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JIMD Rep. 2015; 15: 13–27.
Published online 2014 Apr 10. doi: 10.1007/8904_2014_293
PMCID: PMC4270867
PMID: 24718837

Thiamine-Responsive and Non-responsive Patients with PDHC-E1 Deficiency: A Retrospective Assessment

Sanne van Dongen, Ruth M. Brown, Garry K. Brown, David R. Thorburn, and Avihu Bonehcorresponding author
Metabolic Research, Murdoch Children’s Research Institute, Royal Children’s Hospital, Flemington Road, Parkville, Melbourne, VIC 3052 Australia
Nijmegen Medical Centre, Institute for Genetic and Metabolic Disease, Radboud University, Nijmegen, The Netherlands
Genetics Unit, Department of Biochemistry, University of Oxford, Oxford, UK
Department of Paediatrics, University of Melbourne, Melbourne, VIC Australia
Avihu Boneh, ua.gro.sgcv@henob.uhiva.
Collaborators: Johannes Zschocke and K Michael Gibson
Division of Human Genetics, Medical University Innsbruck, Innsbruck, Austria
WSU Division of Health Sciences Clinical Pharmacology Unit, Spokane, Washington USA

Abstract

Pyruvate dehydrogenase complex (PDHC) deficiency is a disorder of energy metabolism that leads to a range of clinical manifestations. We sought to characterise clinical manifestations and biochemical, neuroimaging and molecular findings in thiamine-responsive and nonresponsive PDHC-deficient patients and to identify potential pitfalls in the diagnosis of PDHC deficiency. We retrospectively reviewed all medical records of all PDHC-deficient patients (n = 19; all had PDHA1 gene mutations) and one patient with severe PDHC deficiency secondary to 3-hydroxyisobutyryl-CoA hydrolase deficiency managed at our centre between 1982 and 2012. Responsiveness to thiamine was based on clinical parameters. Seventeen patients received thiamine treatment: eight did not respond, four showed sustained response and the others responded temporarily/questionably. Sustained response was noted at thiamine doses >400 mg/day. Age at presentation was 0–6 and 12–27 months in the nonresponsive (n = 8) and responsive (n = 4) patients, respectively. Corpus callosum abnormalities were noted in 4/8 nonresponsive patients. Basal ganglia involvement (consistent with Leigh disease) was found in four patients (including 2/4 thiamine-responsive patients). Diagnosis through mutation analysis was more sensitive and specific than through enzymatic analysis. We conclude that patients presenting at age >12 months with relapsing ataxia and possibly Leigh syndrome are more likely to be thiamine responsive than those presenting with neonatal lactic acidosis and corpus callosum abnormalities. However, this distinction is equivocal and treatment with thiamine (>400 mg/day) should be commenced on all patients suspected of having PDHC deficiency. Mutation analysis is the preferable first-line diagnostic test to avoid missing thiamine-responsive patients and misdiagnosing patients with secondary PDHC deficiency.

Short Summary: Thiamine responsiveness is more likely in patients presenting at age >12 months with relapsing ataxia and possibly Leigh syndrome than in those presenting with neonatal lactic acidosis and corpus callosum abnormalities. Thiamine doses >400 mg/day are required for sustained response. Mutation analysis is more sensitive and specific than enzymatic analysis as a first-line diagnostic test.

Introduction

The pyruvate dehydrogenase complex (PDHC) is pivotal for energy metabolism in that it catalyses the oxidative decarboxylation of pyruvate into acetyl-CoA, linking glycolysis to the tricarboxylic acid cycle. The PDHC is comprised of several copies of enzymatic subunits, E1 (pyruvate dehydrogenase), E2 (dihydrolipoamide transacetylase) and E3 (dihydrolipoamide dehydrogenase) and an E3 binding protein (E3BP), and its activity is regulated by several isoforms of kinases and phosphatases through reversible phosphorylation (Linn et al. 1969; Pagliarini and Dixon 2006). The E1 enzyme consists of two α- and two β-subunits that share a binding site for thiamine pyrophosphate (TPP), a metabolite of thiamine, which is an essential cofactor for the enzymatic reaction and also helps to maintain the PDHC in an activated state by inhibiting its phosphorylation (Roche and Reed 1972).

PDHC deficiency causes impairment of energy metabolism and leads to a broad range of symptoms. Four main neurological presentations have been reported: neonatal encephalopathy with lactic acidosis, non-progressive infantile encephalopathy, Leigh syndrome and relapsing ataxia (Robinson et al. 1987; Brown et al. 1988, 1989a; Barnerias et al. 2010; Patel et al. 2012). The majority of patients have a mutation located in the PDHA1 gene encoding the E1α subunit, which is located on the X chromosome (Robinson and Sherwood 1984; McKay et al. 1986; Wicking et al. 1986; Brown et al. 1989b; Lissens et al. 2000). The differences in presentation result from variations in mutations and from the degree of X inactivation in females (Brown et al. 1989b; Dahl et al. 1992).

