracil and thymine
in DPD deficiency, with the exception of HPLC analysis with UV detection.
Indeed, since dihydropyrimidinase (DHP) catalyzes the opening of
the pyrimidine ring, its products, dihydrouracil and dihydrothymine,
are not UV-detectable. Nevertheless, moderate (two to twentyfold)
elevations of uracil and thymine are also found in the urine of
DHP-deficient individuals and may provide a diagnostic clue. Enzyme
assay requires liver biopsy, since more accessible tissues do not
possess DHP activity.120
+++
Treatment and
Prognosis
++
There is no therapy, and prognosis seems unpredictable. The first
reported patient recovered completely and apparently displays normal
physical and mental development.130 In contrast,
another patient had a progressive neurodegenerative clinical course.135
+++
Ureidopropionase Deficiency
++
Ureidopropionase deficiency (OMIM 606673) was first identified
in a 17-month-old girl of consanguineous parents. She presented with
muscle hypotonia, dystonic movements, and severe developmental delay.136 To
date, less than 10 patients have been diagnosed.122,137,138
+++
Clinical Presentation
++
Symptoms of the disorder are widely variable. They include early
onset dystonic movement disorder, severe developmental delay with marked
impairment of visual responsiveness combined with severely delayed
myelination, optic atrophy, pigmentary retinopathy, and cerebellar
hypoplasia.139 Another patient presented at 4 months
of age with an acute life-threatening febrile status epilepticus.140 At
11 months, he showed severe psychomotor retardation with muscular
hypotonia, extremely limited visual contact, and poorly controlled epilepsy.
MRI revealed large subdural hematomata and global supratentorial
atrophy. Another patient showed congenital urogenital and colorectal
anomalies but reached normal developmental milestones.138
+++
Metabolic Derangement
++
Ureidopropionase (also termed β-alanine
synthase) catalyzes the last step of the pyrimidine degradative
pathway, the conversion of β-ureidopropionate and β-ureidoisobutyrate
into β-alanine and β-aminoisobutyrate,
respectively (Fig. 168-3). Its deficiency
provokes elevations of ureidopropionic acid (also called ββ) and
ureidoisobutyric acid (also called ββ). Studies
with cultured chick neurons suggest that β-ureidopropionate
may act as a neurotoxin by inhibiting mitochondrial energy metabolism.141 Also,
as in DPD and DHP deficiency, a reduction of the concentration of β-alanine
may play a role.
++
Genetic analysis of the human β-ureidopropionase
gene, mapped to chromosome 22q11.2, has revealed the presence of
two splice-site and one missense mutation in the first four patients
with β-ureidopropionase deficiency.142
++
As with dihydrouracil and dihydrothymine, ureidopropionic acid
and ureidoisobutyric acid are not UV-detectable but can be easily detected
with HPLC-electrospray tandem mass spectrometry.137 Assay
of β-ureidopropionase in liver confirms the diagnosis.122,137
+++
Treatment and
Prognosis
++
A therapeutic trial with β-alanine over 1.5 years
has shown no convincing clinical improvement.139
+++
Pyrimidine 5′-Nucleotidase Deficiency
++
Purine and pyrimidine nucleoside monophosphates are catabolized
to their corresponding nucleosides (Figs. 168-1 and 168-3) by a variety of cytosolic and membrane
5′-nucleotidases that may be more specific
for one or another type of nucleoside monophosphate. Several pyrimidine 5′-nucleotidases
(P5N) exist that dephosphorylate CMP, UMP, their deoxy counterparts,
and TMP to the corresponding nucleosides (Fig. 168-3).
The deficiency of one these, P5N-1, a 52 kDa an isozyme restricted
to erythrocytes that catalyzes the dephosphorylation of UMP and CMP
but not of TMP, was identified in patients with severe chronic hemolytic
anemia.143
+++
Cytosolic 5′-Nucleotidase Superactivity
++
This disorder was first identified in the cultured skin fibroblasts
of a 3-year-old girl with frequent infections, seizures, developmental delay,
severe language deficit, hyperactivity with short attention span,
and poor interaction with other children.144 Later
on, three additional unrelated patients were diagnosed,145 and
at least five other patients have reportedly been identified. Routine
laboratory tests were unremarkable except for persistent hypouricosuria.
Biochemical studies in the patients’ fibroblasts showed
six- to tenfold elevations of the activity of cytosolic 5′-nucleotidase,
measured either with a pyrimidine (UMP) or a purine (AMP) as substrate. Electrophoretical
abnormalities were also observed.146 The hypouricosuria
accompanying the syndrome has been tentatively explained by a 30% to
50% lower concentration of PRPP, as found in patients’ fibroblasts;
this is caused by increased salvage of the purine bases derived
from accelerated nucleotide catabolism. Decreased PRPP would result
in decreased de novo purine synthesis and hence of uric acid production. Based
on the possibility that increased catabolism might cause a deficiency
of pyrimidine nucleotides, the patients were treated with oral uridine
at the dose of 200 to 1000 mg/kg per day. Remarkable developmental
improvement and a decrease in frequency of seizures and infections
were recorded. Upon replacement of uridine by placebo, all patients rapidly
reverted to their pretreatment states.
+++
Thymidine Phosphorylase Deficiency
++
Thymidine phosphorylase (TP) catalyzes the phosphorolytic cleavage
of thymidine (eFig. 168.1 and Fig.
168-3) and deoxyuridine to thymine and uracil, respectively.
Deficiency of this enzyme was found147 to cause
mitochondrial neurogastrointestinal encephalomyopathy (MNGIE), an
autosomal recessive disease associated with multiple deletions and
depletion of skeletal muscle mitochondrial DNA (mtDNA). MNGIE is
characterized by ptosis, progressive external ophthalmoplegia, severe
gastrointestinal dysmotility, cachexia, myopathy, peripheral neuropathy,
and lactic acidosis.