Chapter 165. Disorders of Bile Acid Synthesis

Metabolic Pathways

The major bile acids produced in the liver are the taurine and glycine conjugates (amidates) of chenodeoxycholic acid (CDCA) and cholic acid (CA). Secretion of these conjugated bile acids into the canaliculi by the bile salt export pump fuels bile flow out of the liver. They are powerful detergents. This property is important in keeping cholesterol in solution in bile, and it is also essential for the digestion and absorption of fats and fat-soluble vitamins. The conversion of cholesterol to bile acids and the secretion of cholesterol into bile represent the major routes for elimination of excess cholesterol from the body. Thus, individuals with disorders of bile acid synthesis can present with liver disease due to impaired bile secretion (cholestasis), with gallstones, steatorrhea, symptoms of fat-soluble vitamin malabsorption, and buildup of cholesterol in tissues (causing neurological impairment, atherosclerosis, and xanthomata).

The conversion of cholesterol to bile acids requires modifications to the sterol nucleus and to the sterol side chain.1 The major pathway for bile acid synthesis in adults (the “neutral” pathway) starts with conversion of cholesterol to 7α-hydroxycholesterol. A second pathway (the “acidic” pathway) starts with the conversion of cholesterol to 27-hydroxycholesterol, and the early steps can take place outside the liver. The neutral and acidic pathways share several enzymes; thus, inborn errors can disrupt both routes for the synthesis of bile acids. A simplified version of the major bile acid synthesis pathways is shown in Figure 165-1.

The modifications to the cholesterol side chain involve 27-hydroxylation, further oxidation to a C27 bile acid, alteration of the stereochemistry of the C27 bile acid CoA ester (racemization), and then β-oxidation in the peroxisomes to produce a C24 bile acid CoA ester that is conjugated with glycine or taurine.

Disorders of bile acid synthesis that affect transformation of the cholesterol nucleus produce hepatobiliary disease; those that affect the oxidation of the cholesterol side chain often also cause extrahepatic (particularly neurological) disease. This is probably because the early steps of the acidic pathway for bile acid synthesis play an important role in controlling cholesterol concentrations in extrahepatic tissues.

Unusual bile acids and bile alcohols in urine are best detected by fast atom bombardment mass spectrometry (FAB-MS)2 or electrospray ionization tandem mass spectrometry (ESI-MS/MS).3 Definitive identification of metabolites is most easily achieved using gas chromatography-mass spectrometry (GC-MS).4eFigure 165.1 shows some diagnostic urinary bile acid and bile alcohol profiles.

Clinical Presentation

This disorder has not yet been detected in children. Homozygous cholesterol 7α-hydroxylase deficiency was detected in three adults with hypercholesterolemia, hypertriglyceridemia, and premature gallstone disease.5 One suffered from premature coronary and peripheral vascular disease. Their LDL cholesterol levels were noticeably resistant to treatment with inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase (statins). A study of the kindred revealed that individuals heterozygous for the mutation were also hyperlipidemic, indicating that this is a codominant disorder.

Metabolic Derangement and Pathophysiology

A block at the level of cholesterol 7α-hydroxylase prevents the neutral pathway from synthesizing bile acids. However, this is probably compensated for by an upregulation of the acidic pathway, which increases the synthesis of CDCA. In one homozygote, the cholesterol content of a liver biopsy was increased. Fecal bile acid output was reduced, and the ratio of chenodeoxycholic acid derived fecal bile acids/cholic acid derived fecal bile acids was increased.

Genetics

Cholesterol 7α-hydroxylase deficiency is a codominant disorder caused by mutations in the CYP7A1 gene on chromosome 8q11-q12. The only mutation described to date is a frameshift mutation (L413fsX414) that results in loss of the active site and enzyme function.

Diagnostic Tests

There is no simple blood or urine test for cholesterol 7α-hydroxylase deficiency. Analysis of CYP7A1 can be undertaken in a patient with statin-resistant hypercholesterolemia.

Treatment and Long-Term Outcome

Control of plasma cholesterol and triglyceride levels requires treatment with a combination of powerful HMG-CoA reductase inhibitor (atorvastatin) and niacin. The variability of the disorder and long-term prognosis is not known.