A high-fat diet, with the amount of calories provided from fat exceeding 50%, has been shown to be effective in many patients with PDHC deficiency. It leads to decreased blood lactate and pyruvate concentrations and provides an alternative source of energy in the form of ketones (Wexler et al. 1997) and has been shown to be helpful in reducing childhood onset epilepsy and paroxysmal dystonia (Barnerias et al. 2010). However, its efficacy has been variable (Weber et al. 2001). The administration of thiamine is an additional treatment that can potentially be effective, given its role as a cofactor (as TPP) of the enzyme-complex activity (Di Rocco et al. 2000; Lee et al. 2006; Barnerias et al. 2010; Giribaldi et al. 2012).

The diagnostic process usually includes measuring PDHC activity in cultured fibroblasts or skeletal muscle, but there may be great variation in fibroblast activities reported by different laboratories. Thus, it is likely that not all patients are correctly diagnosed and therefore the exact prevalence of PDHC deficiency is not known (Barnerias et al. 2010; Patel et al. 2012). Moreover, thiamine-responsive patients might be missed, because enzyme analysis is usually performed with high TPP concentrations and will not show a decreased PDHC enzyme activity in these patients (Di Rocco et al. 2000; Naito et al. 2002a; Lee et al. 2006). Amongst patients with thiamine-responsive PDHC deficiency, several different mutations have been found; the majority located in the PDHA1 gene, encoding the E1α subunit, some in the region encoding the TPP-binding site, some outside the TPP-binding site (Naito et al. 1994, 2002a, b; Benelli et al. 2002; Debray et al. 2008). At present, it is not known whether thiamine-responsive patients present differently from those not responsive to thiamine and what doses of the vitamin are required to gain significant therapeutic benefits.

The aim of this study was to review all clinical and laboratory data on our patients with PDHC deficiency in order to elucidate whether a prediction of thiamine responsiveness can be made in patients with PDHC deficiency based on clinical, biochemical or neuroimaging studies and to demonstrate the difficulties in diagnosing PDHC deficiency accurately and reliably by enzymatic analysis. To this end, we included in this study a patient with secondary PDHC deficiency, presumed to have primary PDHC deficiency at the time of presentation.

Methods

We conducted a retrospective review of all medical records of all patients with symptoms and biochemical findings consistent with PDHC deficiency at the Royal Children’s Hospital in Melbourne, Australia. Included patients were born in Victoria or Tasmania between 1 January 1982 and 2012. Available clinical, laboratory and neuroimaging characteristics were collected into a database. We defined patients with PDHC deficiency as those with a pathogenic mutation in one of the genes encoding the PDHC subunits or a combination of clinical presentation consistent with PDHC deficiency (e.g. neonatal lactic acidosis, Leigh disease, relapsing ataxia etc.) and diagnostically low PDHC enzyme activity with no other confirmed diagnosis. We also included in this study one patient with severe PDHC deficiency secondary to 3-hydroxyisobutyryl-CoA hydrolase (HIBCH) deficiency determined enzymatically. The study was approved by the Human Research Ethics Committee of the Royal Children’s Hospital (HREC # 32087A).

Enzyme analyses were performed usually in cultured skin fibroblasts as previously published (Rahman et al. 1996). Briefly, total (dichloroacetate-activated) PDHC activity was assayed by measuring the rate of 14C–CO2 formation from [1-14C]pyruvate. PDHC activity was also measured in lymphocytes and/or skeletal muscle biopsies from some patients. Activities were expressed as percentages of normal control mean relative to protein concentration and relative to citrate synthase. Mutations were identified by sequencing the coding region of the PDHA1 gene in cDNA prepared from patient fibroblasts and then confirmed in genomic DNA as described previously (Ridout et al. 2008).

Clinical characteristics included pregnancy and perinatal history, age at onset, presenting symptoms and signs, details on thiamine treatment and outcome. Responsiveness to thiamine treatment was defined as sustained clinical improvement (i.e. regaining spontaneous breathing after a period of intubation; decline in seizure activity; developmental gain with progress in communication, intellectual or motor skills; improvement in mobility, tone or coordination; or a decrease in movement disorders (Pastoris et al. 1996; Di Rocco et al. 2000; Naito et al. 2002a, b)) and/or improvement suggested by neuroimaging after the start of thiamine administration. Findings of brain computerised tomography (CT), ultrasonography (US) or magnetic resonance imaging (MRI) before and after the start of thiamine treatment were reviewed, as available. The lowest and highest concentrations of lactate, pyruvate and alanine in blood (arterial, venous and capillary) and cerebrospinal fluid (CSF) before and after start of thiamine treatment were noted but are not reported here. We did not consider biochemical improvement in the definition of responsiveness to thiamine because values of lactate and pyruvate range broadly, are inconsistent and can be influenced by a variety of factors, as has been well documented previously (Chariot et al. 1994). Moreover, lack of correlation between lactate and pyruvate concentrations and clinical outcome has been shown in previous studies on treatment of patients with PDHC deficiency (Barnerias et al. 2010).

