Biochemical Genetics

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CHAPTER 11 Biochemical Genetics

Life … is a relationship between molecules.

LINUS PAULING

The existence of chemical individuality follows of necessity from that of chemical specificity, but we should expect the differences between individuals to be still more subtle and difficult of detection.

ARCHIBALD GARROD (1908)

In this chapter, we consider single-gene biochemical or metabolic diseases, including mitochondrial disorders. The range of known disorders is vast, so only an overview is possible, but it is hoped that the reader will gain a flavor of this fascinating area of medicine. At the beginning of the twentieth century, Garrod introduced the concept of ‘chemical individuality’, leading in turn to the concept of the inborn error of metabolism (IEM). Beadle and Tatum later developed the idea that metabolic processes, whether in humans or any other organism, proceed by steps. They proposed that each step was controlled by a particular enzyme and that this, in turn, was the product of a particular gene. This was referred to as the one gene–one enzyme (or protein) concept.

Inborn Errors of Metabolism

In excess of 200 IEMs are known that can be grouped by either the metabolite, metabolic pathway, function of the enzyme, or cellular organelle involved (Table 11.1). Most follow autosomal recessive or X-linked recessive inheritance, with only a few being autosomal dominant. This is because the defective protein in most cases is a diffusible enzyme, and there is usually sufficient residual activity in the heterozygous state (loss-of-function, see p. 26) for the enzyme to function normally in most situations. If, however, the reaction catalysed by an enzyme is rate limiting (haploinsufficiency, see p. 26) or the gene product is part of a multimeric complex (dominant-negative, see p. 26), the disorder can manifest in the heterozygous state and follow dominant inheritance (p. 109).

Table 11.1 Characteristics of Some Inborn Errors of Metabolism

Disorders of Amino Acid Metabolism

There are a number of disorders of amino acid metabolism, the best known of which is phenylketonuria.

Phenylketonuria

Children with phenylketonuria (PKU), if untreated, are severely intellectually impaired and often develop seizures. There is a deficiency of the enzyme required for the conversion of phenylalanine to tyrosine, phenylalanine hydroxylase (PAH)—causing a ‘genetic block’ in the metabolic pathway (Figure 11.1).

FIGURE 11.1 Sites of ‘biochemical block’ in phenylketonuria, alkaptonuria, congenital hypothyroidism, and oculocutaneous albinism.

PKU was the first genetic disorder in humans shown to be caused by a specific enzyme deficiency, by Jervis in 1953. As a result of the enzyme defect, phenylalanine accumulates and is converted into phenylpyruvic acid and other metabolites that are excreted in the urine. The enzyme block leads to a deficiency of tyrosine, with a consequent reduction in melanin formation, and children therefore often have blond hair and blue eyes (Figure 11.2). In addition, areas of the brain that are usually pigmented, such as the substantia nigra, may also lack pigment.

FIGURE 11.2 Facies of a male with phenylketonuria; note the fair complexion.

Treatment of PKU

An obvious method of treating children with PKU would be to replace the missing enzyme, but this is not simply achieved (p. 349). Bickel, just 1 year after the enzyme deficiency had been identified, suggested that PKU could be treated by removal of phenylalanine from the diet and this has proved effective. If PKU is detected early enough in infancy, intellectual impairment can be prevented by giving a phenylalanine restricted diet. Phenylalanine is an essential amino acid and therefore cannot be removed entirely from the diet. By monitoring the level of phenylalanine in the blood, it is possible to supply sufficient amounts to meet normal requirements but avoiding toxic levels, resulting in mental retardation. After brain development is complete, dietary restriction can be relaxed—from adolescence onward.

The intellectual impairment seen in children with phenylketonuria is likely due to toxic levels of phenylalanine, and/or its metabolites, rather than a deficiency of tyrosine, of which adequate amounts are present in a normal diet. Both prenatal and postnatal factors may be responsible for developmental delay in untreated PKU.