Clinical Presentation

Deficiency of 3β-HSD typically presents in the neonatal period with conjugated hyperbilirubinemia, elevated transaminases, and a normal γ-gutamyl transpeptidase.6,7 There is often associated steatorrhea. The liver biopsy shows a giant-cell hepatitis with evidence of cholestasis (retained bile pigment). Fat-soluble vitamin malabsorption may become evident from rickets (or even severe symptomatic hypocalcemia), vitamin K–responsive coagulopathy, or low plasma concentrations of vitamin E or vitamin A.6,8,9 If left untreated, the liver disease progresses with hepatomegaly, splenomegaly, and evidence of increasing fibrosis on a liver biopsy; the picture can resemble progressive familial intrahepatic cholestasis (PFIC), although pruritus is perhaps less common in 3β-HSD deficiency than in PFIC.10 Occasionally the disorder is first detected in a patient with chronic hepatitis in the second decade or beyond.

Metabolic Derangement and Pathophysiology

3β-HSD is thought to be involved in both the neutral and the acidic pathway of bile acid synthesis, so a severe reduction in enzyme activity leads to very marked impairment of CDCA and CA synthesis. The accumulating 3β-hydroxy-Δ5intermediates (such as 7α-hydroxycholesterol) can undergo the other reactions required for conversion to C24 bile acids (eFig. 165.2). Thus, these patients synthesize considerable amounts of bile acids that resemble CDCA and CA, except that they retain the 3β-hydroxy-Δ5 configuration of cholesterol. These bile acids are readily sulfated, so the major urinary bile acids are sulfated 3β,7α-dihydroxy-5-cholenoic acid, sulfated 3β,7α,12α–trihydroxycholenoic acid, and their glycine conjugates. The bile acids that are produced in large amounts are not substrates for the bile salt export pump and probably actually inhibit it. Thus, transport of bile acids into bile is very much reduced and with it bile flow (cholestasis). Bile acid deficiency in the gut contributes to malabsorption of fat and fat-soluble vitamins.

Genetics

3β-HSD deficiency is an autosomal recessive trait caused by reduced activity of 3β-hydroxy-Δ5-C27-steroid dehydrogenase (OMIM 607764) which is encoded by the HSD3B7 (C27-3BETA-HSD) gene on chromosome 16p11.2-12. Twelve mutations have been described to date.11 Prenatal diagnosis by DNA analysis is possible, but this is a very treatable disease.

Diagnostic Tests

Urine bile acid analysis shows peaks attributable to the sulfates and glycine-conjugated sulfates of 3β,7α-dihydroxy-5-cholenoic acid and 3β,7α,12α-trihydroxy-5-cholenoic acid: (m/z 469,485,526,542). Plasma bile acid analysis shows the presence of these two bile acids and C27 analogues, particularly 3β,7α-dihydroxy-5-cholestenoic acid. The plasma concentrations of CDCA and CA are strikingly low for a child with cholestasis. The activity of 3β-HSD is reduced in cultured skin fibroblasts,12 and mutations can be found in the HSD3B7 gene.

Treatment and Long-Term Outcome

Emergency Treatments

Severe coagulopathy requires intravenous vitamin K. Seizures and tetany due to hypocalcemia require intravenous calcium and correction of vitamin D deficiency. Both of these problems will not recur once bile acid replacement has been started.

Bile Acid Replacement

The first patients were treated with chenodeoxycholic acid13,8at a dose of 12 to 18mg/kg per day for 2 months, followed by 9 to 12 mg/kg per day for maintenance. This led to improvement in symptoms, rapid normalization of liver function tests, and improvement in the appearances in the liver biopsy. Follow-up has shown that treatment with CDCA fully controls the disease into adult life (whereas untreated children with 3β-HSD deficiency may die from the complications of cirrhosis before age 5). Some children with severe liver disease (bilirubin > 120 μM and AST > 260 i.u/L) have shown an initial rise in transaminases with CDCA treatment14; in such patients, changing to a combination of cholic acid (7 mg/kg per day) plus CDCA (7 mg/kg per day) has produced a gradual normalization of liver function tests. Some hepatologists advocate using cholic acid alone.7

Clinical Presentation

Δ4-3-oxosteroid 5β-reductase deficiency leads to increased urinary excretion of bile acids with a 3-oxo-Δ4 structure, principally the glycine and taurine conjugates of 7α-hydroxy-3-oxo-4-cholenoic acid and 7α,12α-dihydroxy-3-oxo-4-cholenoic acid. However, patients who excrete these 3-oxo-Δ4 bile acids as the major urinary bile acids can be divided into three groups: those with proven mutations in SRD5B1 (AKR1D1, the gene encoding the 5β-reductase enzyme),15,16 those in whom this has been excluded,17 and those in whom the results of SRD5B1 mutation analysis have not been published.18,19 In the second and third groups, the cause of excretion of 3-oxo-Δ4 bile acids remains uncertain, and because this pattern of urinary bile acid excretion can be a nonspecific consequence of severe liver disease,20,21 the description in this chapter will focus on the five patients with proven SRD5B1 mutations.