Results

We identified 19 patients with primary PDHC deficiency and one patient with secondary PDHC deficiency due to 3-hydroxyisobutyryl-CoA hydrolase (HIBCH) deficiency. Details regarding the presenting symptoms and signs and confirmation of diagnosis are presented in Table 1. In the group of patients with PDHC deficiency feeding difficulties were a common presenting symptom in the neonatal period (n = 7/9) as well as lethargy (5/9), hypotonia (4/9) and abnormal respiration (3/9). Seizures were noted in two patients who presented in the neonatal period and two who presented later. Intermittent ataxia was noted in two patients (or possibly three, including patient 15). The patient with HIBCH deficiency also presented with feeding difficulties.

Table 1

Presenting symptoms/signs and results of confirmatory tests

Patient number/sexAge at onsetPresenting symptoms/signsPDHC activity (%)Mutation
1/maleNeonatalFeeding difficultiesF: 39PDHA1: c.1133G > A, p.Arg378His
Lethargy
Irregular breathing
Hypertonia
2/femaleNeonatalFeeding difficultiesF: 36 (PDH E1: 28)PDHA1: c.419-(17_14)delTGTT
LethargySM: 39
Hypotonia
3/femaleNeonatalFeeding difficultiesF: 21PDHA1: c.927_933delAGTAAGA, p.Ser312Valfs11
Lethargy
Hypotonia
Seizures
Facial asymmetry
4/maleNeonatalFeeding difficultiesF: 32PDHA1: c.483C > T, p.Tyr161Tyr
Lethargy
Respiratory distress
Hypotonia
Seizures
5/maleNeonatalFeeding difficultiesF: 35PDHA1: c.604-10C > G
Lethargy
Respiratory distress
Hypotonia
6/femaleNeonatalFeeding difficultiesF: 19PDHA1: c.511G > A, p.Val171Met
7/femaleNeonatalFeeding difficultiesF: 50PDHA1: c.947C > T, p.Pro316Leu
Hyperextension neck
8/femaleNeonatalMild lethargyF: 33 (relative to protein)PDHA1: c.1139_1142dupCCAA, p.Trp383Serfs5
9/femaleNeonatalCongenital retinal dystrophyF: 40 (PDH E1: 37)PDHA1: c.853C > T, p.Gln285*
10/maleM2SeizuresF: 43PDHA1: c.628A > G, p.Met210Val
Respiratory distressL: 53
11/maleM2Episodes of irregular breathingF: 11PDHA1: c.1132C > T, p.Arg378Cys
Seizures
12/maleM4Feeding difficultiesF: 43PDHA1: c.1156_7insT, p.Lys387*
HypotoniaL: 18
SM: <5
13/femaleM6Feeding difficultiesL: 39PDHA1 c.1156_7insT, p.Lys387*
Lethargy
Hypotonia
Developmental delay
14/femaleM12Developmental delayF: 64PDHA1: c.858_861dupTTAC, p.Arg288Leufs9
Progressive microcephaly
15/maleM12TremorF: 37PDHA1: c.787C > G, p.Arg263Gly
Frequent falling
General weakness
16/maleM14Intermittent ataxiaF: 75PDHA1: c.787C > G, p.Arg263Gly
17/femaleM24Developmental delayF: 10PDHA1: c.1156_7insT, p.Lys387*
Right hemiparesis
18/maleY2M3Intermittent ataxiaF: 41PDHA1: c.261T > G, p.Ile87Met
19a/female(Cascade testing)No symptomsNDPDHA1: c.261T > G, p.Ile87Met
20b/femaleNeonatalFeeding difficultiesF: 26

Y years, M months, Sz seizures, F fibroblasts, L lymphocytes, SM skeletal muscleaAsymptomatic female patient diagnosed through cascade testingbPDHC deficiency secondary to HIBCH deficiency

Other neurological and non-neurological symptoms and signs, noted through follow-up reviews, are summarised in Table 2. Ataxia was noted in 8/19 and dystonia in 4/19 patients, combined with chorea in two. Tremor was noted in 3/19 patients. Microcephaly was documented in 4/19 patients, all female. One female patient (patient 19, sister of patient 18) was asymptomatic. Only 4/19 patients had normal development.