Alkaptonuria

Alkaptonuria was the original autosomal recessive IEM described by Garrod. Here there is a block in the breakdown of homogentisic acid, a metabolite of tyrosine, because of a deficiency of the enzyme homogentisic acid oxidase (see Figure 11.1). As a consequence, homogentisic acid accumulates and is excreted in the urine, which then darkens on exposure to air. Dark pigment is also deposited in certain tissues, such as the ear wax, cartilage, and joints, where it is known as ochronosis, which in joints can lead to arthritis later in life.

Oculocutaneous Albinism

Oculocutaneous albinism (OCA) is an autosomal recessive disorder resulting from a deficiency of the enzyme tyrosinase, which is necessary for the formation of melanin from tyrosine (see Figure 11.1). In OCA there is a lack of pigment in the skin, hair, iris, and ocular fundus (Figure 11.3), and the lack of eye pigment results in poor visual acuity and uncontrolled pendular eye movements—nystagmus. Reduced fundal pigmentation leads to underdevelopment of part of retina for fine vision—the fovea—and abnormal projection of the visual pathways to the optic cortex.

FIGURE 11.3 Oculocutaneous albinism in a child of Afro-Caribbean origin.

(Courtesy of Dr V. A. McKusick.)

Heterogeneity of OCA

OCA is genetically and biochemically heterogeneous. Cells from those with classic albinism have no measurable tyrosinase activity, the so-called tyrosinase-negative form. However, cells from some persons with albinism show reduced but residual tyrosinase activity and are termed tyrosinase positive. This is usually reflected clinically by variable development of pigmentation of their hair and skin with age. Both types are known as tyrosinase gene-related OCA type 1.

OCA type 1 is due to mutations in the tyrosinase gene on chromosome 11q. However, linkage studies in some families with tyrosinase-positive OCA have excluded the tyrosinase gene. Some of these have a mutation in the P gene, the human homolog of a mouse gene called pink-eyed dilution, or ‘pink-eye’, located on chromosome 15q. This has been termed OCA type 2. In addition, in a proportion of families with OCA, linkage to both of these two loci has been excluded, consistent with the existence of a third locus responsible for OCA.

Homocystinuria

Homocystinuria is a recessively inherited sulfur amino-acid IEM characterized by learning disability, seizures, thrombophilia, osteoporosis, scoliosis, pectus excavatum, long fingers and toes (arachnodactyly), and a tendency to dislocation of the lenses. The somatic features therefore resemble the autosomal dominant disorder Marfan syndrome (p. 300).

Homocystinuria results from deficiency of the enzyme cystathionine β-synthase and can be screened for by means of a positive cyanide nitroprusside test, which detects the presence of increased levels of homocystine in the urine. The diagnosis is confirmed by raised plasma homocystine levels. Treatment involves a low-methionine diet with cystine supplementation. A proportion of individuals with homocystinuria are responsive to the enzyme cofactor pyridoxine (i.e., the pyridoxine-responsive form). A small proportion of affected individuals have mutations in genes leading to deficiencies of enzymes involved in the synthesis of cofactors for cystathionine β-synthase.

Disorders of Branched-Chain Amino Acid Metabolism

The essential branched-chain amino acids leucine, isoleucine and valine have a part of their metabolic pathways in common. Deficiency of the enzyme involved results in maple syrup urine disease.

Maple Syrup Urine Disease

Newborn infants with this autosomal recessive disorder present in the first week of life with vomiting, then alternating tone, leading to death within a few weeks if untreated. The name derives from the odor of the urine—likened to that of maple syrup. The disorder is caused by a deficiency of the branched-chain ketoacid decarboxylase, producing increased urinary excretion of the branched-chain amino acids valine, leucine, and isoleucine, the presence of which suggests the diagnosis, confirmed by demonstration of the three essential branched-chain amino acids in blood. Treatment involves a diet restricting the intake of these three amino acids to the amounts necessary for growth. Affected individuals are particularly susceptible to deterioration, particularly in association with intercurrent illnesses leading to catabolic protein degradation.