All five patients presented as neonates with cholestatic jaundice with raised transaminases but normal γ-GT, low vitamin E, and prolonged clotting times that improved with parenteral vitamin K.15,16 Their liver biopsies showed giant cell transformation, canalicular and hepatocellular cholestasis, portal inflammation, septal fibrosis, occasional necrotic foci, and in some cases, increased extramedullary hemopoiesis. Cholestasis persisted in all cases. One infant progressed to liver failure, failed to respond to treatment with ursodeoxycholic acid or chenodeoxycholic acid, and had a liver transplant at 19 weeks.15 One child failed to respond to ursodeoxycholic acid treatment but responded extremely well to treatment with chenodeoxycholic acid and cholic acid (started at 8 months) and was asymptomatic at the age of 10 years.15,22,23 One patient showed an initial response to chenodeoxycholic acid plus cholic acid but then deteriorated (possibly due to cytomegalovirus infection) and required transplantation.15 Twin patients responded to treatment with cholic acid started at 8 months, and they were well at age 5 years.16

Metabolic Derangement and Pathophysiology

Δ4-3-oxosteroid 5β-reductase is thought to be involved in the neutral and the acidic pathways of bile acid synthesis and in one route for the degradation of steroid hormones with a 3-oxo-Δ4 structure (including glucocorticoids, mineralocorticoids, testosterone, and progesterone). The impaired function of the bile acid synthetic pathways results in very markedly reduced synthesis of CDCA and CA; these bile acids are present at unusually low concentration in the plasma and urine (for a child with cholestasis). The bile acid precursors with a 3-oxo-Δ4structure undergo side chain oxidation to produce 7α-hydroxy-3-oxo-4-cholenoic acid and 7α,12α-dihydroxy-3-oxo-4-cholenoic acid (eFig. 165.3). The glycine and taurine conjugates of these bile acids are prominent in plasma and are the major bile acids in urine. Bile acids with a 3-oxo-Δ4 structure are not substrates for the bile salt export pump and may actually inhibit it; hence the reduced bile flow (cholestasis).

Accumulating 3-oxo-Δ4intermediates (such as 7α-hydroxy-cholest-4-en-3-one) can be converted by a 5α-reductase enzyme to 5α(H) derivatives (eg, 7α-hydroxy-5α-cholestan-3-one) and hence to 5α(H)- or allo-bile acids (eg, allochenodeoxycholic acid, 3α,7α-dihydroxy-5α-cholanic acid). In one patient, while 3-oxo-Δ4 bile acids were the major bile acids in infancy, by age 11, the major plasma bile acids were allo bile acids. At the age of 11, this child had no evidence of liver disease or malabsorption despite having stopped treatment with CDCA and CA for a year. This suggests that, as childhood progresses, the formation of allo bile acids may actually compensate for the 5β-reductase deficiency. The same patient (with proven mutations in the AKR1D1 gene) was shown to have only trace amounts of 5β-reduced steroid metabolites in the urine. Despite the impairment of degradation of 3-oxo-Δ4steroid hormones, there was no evidence of Cushingoid features, hypertension, or a sex hormone problem, again suggesting that the 5α-reductase pathway can compensate.23

Genetics

Some cases showing a urine bile acid profile suggestive of 5β-reductase deficiency are due to homozygous mutations in the SRD5B1 (AKR1D1) gene (OMIM 604741). In others, abnormalities in the cDNA of this gene have been excluded, and in others, no information has been published (OMIM 235555).

Diagnostic Tests

Urine bile acid analysis shows that the main bile acids in the urine are 3-oxo-Δ4 bile acids and that excretion of the corresponding saturated bile acids is markedly reduced: 444>>448, 460>>464, 494>>498, 510>>514. Plasma bile acid analyses in infancy show markedly elevated concentrations of 7α-hydroxy-4-cholenoic acid and 7α,12α-dihydroxy-4-cholenoic acid. In an older child, the major plasma bile acids are allo bile acids.23

Treatment and Long-Term Outcome

Treatment of a bleeding diathesis with parenteral vitamin K may be required. Vitamin D may be needed for rickets. Deficiency of 5β-reductase can progress rapidly to liver failure. However, treatment with bile acid replacement therapy can normalize liver function and lead to good long-term health. Successful regimes include chenodeoxycholic acid plus cholic acid (8 mg/kg per day of each) and cholic acid alone. It is currently unknown whether there is spontaneous improvement with age,23 which would make continued bile acid replacement unnecessary. Any such tendency for spontaneous resolution will make assessments of long-term bile acid replacement regimes difficult.