Table 2

Follow-up: neurological and non-neurological manifestations

Patient/sexSeizures (age)Ataxia (age)Movement disorder (age)Tone/tendon reflexesOther neurological symptoms and signsOphthalmological findingsOther
1/maleM1Axial hypotonia, distal hypertonia/hyperreflexiaAspiration
2/femaleAxial hypertonia, distal hypertoniaEpisodes of cyanosis
3/femaleD6Axial hypotonia, fluctuating distal tone
4/maleD1Dystonia (D13)Axial hypotonia, distal hypertoniaAutonomic dysfunction
5/maleM19Generalised hypotonia/hyperreflexiaBulbar dysfunctionFluctuating ptosis
6/femaleM4M14Chorea/dystonia (Y9M10)Axial hypotonia, distal hypertoniaMicrocephalyAlternating exotropia (Lt)Skeletal deformities
DysphagiaPale optic disksFatigue
Peripheral oedema
7/femaleSpastic tetraparesis/hyperreflexiaMicrocephalySkeletal deformities
Cortical blindnessIrritability hepatomegaly
8/femaleY5M2Axial hypotonia, distal hypertonia/hyperreflexiaMicrocephaly
Mild pseudobulbar dysfunction
Sensorineural hearing loss
Ptosis (Lt)
Retinal changes
Skeletal deformities
9/femaleY3Axial hypotonia, distal hypertonia/hyperreflexiaMicrocephalyDivergent squintHyperlaxity of small joints
DysphagiaSleep disturbance
Lethargy
Contractures
10/maleM2Intermittent dystonia (Y10)Axial hypotoniaLethargyEpisode of unequal pupilsFeeding difficulties
Developed spastic tetraparesis/hyperreflexiaAxonal neuropathy
Aspiration
Episodes of respiratory distress
Skeletal deformities
Growth retardation
Anxiety/panic attacks
Easy bruising
11/maleM2Chorea (M6)Axial hypotonia, distal hypertoniaLethargyPale optic disks; nystagmusFeeding difficulties
Distal weaknessSkeletal deformities
Bulbar dysfunction
Episodes of irregular breathing
Fluctuating consciousness
12/maleGeneralised hypotoniaEpisodes lethargy and sweating
Quadriplegia
Bulbar dysfunction
Respiratory decompensation
13/femaleM22Y3M9Tremor (Y2M1)Axial hypotonia, distal hypertoniaEpisodes of irregular breathingAlternating divergent squintSkeletal deformities
ContracturesPale optic disksOsteopenia
Patella alta
Premature pubarche
Prolonged QT interval
Fatigue
Headaches
Mild memory loss
Episodic chest pain
Sleep disturbance
14/femaleY4M11Spastic tetraparesisGrowth retardation
Autistic features
15/maleM12Tremor (M12)Generalised hypotonia/areflexiaLethargy
Episodic respiratory decompensation
Bulbar dysfunction
Fasciculations
16/maleM20M14Tremor (M20)Generalised hypotonia/areflexiaDysphagiaIntermittent ophthalmoplegiaFeeding difficulties
Respiratory decompensationConvergent strabismusFatigue
DysarthriaHypertension
Sixth nerve palsyTachycardia
Prolonged QT interval orthopnoea
Thumb hypermobility
Episodes of hyperacusis
17/femaleY8M0Distal hypertoniaIntermittent hemiplegiaSkeletal deformities
Dysdiadochokinesia later persistent hemiplegiaOsteopenia
Sensorineural hearing lossFatigue
Headaches
Short concentration span
18/maleY2M3Tics/grunts/choreaEpisodic axial hypotoniaHyperreflexia Episodic lethargy/weaknessAstigmatismSkeletal deformities
(Y3M1)Distal hypertoniaPeripheral neuropathyGrowth retardation
DystoniaAcute hemiparesisHeadaches
(Y5M0)DysarthriaDaytime urine incontinence
19a/femaleNormal
20b/femaleM3Distal hypotoniaEpisodic respiratory distressOptic atrophy
Fluctuating distal toneAspiration

Y years, M months, D days, Sz seizures

aAsymptomatic female patient diagnosed through cascade testing

bPDHC deficiency secondary to HIBCH deficiency

Details on treatment and responsiveness to treatment are summarised in Table 3. High-fat diet (50–60% of total calorie intake) was prescribed routinely or as part of a “sick day regime” in 15 patients, alone or in combination with thiamine. Two patients (patients 2 and 5) did not receive thiamine treatment and one was lost to follow-up (patient 14). Eight patients (patients 3, 4, 6, 7, 9, 10, 12, 13) received thiamine at doses ranging from 50 mg to 300 mg/day with no clinical effect. All were on a high-fat diet (including one on expressed breast milk) and five were on anticonvulsants. The age at onset in these patients ranged from the neonatal period (five patients) to 6 months. Seizures occurred in 6/8, dystonia and bulbar dysfunction were noted in 3/8 and chorea in 2/8 patients. Ataxia was reported in only 2/8 patients. Five of these patients have deceased. Patients 1 and 11 (age at presentation: neonatal and 2 months, respectively) seemed to respond favourably but temporarily and deceased later (one at age 1 year; ~25 years ago). Both had seizures and one had chorea.