Urea Cycle Disorders

The urea cycle is a five-step metabolic pathway that takes place primarily in liver cells for the removal of waste nitrogen from the amino groups of amino acids arising from the normal turnover of protein. It converts two molecules of ammonia and one of bicarbonate into urea (Figure 11.4). Deficiencies of enzymes in the urea cycle result in intolerance to protein from the accumulation of ammonia in the body—hyperammonemia. Increased ammonia levels are toxic to the central nervous system and can lead to coma and, with some untreated urea cycle disorders, death. They are collectively and individually rare and, with the exception of X-linked ornithine transcarbamylase deficiency, inherited as autosomal recessive traits.

FIGURE 11.4 Diagram indicating the position of the various inborn errors of the urea cycle.

Disorders of Carbohydrate Metabolism

The inborn errors of carbohydrate metabolism can be considered in two main groups: disorders of monosaccharide metabolism and the glycogen storage disorders.

Disorders of Monosaccharide Metabolism

Two examples of disorders of monosaccharide metabolism are galactosemia and hereditary fructose intolerance.

Galactosemia

Galactosemia is an autosomal recessive disorder resulting from a deficiency of the enzyme galactose 1-phosphate uridyl transferase, necessary for the metabolism of the dietary sugar galactose. Newborns with galactosemia present with vomiting, lethargy, failure to thrive, and jaundice in the second week of life. If untreated, they develop complications that include mental retardation, cataracts, and liver cirrhosis. Complications can be prevented by early diagnosis and feeding infants with milk substitutes that do not contain galactose or lactose—the sugar found in milk that is broken down into galactose. Early diagnosis is essential and galactosemia can be screened for by the presence of reducing substances in the urine with specific testing for galactose.

Hereditary Fructose Intolerance

Hereditary fructose intolerance is an autosomal recessive disorder resulting from a deficiency of the enzyme fructose 1-phosphate aldolase. Dietary fructose is present in honey, fruit, and certain vegetables, and in combination with glucose in the disaccharide sucrose in cane sugar. Individuals with hereditary fructose intolerance present at different ages, depending on when fructose is introduced into the diet. Symptoms can be minimal but might also be as severe as those seen in galactosemia, which include failure to thrive, vomiting, jaundice, and seizures. The diagnosis is confirmed by the presence of fructose in the urine and enzyme assay on an intestinal mucosal or a liver biopsy sample. Dietary restriction of fructose is associated with a good long-term prognosis.

Glycogen Storage Diseases

Glycogen is the form in which the sugar glucose is stored in muscle and liver as a polymer, acting as a reserve energy source. In the glycogen storage diseases (GSDs) glycogen accumulates in excessive amounts in skeletal muscle, cardiac muscle, and/or liver because of a variety of inborn errors of the enzymes involved in synthesis and degradation of glycogen. In addition, because of the metabolic block, glycogen is unavailable as a normal glucose source. This can result in hypoglycemia, impairment of liver function and neurological abnormalities.

In each of the six major types of GSD, there is a specific enzyme defect involving one of the steps in the metabolic pathways of glycogen synthesis or degradation. The various types can be grouped according to whether they affect primarily the liver or muscle. All six types are inherited as autosomal recessive disorders, although there are variants of the hepatic phosphorylase that are X-linked.

Glycogen Storage Diseases that Primarily Affect Liver

von Gierke disease (GSD-I)

von Gierke disease was the first described disorder of glycogen metabolism and results from a deficiency of the enzyme glucose-6-phosphatase, which is responsible for degradation of liver glycogen to release glucose. Infants present with an enlarged liver (hepatomegaly) and/or sweating and a fast heart rate due to hypoglycemia, which can occur after fasting of only 3 to 4 hours duration. Treatment is simple—frequent feeding and avoidance of fasting to maintain the blood sugar concentration.

Cori disease (GSD-III)

Cori disease is caused by deficiency of the enzyme amylo-1,6-glucosidase, which is also known as the debrancher enzyme. Deficiency results in glycogen accumulation in the liver and other tissues because of the inability to cleave the ‘branching’ links of the glycogen polymer. Infants may present with hepatomegaly because of glycogen accumulation and/or muscle weakness. Treatment involves avoiding hypoglycemia by frequent feeding and avoiding prolonged periods of fasting.