Clinical Presentation

CTX has a range of clinical presentations, including cholestatic jaundice in early infancy,24-26 diarrhea and cataracts in the preschool child,27 learning difficulties in childhood, dementia in the teens/early adult life, and the development of tendon xanthomata and early atherosclerosis. The early liver disease can be fatal, and the early dementia can progress to crippling neurological dysfunction. These devastating outcomes are preventable with bile acid replacement therapy. For these reasons, early diagnosis, based on a high index of suspicion and appropriate tests, is extremely important.

It is important to recognize that the neurological deterioration is very variable.28 For example, some patients are normal intellectually but suffer from a neuropathy or mild spastic paresis; others have no neurological signs but present with psychiatric symptoms resembling schizophrenia. Motor dysfunction (spastic paresis, ataxia, expressive dysphasia) develops in approximately 60% of patients in the second or third decade of life.

Tendon xanthomata may be detectable during the second decade of life but usually appear in the third or fourth decade. The Achilles tendon is the most common site; other sites include the tibial tuberosities and the extensor tendons of the fingers and the triceps. Premature atherosclerosis leading to death from myocardial infarction occurs in some patients. In others, death is caused by progression of the neurological disease with increasing spasticity, tremor and ataxia, and pseudobulbar palsy.

Magnetic resonance imaging (MRI) of the brain may show diffuse cerebral atrophy and increased signal intensity in the cerebellar white matter on T2-weighted scans.29 Osteoporosis is common in CTX and may produce pathological fractures; it is associated with low plasma concentrations of 25-hydroxy-vitamin D and 24,25-dihydroxy-vitamin D.30

Metabolic Derangement and Pathophysiology

Sterol 27-hydroxylase is present in the liver and in extrahepatic tissues. It is the first step in the acidic pathway for bile acid synthesis (eFig. 165.4) and provides a route for removing cholesterol from extrahepatic tissues.31 In CTX, accumulating cholesterol is converted partly to cholestanol. Sterol 27-hydroxylase also participates in the neutral pathway of bile acid synthesis.32 5β-Cholestane-3α,7α,12α-triol cannot be hydroxylated in the C27 position and accumulates in the liver. Consequently, it is metabolized by an alternative pathway, starting with hydroxylation in the C25 position (in the endoplasmic reticulum). Further hydroxylations (eg, in the C22 or C23 position) result in the synthesis of the characteristic bile alcohols that are found (as glucuronides) in the urine (eFig. 165.4). Bile-acid precursors other than 5β-cholestane-3α,7α,12α-triol also accumulate. Some of these (eg, 7α-hydroxy-cholest-4-en-3-one) are probably converted to cholestanol by a pathway involving 7α-dehydroxylation. Because patients with CTX have a reduced rate of bile-acid synthesis, the normal feedback inhibition of cholesterol 7α-hydroxylase by bile acids is disrupted. This further enhances the production of bile alcohols and cholestanol from bile-acid precursors. Disruption of bile acid synthesis is probably responsible for the cholestatic liver disease of infancy, but the major symptoms of CTX in older children and adults are produced by accumulation of cholesterol and cholestanol in almost every tissue of the body, particularly in the nervous system, atherosclerotic plaques, and tendon xanthomata.

Genetics

CTX (OMIM 213700) is an autosomal recessive disorder caused by deficiency of sterol 27-hydroxylase (OMIM 604741), which is encoded by the CYP27A1 gene located on chromosome 2q33-qter. Many mutations have been described; prenatal diagnosis is usually undertaken by DNA testing.