Table 3

Treatment and response

Patient number/sexStart/dose/dayResponseHigh-fat diet startAnticonvulsantsOutcome (age)
1/maleD14/400 mgRegained spontaneous breathingD16+Deceased (Y1)
2/femaleNoneN/ANoneNoneDeceased (M1D3)
3/femaleD5/50 mgNo clinical effectD7+Deceased (D27)
4/maleD21/150 mgNo clinical effectExpressed breast milk+Deceased (M1D13)
5/maleNoneN/ANone+Deceased (Y1M9)
6/femaleY9M10/150–300 mgNo clinical effectY10M8+Alive (Y13M6)
7/femaleM9D19/NINo clinical effectY1 (for 1 year)NoneDeceased (Y27M8)
8/femaleM7D21/200–600 mgImproved developmentM9D17NoneAlive (Y7M6)
9/femaleY6M7/300 mgNo clinical effectY2M10D12 (stopped by family)+Alive (Y17M11)
10/maleM3D5/300 mgNo clinical effectM3+Deceased (Y11M3)
11/maleM6D20/150–600 mg (1,200 mg attempted before death)Regained spontaneous breathingM14D20+Deceased (Y2M9)
Improved mobility and development
Decline in seizures
12/maleM6D30/150 mgNo clinical effectM6NoneDeceased (Y1M1)
13/femaleY14M8/100 mgNo clinical effectM6NoneAlive (Y18M4)
14/femaleY4M11/300 mgNo informationY4M11Lost to follow-up (Y6M6)
15/maleY1M3/200–600 mgRegained spontaneous breathingNoneNoneAlive (Y3M10)
Improved mobility and strength
Resumed gait and speech within weeks. Attends regular kinder
16/maleY1M8/300–1,200 mgImproved coordination and toneY1M8 (for 1 year; then when unwell)NoneAlive (Y8M11)
Developmental gains: intellectual, communication and mobility
Improved neuropsychological performance at age Y7M3. Attends mainstream school
17/femaleY13M6/100–500 mgImproved mobility and strengthY16M3 (40% of calories)NoneAlive (Y31M4)
Increased energy level
18/maleY2M5/300–600 mgImproved development: intellectual, communication and mobility (limited by contractures)Y2M5NoneAlive (Y9M8)
Increased energy level
19a/femaleY3M5/300–600 mgRemains asymptomaticNoneNoneAlive (Y9M8)
20b/femaleY5M2/200–300 mgNo clinical effectNone+Deceased (Y8M1)

Y years, M months, D days, Sz seizuresaAsymptomatic female patient diagnosed through cascade testingbPDHC deficiency secondary to HIBCH deficiency

Four patients (patients 15, 16, 17, 18) were treated with thiamine doses 200–1,200 mg/day and showed sustained response at doses above 500 mg/day. Of these, one was not on a high-fat diet, another was only temporarily on the diet and one was on a “moderately” (40%) high-fat diet. The age at presentation of these patients ranged from 12 to 27 months. Ataxia was reported in 3/4, tremor in 2/4 and bulbar dysfunction and chorea/dystonia in one patient, each. Seizures occurred in only one of these patients. Two of the three patients who regained spontaneous breathing after a period of acute deterioration had only transient response; hence, this feature cannot be ascribed unequivocally to thiamine responsiveness and may reflect the natural history of the disorder. Patient 8, who presented in the neonatal period, is severely disabled, and although she seems to have improved on thiamine treatment, it is hard to confirm improvement objectively. The patient with HIBCH deficiency did not respond to thiamine.

Neuroimaging was done in all 20 patients except for one nonresponsive patient and one asymptomatic PDHC-deficient patient (Table 4). Structural abnormality of the corpus callosum was noted in 6/19 patients (4/9 patients who presented in the neonatal period; two who presented in the second month of life, including one prenatal ultrasonographic diagnosis). Of these patients, 4/6 did not respond to thiamine treatment, 1/6 responded temporarily and 1/6 did not receive thiamine treatment. Cystic lesions were noted in 6/9 patients who presented in the neonatal period. Pre-thiamine treatment imaging revealed basal ganglia involvement (consistent with Leigh disease) in 2/4 thiamine-responsive patients, one who was not treated with thiamine and one patient who had transient response to treatment. The patient with HIBCH deficiency had “abnormalities in brainstem and basal ganglia”.