Anderson disease (GSD-IV)

Anderson disease results from deficiency of glycogen brancher enzyme leading to the formation of abnormal glycogen consisting of long chains with few branches that cannot be broken down by the enzymes normally responsible for glycogen degradation. Infants present with hypotonia and abnormal liver function in their first year, progressing rapidly to liver failure. No effective treatment is available apart from the possibility of a liver transplant.

Hepatic phosphorylase deficiency (GSD-VI)

Hepatic phosphorylase is a multimeric enzyme complex with subunits coded for by both autosomal and X-linked genes. Deficiency of hepatic phosphorylase obstructs glycogen degradation, which results in children presenting in the first 2 years of life with hepatomegaly, hypoglycemia, and failure to thrive. Treatment is with carbohydrate supplementation.

Glycogen Storage Diseases that Primarily Affect Muscle

Pompe disease (GSD-II)

Infants with Pompe disease usually present in the first few months of life with floppiness (hypotonia) and delay in the gross motor milestones because of muscle weakness. They then develop an enlarged heart and die from cardiac failure in the first or second year. Voluntary and cardiac muscle accumulates glycogen because of a deficiency of the lysosomal enzyme α-1,4-glucosidase, which is needed to break down glycogen. The diagnosis can be confirmed by enzyme assay of white blood cells or fibroblasts. Early reports of enzyme replacement therapy appear promising.

McArdle disease (GSD-V)

People with McArdle disease present with muscle cramps on exercise in the teenage years. The condition is caused by a deficiency of muscle phosphorylase, which is necessary for degradation of muscle glycogen. There is no effective treatment, although in some the muscle cramps tend to decline if exercise is continued, probably as a result of other energy sources becoming available from alternative metabolic pathways.

Disorders of Steroid Metabolism

The disorders of steroid metabolism include a number of autosomal recessive inborn errors of the biosynthetic pathways of cortisol. Virilization of a female fetus may occur together with salt loss in infants of either sex from a deficiency of the hormone aldosterone. In addition, defects of the androgen receptor result in lack of virilization of chromosomally male individuals (Figure 11.5).

FIGURE 11.5 Steroid biosynthesis indicating the site of the common inborn errors of steroid biosynthesis.

Congenital Adrenal Hyperplasia (Adrenogenital Syndrome)

The diagnosis of congenital adrenal hyperplasia (CAH) should be considered in any newborn female infant presenting with virilization of the external genitalia, because this is the most common cause of ambiguous genitalia in female newborns (p. 288) (Figure 11.6). 21-Hydroxylase deficiency accounts for more than 90% of cases. Approximately 25% have the salt-losing form, presenting in the second or third week of life with circulatory collapse, hyponatremia, and hyperkalemia. Less commonly, CAH is a result of deficiency of the enzymes 11β-hydroxylase or 3β-dehydrogenase, and very rarely occurs as a result of deficiencies of enzymes 17α-hydroxylase and 17,20-lyase. Desmolase deficiency is very rare, with all pathways blocked, causing a reversed phenotype of ambiguous genitalia in males, and severe addisonian crises. Males with the rare 5α-reductase deficiency are significantly under-masculinized but do not suffer other metabolic problems and are likely to be raised as females. At puberty, however, the surge in androgen production is sufficient to stimulate growth of the phallus, making gender identity and assignment problematic.

FIGURE 11.6 A, Virilized external genitalia in a female with congenital adrenal hyperplasia. B, A male baby with hypospadias who clearly has testes in the scrotal sacs.

Affected females with classic CAH are virilized from accumulation of the adrenocortical steroids proximal to the enzyme block in the steroid biosynthetic pathway, many of which have testosterone-like activity (see Figure 11.5). However, they have normal müllerian-derived internal organs. The possibility of CAH should not be forgotten, of course, in male infants presenting with circulatory collapse in the first few weeks of life.