Diagnostic Tests

Analysis of cholanoids (bile acids and bile alcohols) in urine shows a suggestive pattern dominated by the excretion of cholestane-tetrol glucuronides (m/z 611), and cholestane-pentol glucuronides (627). This pattern must be distinguished from a normal pattern in some infants in which the major cholanoids in urine (albeit at much lower concentration) are nor-cholestane pentol glucuronides (613) and cholestane-pentol glucuronides (627). Analysis of the urinary bile alcohols in an infant by GC-MS shows predominance of a 3,7,12,24,25-pentol, whereas in adults the major bile alcohols are 3,7,12,23,25- and 3,7,12,22,25-pentols. Analysis of plasma shows that at all ages, the concentration of 5β-cholestane-3α,7α,12α,25-tetrol is increased. In adults, the other major bile acid precursors detected are 5β-cholestane-3α,7α,12α-triol and 7α,12α-dihydroxy-cholest-4-en-3-one, whereas in infants the profile shows unusual cholestene-triols. In adults, the plasma concentration of cholestanol is usually increased, but this is not a reliable diagnostic marker. The full sterol profile shows increased concentrations of several cholesterol precursors. It is possible to measure 27-Hydroxylation of C27 sterols can be measured in cultured skin fibroblasts, and the enzyme activity is virtually absent in fibroblasts from patients with CTX.33

Treatment and Long-Term Outcome

Without treatment, the neonatal-onset liver disease can be fatal. Adults with untreated CTX usually die from progressive neurological dysfunction or myocardial infarction between the ages of 30 and 60 years. The treatment for CTX that has been thoroughly evaluated is chenodeoxycholic acid. The results of this treatment were first reported in 1984.34 The rates of cholestanol and cholesterol synthesis were reduced, and plasma cholestanol concentrations fell. A significant number of patients showed reversal of their neurological disability, with clearing of the dementia, improved orientation, a rise in intelligence quotient, and enhanced strength and independence. However, the MRI appearances do not show obvious improvement.35 Urinary excretion of bile-alcohol glucuronides is markedly suppressed. Chenodeoxycholic acid almost certainly works by suppressing cholesterol 7α-hydroxylase activity; ursodeoxycholic acid, which does not inhibit the enzyme, is ineffective. Adults have usually been treated with a dose of 750 mg/day of chenodeoxycholic acid. Other treatments include hydroxymethylglutaryl-coenzyme-A-reductase inhibitors (statins such as lovastatin)36 and low-density lipoprotein apheresis.37 There is currently insufficient information available to assess these forms of treatment. The osteoporosis seen in patients with CTX appears to be resistant to chenodeoxycholic-acid therapy.38 Cholestatic liver disease in infancy may be self-limiting, but in those children in whom it is not, bile acid treatment has been successful; cholic acid is probably preferable to chenodeoxycholic acid.24,25

Clinical Presentation

α-methyl-acyl-CoA racemase (AMACR) deficiency was first described in 2000.39 It can present in the neonatal period. In 2001, Van Veldhoven and colleagues described an infant with AMACR deficiency who presented with a coagulopathy due to vitamin K deficiency; a sibling had died of a major bleed with the same cause.40 The infant had mild cholestatic jaundice with raised aspartate aminotransferase and, in contrast to 3β-HSD deficiency, 5β-reductase deficiency, and CTX, had a raised γ-GT. Liver biopsy showed a mild nonspecific lymphocytic portal infiltrate and abundant giant cell transformation. Further details on this patient and the molecular basis of the disorder were published by Setchell and others in 2003.41 AMACR may also present with developmental delay in childhood and with epilepsy and postictal encephalopathy, which may result in permanent neurological deficit (eg, blindness, cognitive decline39,42,43). Adult patients have insidiously developed a pigmentary retinopathy, spastic paraparesis, tremor, and a demyelinating sensory motor neuropathy. MRI scans have revealed high signal in the pons, basal ganglia, thalami, cerebral peduncles, and subcortical white matter.

Metabolic Derangement and Pathophysiology

The neutral bile acid synthesis pathway produces the 25R isomers of DHCA-CoA and THCA-CoA. AMACR is required to convert these to the 25S isomers, which are the substrates for peroxisomal β-oxidation (eFig. 165.5). In patients with AMACR deficiency, the 25R isomers of DHCA and THCA accumulate as the free acids and the taurine conjugates. AMACR is also required for converting the 2R isomer of pristanoyl-CoA to the 2S isomer, which is the substrate for β-oxidation. Plasma concentrations of pristanic acid can be grossly elevated. High levels of phytanic acid cause neurological damage in Refsum’s disease, and it is quite likely that the very high levels of the (structurally similar) pristanic acid contribute to the neurological disease in AMACR deficiency.

Genetics

AMACR deficiency is an autosomal recessive disorder caused by mutations in the AMACR gene on chromosome 5p13.2-q11.1. One mutation (c.154T>C) has been found in four patients, including those with neonatal cholestasis and adult-onset neurological disease. There are some common polymorphisms in the coding region of the gene.