Table 4

Neuroimaging

Patient numberNeuroimaging
Before thiamine RxAfter thiamine Rx
 1US: multiple cystic lesions in anterior aspect choroid plexus. Mild asymmetry and dilatation of ventricular systemCT (8 D after start Rx): no abnormalities
US (14 D later): cystic lesions floor lateral ventricles, possible resolving sub-ependymal haemorrhage
 2US: CC agenesis, right choroid cyst. Gross dilatation lateral ventricles. Diminished amount of cerebral substance. Prominent cisterna magna
 3US: possible cystic structures floor lateral ventricles. Gross hydrocephalus lateral and third ventricles
 4MRI: thin CC, periventricular cysts, mild lateral and third ventriculomegaly. Immature cerebral sulcation. Under opercularisation. Globally abnormal signal white matter, generous extra-axial surface spaces. MRS: No elevated lactate
 5MRI: symmetric signal abnormalities cerebellar hemispheres, brain stem, R occipital pole, L hypothalamus (consistent with Leigh syndrome)
MRI (15D later): interval evolution of signal abnormalities. Resolution of acute oedema, generalised cerebral atrophy. Possible signal changes ganglio-thalamic regions
MRS: elevated choline, lactate and pyruvate
 6MRI: microcephaly, partial CC agenesis, posterior fossa arachnoid cyst. Asymmetrical ventriculomegaly. Cerebral atrophy Lt > Rt
MRI (9Y10M later): myelination of remaining white matter occurred, no other changes
MRS: abnormal lactate peaks over left periventricular white matter
 7
 8MRI: asymmetric lateral ventricles, white matter loss Lt > Rt, cerebral atrophy. Extra-axial collection R frontal region
 9MRI: partial CC agenesis. Probable large arachnoid cyst posterior fossa. Possible stenosis in aqueduct of Sylvius causing hydrocephalus
10Prenatal US: agenesis of the CCMRI (1 M after start Rx): CC agenesis, dysmorphic ventricles. MRS: increase lactate
MRI (5 M after stop Rx): CC agenesis, colpocephaly, signal change in globus pallidus and deep cerebellar white matter. MRS: lactate peak brainstem and basal ganglia
11CTc: mild prominence lateral and third ventriclesCT(7 M after start Rx): stable prominent ventricles and extra-axial spaces
MRI: thin CC. Symmetrical signal abnormalities hippocampi, globus pallidi, lateral thalami (consistent with Leigh Syndrome). Enlarged extra-axial CSF spaces
12
13MRI: brachycephaly. Possible white matter loss, but myelination not yet completed
14CT: ventricular dilatation and cerebral atrophy
MRI: cerebral atrophy
15MRI: no abnormalities
16MRI: symmetrical signal changes in globus pallidi, putamen, dentate nuclei, ventral medulla and around fourth ventricle (consistent with Leigh syndrome)MRId: decrease of signal changes except for increased signal in dentate nucleus. New areas in cerebral peduncles
MRS: raised lactateMRI (1 Y9 M after restart Rx): decrease in signal changes, areas partially replaced by more cystic changes
MRI (5 M later): progression of swelling, patchy signal changes globus pallidi. Slight increase signal change medulla
MRI (4 Y7 M later): Slight increase hyperintensity dorsal medulla. Decrease in signal changes and resolution of swelling, with residual cyst
17MRI: decreased signal in L globus pallidus and small area of increased signal in R globus pallidus and signal change L cerebral peduncle
18MRI: no abnormalitiesMRI (2 Y6 M after start Rx): extensive gliosis, bilateral cystic and asymmetrical signal changes basal ganglia
MRI (6 M later): stable appearance
19a
20b MRI: thin CC, prominence of ventricular system, cerebral atrophy. Delayed myelination, abnormalities brain stem and basal ganglia

Y years, M months, D days, CC corpus callosum, Lt left, Rt right

aAsymptomatic female patient diagnosed through cascade testing

bPDHC deficiency secondary to HIBCH deficiency

c4 days after starting thiamine treatment

d5 days after restarting thiamine treatment, after ceasing treatment for 6 months

Enzyme analysis was done in all but patient 19, who was diagnosed by mutation analysis through cascade screening. Fibroblast PDHC activity, expressed relative to citrate synthase, ranged from 19% to 50% and from 10% to 75% of normal in thiamine-nonresponsive and thiamine-responsive patients, respectively (with no substantial difference between samples from male or female patients in these groups). In two thiamine-responsive patients (patients 14 and 16), enzyme activity was not low enough to be diagnostic for PDHC deficiency, and mutation analysis eventually established the diagnosis. Of particular note is the very low PDHC activity (26% of normal) in cultured fibroblasts of the patient later found to have HIBCH deficiency.

Pathogenic mutations leading to PDHC deficiency were identified in all patients with PDHC deficiency (Table 1). All our patients had mutations in the PDHA1 gene (Table 1) with at least 7/19 having de novo mutations (three mothers were carriers; information regarding the other mothers is not available). Patients 1, 3, 4, 7, 10, 11, 15 and 16 had mutations that had been described previously. All other patients were found to have novel mutations, splicing, insertions and duplications (some in the same region as other similar mutations; e.g. p.Ile87Met in thiamine-responsive patients 18, 19). The patient with HIBCH deficiency was found to be a compound heterozygote for mutations in the HIBCH gene.