Affected infants, in addition to requiring urgent correct assignment of gender, are treated with replacement cortisol, along with fludrocortisone if they have the salt-losing form. Virilized females may require plastic surgery later. Steroid replacement is lifelong and should be increased during intercurrent illness or stress, such as surgery. Menarche in girls with salt-losing CAH is late, menstruation irregular, and they are subfertile.

Androgen Insensitivity Syndrome

Individuals with the androgen insensitivity syndrome have female external genitalia and undergo breast development in puberty (p. 288). They classically present either with primary amenorrhea or with an inguinal hernia containing a gonad that turns out to be a testis. Inguinal hernia is uncommon in girls and if present, especially if bilateral, androgen insensitivity syndrome should be considered. There is often scanty secondary sexual hair and investigation of the internal genitalia reveals an absent uterus and fallopian tubes with a blind-ending vagina. Chromosome analysis reveals a normal male karyotype, 46,XY.

Androgen production by the testes is normal but androgen does not bind normally because of an abnormal androgen receptor (see Figure 11.5)—the androgen receptor gene on the X chromosome is mutated. This can be functionally assayed in skin fibroblasts. Some individuals have incomplete or partial androgen insensitivity, and under-virilization is variable. Affected subjects may have a female sexual orientation and are sterile. Testes must be removed because of an increased malignancy risk, and estrogen given for secondary sexual development and prevention of long-term osteoporosis.

Disorders of Lipid Metabolism

Familial hypercholesterolemia is the most common autosomal dominant single-gene disorder in Western society and is associated with high morbidity and mortality rates through premature coronary artery disease (p. 241).

Familial Hypercholesterolemia

Persons with familial hypercholesterolemia (FH) have raised cholesterol levels with a significant risk of developing early coronary artery disease (p. 241). They can present in childhood or adolescence with subcutaneous deposition of lipid, known as xanthomata (Figure 11.7). Starting with families who presented with early coronary artery disease, Brown and Goldstein unraveled the biology of the low-density lipoprotein (LDL) receptor (p. 241) and the pathological basis of FH.

FIGURE 11.7 Legs of a person homozygous for familial hypercholesterolemia, showing multiple xanthomata.

(Courtesy Dr. E. Wraith, Royal Manchester Children’s Hospital, Manchester, UK.)

Cells normally derive cholesterol from either endogenous synthesis or dietary uptake from LDL receptors on the cell surface. Intracellular cholesterol levels are maintained by a feedback system, with free cholesterol inhibiting LDL receptor synthesis as well as reducing the level of de novo endogenous synthesis.

High cholesterol levels in FH are due to deficient or defective function of the LDL receptors leading to increased levels of endogenous cholesterol synthesis. Four main classes of mutation in the LDL receptor have been identified: (1) reduced or defective biosynthesis of the receptor; (2) reduced or defective transport of the receptor from the endoplasmic reticulum to the Golgi apparatus; (3) abnormal binding of LDL by the receptor; and (4) abnormal internalization of LDL by the receptor (Figure 11.8). Specific mutations are more prevalent in certain ethnic groups because of founder effects (p. 133).

FIGURE 11.8 Stages in cholesterol biosynthesis and in the metabolism of low-density lipoprotein (LDL) receptors, indicating the types of mutation in familial hypercholesterolemia.

(Adapted from Brown MS, Goldstein JL 1986 A receptor-mediated pathway for cholesterol homeostasis. Science 232:34–47.)

The mainstay of management is dietary restriction of cholesterol intake and drug treatment with ‘statins’ that reduce the endogenous synthesis of cholesterol by inhibiting the enzyme 3-hydroxy-3-methylglutaryl coenzyme A (CoA) reductase. Cholesterol levels in affected families are variable and lipid assays do not necessarily identify those with mutations. There is therefore interest in the introduction of widespread genetic testing, though most mutations are missense, which may pose problems of interpretation.