Diagnostic Tests

Analysis of urinary bile acids in a cholestatic infant revealed major peaks attributable to taurine conjugated THCA (m/z 556) and mono- and dihydroxylated derivatives (m/z 572 and 588). Analysis of plasma bile acids by GC-MS reveals increased concentrations of DHCA and THCA and the C29-dicarboxylic acid (3α,7α,12α-trihydroxy-27a,27b-dihomo-5b-cholestan-26,27b-dioic acid). HPLC-ESI-MS/MS can be used to show that it is the (25R) isomer of THCA that is accumulating. GC-MS analysis of fatty acids in plasma shows an elevated concentration of pristanic acid with mildly elevated/normal plasma phytanic acid concentration and normal very-long-chain fatty acids.

Treatment and Long-Term Outcome

The outcome for patients with AMACR deficiency has varied enormously from neonatal death caused by hemorrhage due to vitamin K deficiency to onset with difficulty in walking in the sixth decade. Obviously, parenteral vitamin K may be lifesaving in the neonatal period. One patient who presented with neonatal cholestasis was treated with cholic acid, which was associated with improved liver function tests.41 Reduction of plasma pristanic acid levels seems advisable. This can be achieved with a low phytanic acid diet, which involves, in particular, restriction of dairy products, beef, and lamb. It is unclear whether bile acid replacement therapy has a role in preventing neurological disease.

Clinical Presentation

Peroxisomal D-bifunctional protein (DBP) deficiency usually presents in the neonatal period with profound hypotonia and seizures.42 Other typical features include visual impairment; severe developmental delay; and a characteristic appearance of high forehead, high-arched palate, enlarged fontanel, long philtrum, epicanthic folds, hypertelorism, macrocephaly, retrognathia, and low-set ears. Brain imaging may show gross ventricular dilatation (29%), neocortical dysplasia (27%), cerebral demyelination (17%), and cerebellar atrophy (17%). Liver dysfunction is present in 26% of patients, and 43% have hepatomegaly. Postmortem examination of 11 patients showed polymicrogyria in 64% of them. Renal cysts and adrenal cortical atrophy were seen in 33% and 42% of autopsied cases, respectively. Less common features included delay of bone maturation, skeletal malformations, and calcific stippling.

Metabolic Derangement and Pathophysiology

Some of the early papers that describe DBP deficiency’s impact on bile acid synthesis were written when it was erroneously thought that the patients had a deficiency of the peroxisomal 3-oxoacyl-CoA thiolase.43-46

DBP catalyzes the second and third steps of peroxisomal β-oxidation of fatty acid derivatives; it has a hydroxyacyl-CoA dehydrogenase unit and an enoyl-CoA hydratase unit. It is involved in the β-oxidation of very-long-chain fatty acids, of pristanic acid, and of DHCA and THCA. Thus, the concentrations of all these substrates are often elevated in plasma, although the accumulation of pristanate is dependent upon the dietary intake of phytanate.

DBP-deficient patients can be classified as type I, deficient in both the hydratase and the dehydrogenase components; type II, deficient in only the hydratase unit; and type III, deficient in only the dehydrogenase unit. In patients with type III DBP deficiency, the major bile acid in plasma and bile may be 3α,7α,12α,24-tetrahydroxy-5β-cholestanoic acid (varanic acid; eFig. 165.6).

Genetics

DBP deficiency is an autosomal recessive disorder caused by mutations in the HSD17B4 gene on chromosome 5q2.47

Diagnostic Tests

Sometimes the urine bile acid profile is abnormal, showing the presence of taurine-conjugated tetrahydroxy cholestanoic acid (m/z 570) and taurine-conjugated tetrahydroxy cholestanoic acid (572).45 Analysis of plasma or duodenal bile acids may show unconjugated varanic acid (3α,7α,12α,24-tetrahydroxy-5β-cholestanoic acid) as a major component.45,46,48 This can distinguish patients with DBP deficiency from those with peroxisome biogenesis defects. THCA and DHCA are elevated in plasma in 74% of patients with DBP deficiency.42 A larger percentage of patients (94%) have elevated very-long-chain fatty acids; fewer (44%) have elevated pristanic acid. The diagnosis of DBP deficiency is achieved by measuring a range of peroxisome functions in cultured skin fibroblasts and proceeding to DNA analysis if necessary. These tests distinguish patients with DBP deficiency from patients with disorders of peroxisome biogenesis, who can be similar both clinically and biochemically.