Discussion

We sought to identify clinical, laboratory and imaging findings that may predict clinical responsiveness to thiamine treatment in patients with PDHC deficiency. Despite the limitations of it being a retrospective observational study, with a limited number of patients, our results highlight the pitfalls in the clinical and biochemical diagnosis of PDHC deficiency and the need for high thiamine doses to achieve sustained clinical benefit from this treatment. Prospective studies with larger cohorts of patients are needed to further elucidate the clinical features that may clearly distinguish and enable prediction of responsiveness and the thiamine doses for sustained response.

The spectrum of clinical manifestations in our cohort of patients is in line with previous reports (Barnerias et al. 2010; Giribaldi et al. 2012; Patel et al. 2012). The high prevalence of feeding difficulties and lethargy soon after birth and the high prevalence of skeletal deformities are intriguing, as they have not been previously reported (Barnerias et al. 2010; Patel et al. 2012), possibly because they had not been directly associated with this diagnosis. It is also likely that some of the skeletal deformities were secondary to dystonia. Our observations indicate that patients who present with relapsing ataxia and those with Leigh syndrome are more likely to be responsive to thiamine therapy. Ataxia has been previously described in thiamine-responsive patients (Kinoshita et al. 1997; Di Rocco et al. 2000) and in patients with mutations within the thiamine pyrophosphate domain (Debray et al. 2008). From a neuroimaging perspective, basal ganglia abnormalities were more common in thiamine-responsive patients, in support of the diagnosis of Leigh disease, whereas corpus callosum abnormalities and cystic lesions were found mainly in nonresponsive patients, in support of an early, possibly intrauterine insult to the CNS. However, these characteristics are not absolutely distinctive as they can be present in both responsive and nonresponsive patients. No differences were found in other clinical or biochemical characteristics (plasma and CSF lactate and pyruvate concentrations, PDHC enzyme activity).

Our observations highlight difficulties and potential pitfalls in diagnosing PDHC deficiency. For example, enzyme analysis of patient 16 was not low enough to be diagnostic for PDHC deficiency and a diagnosis could only be made through mutation analysis. Patient 18 was found to have a novel missense mutation adjacent to two previously reported missense mutations associated with thiamine responsiveness. This patient would not have been diagnosed by standard enzymatic assay as the activity in his cultured fibroblasts was borderline low yet within the normal range both with and without TPP in the assay mixture. Similarly, patient 12 had substantial residual PDHC activity in cultured cells but undetectable activity in skeletal muscle, possibly due to different levels of TPP in tissue versus culture media or to other factors affecting PDHC stability in different cell types. Such pitfalls have been previously described (Di Rocco et al. 2000; Lee et al. 2006). Interpretation of results is further complicated in female patients, where enzyme activity reflects the impact of X inactivation (e.g. patient 14). On the other hand, PDHC activity was very low in fibroblasts from the patient with HIBCH deficiency, who presented with a clinical phenotype, blood and CSF biochemistry and neuroimaging findings suggestive of PDHC deficiency, indicating that secondary PDHC deficiency could mistakenly lead to a diagnosis of PDHC deficiency unless molecular analysis is done. It may therefore be concluded that in order to avoid missing thiamine-responsive PDHC deficiency and misdiagnosing secondary PDHC deficiency, mutation analysis would be the preferred diagnostic test. Given that the vast majority of patients with PDHC deficiency have defects in the X-linked PDHA1 gene, it would seem prudent to sequence that gene first.

It is difficult to establish the exact relationship between mutations in the PDHA1 gene and “thiamine responsiveness”. Before the structure of the E1 enzyme was elucidated, a number of mutations were proposed to involve amino acid residues located within the TPP-binding region. These assignments are not consistent with the known structure of the enzyme, so interpretations of their biochemical consequences are difficult to evaluate. None of the mutations that have been well defined as causing “thiamine-responsive” PDHC deficiency have involved amino acid residues that are directly involved in TPP binding; most are adjacent to the binding site and probably alter the position of the actual side chains that interact with the cofactor, weakening their binding. There are two groups of these: p. Ile87Met (present report), p.Arg88Ser (Marsac et al. 1997), p.Arg88Cys (Fujii et al. 2006) and p.Gly89Ser (Naito et al. 1999) on one side and p.Phe205Leu (Naito et al. 2002a), p.Met210Val (Tripatara et al. 1999), p.Trp214 (Lissens et al. 2000), p.Leu216Phe (Naito et al. 2002a) and p. Pro217Leu (also denoted as Pro188Leu) (Hemalatha et al. 1995) on the other. A number of patients have been reported as thiamine responsive but have mutations involving amino acid residues that are located well away from the TPP-binding site. These include p.Val71Ala and p.Cys101Phe (Naito et al. 2002b), p.Tyr161Cys (Lee et al. 2006), p.Tyr243Ser (Benelli et al. 2002), p.Arg263Gly (Bachmann-Gagescu et al. 2009) and other C-terminal residues (Narisawa et al. 1992). At present, there is no structural explanation as to why these should influence TPP binding.