Lysosomal Storage Disorders

In addition to the IEMs in which an enzyme defect leads to deficiency of an essential metabolite and accumulation of intermediate metabolic precursors, there are a number of disorders in which deficiency of a lysosomal enzyme involved in the degradation of complex macromolecules leads to their accumulation. This accumulation occurs because macromolecules are normally in a constant state of flux, with a delicate balance between their rates of synthesis and breakdown. Children born with lysosomal storage diseases are usually normal initially but with the passage of time commence a downhill course of variable duration owing to the accumulation of one or more of a variety or type of macromolecules.

Mucopolysaccharidoses

Children with one of the mucopolysaccharidoses (MPSs) present with skeletal, vascular, or central nervous system findings along with coarsening of the facial features. These features are due to progressive accumulation of sulfated polysaccharides (also known as glycosaminoglycans) caused by defective degradation of the carbohydrate side-chain of acid mucopolysaccharide.

Six different MPSs are recognized, based on clinical and genetic differences. Each specific MPS type has a characteristic pattern of excretion in the urine of the glycosaminoglycans, dermatan, heparan, keratan, and chondroitin sulfate. Subsequent biochemical investigation has revealed the various types to be due to deficiency of different individual enzymes. All but Hunter syndrome, which is X-linked, are autosomal recessive disorders.

Hurler Syndrome (MPS-I)

Hurler syndrome is the most severe MPS. Infants present in the first year with corneal clouding, a characteristic curvature of the lower spine and subsequent poor growth. They develop hearing loss, coarse facial features, an enlarged liver and spleen, joint stiffness, and vertebral changes in the second year. These features progress together with mental deterioration and eventually death by mid-adolescence from a combination of cardiac failure and respiratory infections.

The diagnosis of Hurler syndrome was initially made by demonstrating the presence of metachromatic granules in the cells (i.e., lysosomes distended by the storage material that is primarily dermatan sulfate). Increased urinary excretion of dermatan and heparan sulfate is commonly used as a screening test, but confirmation of the diagnosis involves demonstration of reduced activity of the lysosomal hydrolase, α-l-iduronidase, and direct gene analysis. Less severe allelic forms of Hurler syndrome, caused by varying levels of residual α-l-iduronidase activity, were previously classified separately as Scheie disease (MPS-I S) and Hurler/Scheie disease (MPS-I H/S).

Hunter Syndrome (MPS-II)

Males with Hunter syndrome usually present between 2 and 5 years with hearing loss, recurrent infections, diarrhea, and poor growth. Facial features are characteristic with coarsening (Figure 11.9), the liver and spleen are enlarged, and joint stiffness occurs. Spinal radiographs show abnormal shape of the vertebrae. Progressive physical and mental deterioration occurs with death usually in adolescence.

FIGURE 11.9 Facies of a male with the mucopolysaccharidosis, Hunter syndrome.

(Courtesy Dr. E. Wraith, Royal Manchester Children’s Hospital, Manchester, UK.)

The diagnosis is confirmed by the presence of excess amounts of dermatan and heparan sulfate in the urine, deficient or decreased activity of the enzyme iduronate sulfate sulfatase in serum or white blood cells, and direct gene analysis.

Sanfilippo Syndrome (MPS-III)

Sanfilippo syndrome is the most common MPS. Affected individuals present in their second year with mild coarsening of features, skeletal changes, and progressive intellectual loss with behavioral problems, seizures, and death in early adult life. The diagnosis is confirmed by the presence of increased urinary heparan and chondroitin sulfate excretion, and deficiency of one of four enzymes involved in the degradation of heparan sulfate: sulfaminidase (MPS-III A), N-acetyl-α-d-glucosaminidase (MPS-III B), acetyl-CoA-α-glucosaminidase-N-acetyltransferase (MPS-III C), or N-acetyl-glucosamine-6-sulfate sulfatase (MPS-III D). Individuals with these different enzyme deficiencies cannot be distinguished clinically. Direct gene testing is not so reliable.

Morquio Syndrome (MPS-IV)

Children with Morquio syndrome present age 2 to 3 years with short stature, thoracic deformity, and curvature of the spine (kyphoscoliosis). Intelligence is normal and survival is long term, but there is a risk of spinal-cord compression from progression of the skeletal involvement. The diagnosis is confirmed by the presence of keratan sulfate in the urine and deficiency of either galactosamine-6-sulphatase (MPS-IV A) or β-galactosidase (MPS-IV B).