Treatment and Long-Term Outcome

Most infants with DBP deficiency die before age 2 years; however, 12 reported patients survived beyond 2 years; five of these survived beyond 7.5 years.44 Biochemical analysis showed a clear correlation between peroxisomal beta-oxidation activity and survival.

Clinical Presentation

Only one patient has been described with this disorder49—a 45-year-old man who started stuttering at age 7 and who developed torticollis with a dystonic head tremor in stressful situations at age 17. At age 29, he was diagnosed with hypergonadotropic hypogonadism and azoospermia. Dystonia became progressively worse, and at age 44, cranial MRI showed bilateral hyperintense signals, butterfly-like lesions in the pons, and lesions in the occipital region.

Metabolic Derangement and Pathophysiology

Sterol carrier protein X is a thiolase active on branched chain CoA esters in the peroxisomes. It is involved in the β-oxidation of C27 bile acids and pristanic acid. Thus, it converts 3α,7α,12α-trihydroxy-24-oxo-5β-cholestanoyl-CoA and coenzyme A to cholyl-CoA and propionyl-CoA (eFig. 165.7). In SCPx deficiency, accumulating 3α,7α,12α-trihydroxy-24-oxo-5β-cholestanoic acid is thought to undergo decarboxylation to 3α,7α,12α-trihydroxy-27-nor-5β-cholestan-24-one. This in turn is thought to be hydroxylated two or three times to generate pentahydroxy-27-nor-5β-cholestan-24-one glucuronides and hexahydroxy-27-nor-5β-cholestan-24-one glucuronides. Reduction of the 24-oxo group probably generates 27-nor-5β-cholestane pentol and hexol glucuronides.

Genetics

Peroxisomal sterol carrier protein X deficiency is caused by mutations in the SCP2 gene on chromosome 1p32. The one patient described was homozygous for a one-nucleotide insertion (545_546insA).

Diagnostic Tests

The plasma concentrations of DHCA and THCA were very slightly elevated in plasma. Pristanate was considerably more elevated. The urine cholanoid profile was dominated by m/z 613 (attributed to 27-nor-5β-cholestane pentol glucuronides), 611 (pentahydroxy-27-nor-5β-cholestan-24-one glucuronides), 627 (hexahydroxy-27-nor-5β-cholestan-24-one glucuronides), and 629 (27-nor-5β-cholestane pentol glucuronides). These identities need to be confirmed by a method such as gas chromatography-mass spectrometry. It should be noted that the bile alcohol glucuronides with mass/charge ratio of 627 and 613 are excreted in quite large amounts by some normal infants. Care should be taken when interpreting the results from analysis of urinary cholanoids by electrospray/fast atom bombardment mass spectrometry.

Treatment and Long-Term Outcome

In the one described case, onset of neurological disease was in childhood, but progression was slow. No treatment has been attempted, but dietary control of high pristanate levels would seem warranted.

Clinical Presentation

The author’s experience with patients who have proven mutations in the bile acid-CoA: amino acid N-acyltransferase (BAAT) gene is limited to observations on a 3-month-old infant who had cholestatic jaundice, vitamin D deficiency, and a liver biopsy showing mild portal and focal lobular hepatitis. Among the Amish, the presentation is with failure to thrive, sometimes with pruritus and occasionally with coagulopathy but not jaundice.50 Two out of four affected patients suffered chronic upper respiratory infection. There are patients in the literature whose biochemical investigations indicate defects in amidation but in whom the molecular basis is uncertain; the defect could be in the bile acyl-CoA synthase (BACS) or in the bile acid-CoA: amino acid N-acyltransferase (BAAT).7

Metabolic Derangement and Pathophysiology

In BAAT deficiency, the liver produces unconjugated (nonamidated) bile acids instead of the usual glycine and taurine conjugates of CA and CDCA. The major urinary bile acid is unconjugated cholic acid. CA and CDCA are also excreted as sulfate and glucuronide conjugates.

Genetics

BAAT deficiency is an autosomal recessive disorder caused by mutations in the BAAT gene on chromosome 9q22.3. The disorder in the Amish is caused by homozygosity for a missense mutation (226 A>G; M76V) in the BAAT gene.