As the TPP-binding site is in the interface between the E1α and β subunits, it might be expected that defects in either one could be thiamine responsive. However, given that the total number of E1 β patients is very small, it is not surprising that no such patients have been described so far. Patients with defects in thiamine transport or the activation of thiamine to TPP would also be expected to be thiamine responsive.

The proportion of thiamine-responsive patients in our cohort (4–5/19) is high in comparison with previous reports (Barnerias et al. 2010; Patel et al. 2012). This difference could be due to differing definitions of responsiveness, namely clinical, biochemical or other. In this regard, we found that clinical response does not correlate with biochemical response (e.g. lactate, pyruvate; data not shown). Another possible reason for our relatively high number of thiamine-responsive patients is an increasing awareness to the possibility of thiamine-responsive PDHC deficiency, coupled with molecular testing. Indeed, there has been an increase in our diagnosis of thiamine-responsive patients within the last 5 years. A third possibility is the use of high (>400 mg/day) or very high (>1,000 mg/day) thiamine doses in our patients. Previous reports have highlighted the discrepancy between in vitro and in vivo response to thiamine. In some reports, recovery of activity has been demonstrated in vitro when the enzyme was assayed in high concentrations of TPP, but there has been no clear clinical response to vitamin supplementation. Debray et al. reported two siblings who were found to have a mutation in the thiamine-binding domain but did not show any clinical benefit from thiamine treatment, at 150 and 250 mg/daily, respectively (Debray et al. 2008). Likewise, Barnerias et al. reported only 1 of 22 patients who was thiamine responsive, but the daily thiamine dose was low (50 mg) (Barnerias et al. 2010). Patel et al. reviewed data on 371 patients and reported that only 73 received thiamine (from a few milligrams to >1,000 mg daily), but no information was given regarding the correlation between doses and responsiveness (Patel et al. 2012). Thus, there are insufficient reported data on the optimal daily thiamine dose that patients with PDHC deficiency should receive. In our cohort, the lowest effective daily dose for sustained benefit was 400 mg/day, but we noted that higher doses should be used in order to maximise the clinical response.

We conclude that patients with PDHC deficiency who present with relapsing ataxia or Leigh disease are more likely to respond to thiamine treatment than those who present with neonatal lactic acidosis and have structural abnormalities of the corpus callosum on imaging. However, responsiveness to thiamine cannot be reliably predicted based on these clinical and neuroimaging differences since some overlap between the two groups exists. We suggest that high-dose (>400 mg/day) thiamine treatment be initiated as soon as PDHC deficiency is suspected. Early treatment will quickly reverse the manifestations of thiamine deficiency and might prevent irreversible complications (e.g. contractures). Plasma thiamine concentration and mutation analysis of the PDHA1 gene should be done as first-line investigations (along with the usual metabolites analysis), in order to avoid missing thiamine-responsive patients and misdiagnosing patients with secondary PDHC deficiency. If negative, enzymatic analysis, molecular analysis of other PDHC genes and a search for other diagnoses should be done. New molecular tests for defects in thiamine transport will enable the correct diagnosis in more patients who present with manifestations suggestive of PDHC deficiency.

Acknowledgements

This work was presented at the SSIEM annual symposium, Birmingham, 2012. J. Inherit. Metab. Dis. 35 (Supp. 1): 122, 2012. We thank Denise Kirby, Wendy Hutchison and Henrik Dahl (Melbourne) and Cheryl Ridout (Oxford) for their contributions to enzyme and molecular diagnosis of the patient cohort. This work was supported by the Victorian Government’s Operational Infrastructure Support Program.

Glossary

Abbreviations

HIBCH3-Hydroxyisobutyryl-CoA hydrolase
MRIMagnetic resonance imaging
PDHCPyruvate dehydrogenase complex
TPPThiamine pyrophosphate

Compliance with Ethics Guidelines

Conflict of Interest

All the authors of this chapter declare that there are no conflicts of interest.

Details of the Contributions of Individual Authors

Sanne Van Dongen: Reviewed the literature, reviewed all patients’ records, collected data into Excel spreadsheets, wrote first manuscript and corrected and rewrote subsequent versions. Reviewed the final version and approved it.

Ruth Brown: Performed mutation analysis, reviewed mutations results and participated in writing the methods, results and discussion sections. Critically reviewed the manuscript and approved its final version.

Garry Brown: Reviewed mutations results and participated in writing the methods, results and discussion sections. Critically reviewed the manuscript and approved its final version.

David Thorburn: Reviewed enzymology and mutation results and participated in writing the methods, results and discussion sections. Critically reviewed the manuscript and approved its final version.

Avihu Boneh: Initiated the study, supervised and reviewed all clinical and laboratory data collection and tabulation, participated in writing all versions of the manuscript and wrote the final version of the manuscript.

Footnotes

Competing interests: None declared

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