Diagnostic Tests

Analysis of urine by negative ion FAB-MS or ESI-MS shows that the major urinary bile acid is an unconjugated trihydroxy-cholanoic acid (m/z 407); GC-MS shows that it is unconjugated cholic acid. Other bile acids that may be detected include sulfated dihydroxycholanoic acid(s) (m/z 471), trihydroxycholanoic acids (m/z 487), dihydroxycholanoic acid(s), and trihydroxycholanoic acid(s) (m/z 567 and 583).

Treatment and Long-Term Outcome

Treatment of vitamin K deficiency may be lifesaving, and treatment of rickets may require 1α-hydroxycholecalciferol or 1,25-dihydroxycholecalciferol. The Amish patients probably had improvement in symptoms with ursodeoxycholic acid, but it is important to note that familial hypercholanemia of the Amish can be caused by defects in a gene responsible for integrity of tight junctions (TJP2) and by mutations in the BAAT gene.50 The one patient monitored by the author had complete resolution of hepatitis in infancy with the help of ursodeoxycholic acid treatment and is asymptomatic at age 3 years.

Bile acyl-CoA synthase (BACS) deficiency is a recently discovered disorder; details will be added to the online version of this chapter when the molecular basis is firmly established.

Clinical Presentation

The first case of oxysterol 7α-hydroxylase deficiency was described in 1998 by Setchell and colleagues.51 This infant, who had severe cholestatic liver disease but normal γ-GT, did not respond to bile acid replacement therapy, required liver transplantation, and died of complications of the transplant.

Recently, mutations in the CYP7B1 gene encoding oxysterol 7α-hydroxylase have been shown in patients with hereditary spastic paraplegia type 5 (HSP5).52 Affected patients have lower-extremity weakness and spasticity, posterior column sensory impairment (as evidenced by diminished vibration sensation and proprioception), and some degree of bladder dysfunction. The age of onset varies from 1 to 40 years. Thus, this seems to be another disorder that can cause cholestasis in the neonate and neurological disease later in life.

Metabolic Derangement and Pathophysiology

In the liver, oxysterol 7α-hydroxylase is essential for the acidic pathway of bile acid synthesis. Deficiency of the enzyme leads to synthesis of 3β-hydroxy-5-cholenoic acid (eFig. 165.8), which has cholestatic and hepatotoxic properties. In extrahepatic tissues, oxysterol 7α-hydroxylase contributes to a pathway for cholesterol degradation, and it provides the primary metabolic route for the modification of dehydroepiandrosterone neurosteroids in the brain. Loss of one or both of these functions may explain the neurological disease.

Genetics

Oxysterol 7α-hydroxylase deficiency is an autosomal recessive disorder caused by mutations in the CYP7B1 gene on chromosome 8q21.3. The child with liver disease was homozygous for an R388X nonsense mutation,51 as was one patient with HSP5. Other patients with HSP5 have had missense mutations.52

Diagnostic Tests

In the reported neonatal case, FAB-MS analysis of urine revealed major peaks of m/z ratio 453 and 510 attributable to 3β-hydroxy-5-cholenoic acid 3-sulfate and its glycine conjugate. GC-MS analysis of plasma indicated that the main bile acids were 3β-hydroxy-5-cholenoic acid and 3β-hydroxy-5-cholestenoic acid the concentration of 27-hydroxycholesterol in plasma was very markedly increased. We do not yet know whether these metabolic abnormalities will be present in adults with HSP5.

Treatment and Long-Term Outcome

The one reported patient who presented with cholestatic liver disease in infancy showed a deterioration with ursodeoxycholic acid and no improvement with cholic acid and required a liver transplant for hepatic failure at 4.5 months. The fact that homozygous mutations have been found in patients with HSP5 suggests that some individuals with oxysterol 7α-hydroxylase deficiency either do not develop cholestatic liver disease or have mild and self-limiting liver dysfunction. No specific treatment has been described.

Disorders of peroxisome biogenesis are discussed in Chapter 162. As inborn errors affecting bile acid synthesis, they can be detected in infancy by analysis of urinary bile acids; this usually shows substantial excretion of taurine-conjugated tetrahydroxy cholestanoic acids (mz 572). Analysis of plasma bile acids is a useful test for diagnosing an older child with a suspected peroxisomal disorder; the plasma typically contains the C27 bile acids, THCA and DHCA, and the C29-dicarboxylic acid. The analysis of plasma bile acids and pristanate in a child with normal very-long-chain fatty acids may reveal a peroxisome biogenesis defect. For example, the patient described in reference 53 was eventually shown to have mutations in the Pex 10 gene (eTable 165.1).53