Maroteaux–Lamy Syndrome (MPS-VI)

This MPS presents with Hurler-like features in early childhood, including coarse facial features, short stature with thoracic deformity, kyphosis, and restriction of joint mobility. In addition, corneal clouding and cardiac valve abnormalities develop; intelligence is normal. A milder form presents later with survival into late adulthood, in contrast to the severe form in which survival is usually only to the third decade. The diagnosis is confirmed by the presence of increased urinary dermatan sulfate excretion and arylsulfatase B deficiency in white blood cells or fibroblasts.

Sly Syndrome (MPS-VII)

This is an extremely variable MPS. Presentation ranges from skeletal features that include mild kyphoscoliosis and hip dysplasia to coarse facial features, hepatosplenomegaly, corneal clouding, cardiac abnormalities, and mental retardation, with death in childhood or adolescence. Increased urinary glycosaminoglycans excretion and β-glucuronidase deficiency in serum, white blood cells, or fibroblasts confirm the diagnosis.

Treatment of the MPSs

Treatment of these disorders by enzyme replacement has proved difficult in practice (p. 349). However, bone marrow transplantation has been attempted with varying success, biochemically, and clinically in relation to the skeletal and cerebral features.

Sphingolipidoses (Lipid Storage Diseases)

In the sphingolipidoses, there is an inability to degrade sphingolipid, resulting in the progressive deposition of lipid or glycolipid, primarily in the brain, liver, and spleen. Central nervous system involvement results in progressive mental deterioration, often with seizures, leading to death in childhood. There are at least 10 different types, with specific enzyme deficiencies, Tay-Sachs, Gaucher, and Niemann-Pick diseases being the most common.

Tay-Sachs Disease

This well-known sphingolipidosis has an incidence of ~1:3600 in Ashkenazi Jews (pp. 313–314). Infants usually present by 6 months of age with poor feeding, lethargy, and floppiness. Developmental regression usually becomes apparent in late infancy, feeding becomes increasingly difficult, and the infant progressively deteriorates, with deafness, visual impairment, and spasticity, which progresses to rigidity. Death usually occurs by the age of 3 years from respiratory infection. Less severe juvenile, adult, and chronic forms are reported.

The diagnosis is supported clinically by the presence of a ‘cherry-red’ spot in the center of the macula of the fundus. Biochemical confirmation of Tay-Sachs disease is by demonstration of reduced hexosaminidase A levels in serum, white blood cells or cultured fibroblasts, and direct gene analysis is available. Reduced hexosaminidase A activity is due to deficiency of the a subunit of the enzyme β-hexosaminidase that leads to accumulation of the sphingolipid GM₂ ganglioside. This deficiency leads to reduced activity of the isozyme, hexosaminidase B, causing the other GM₂ gangliosidosis, Sandhoff disease, which presents with similar clinical features.

Gaucher Disease

This is the most common sphingolipidosis and, as with Tay-Sachs, is relatively more frequent among Ashkenazi Jews. There are two main types based on the age of onset.

Type I, with adult onset, is the more common form and presents with febrile episodes, pain in limbs, joints, or trunk, and a tendency to pathological fractures. Clinical examination usually reveals hepatosplenomegaly and investigations show mild anemia and radiological changes in the vertebrae and proximal femora. The central nervous system is spared.

In type II, infantile Gaucher disease, central nervous system involvement is a major feature and presents age 3 to 6 months with failure to thrive and hepatosplenomegaly. By 6 months, developmental regression and neurological deterioration occur with spasticity and seizures. Recurrent pulmonary infections cause death in the second year.

The diagnosis is confirmed by reduced activity of the enzyme glucosylceramide β-glucosidase in white blood cells or cultured fibroblasts.

Treatment in type 1 involves symptomatic analgesia, and sometimes splenectomy to prevent premature sequestration of red blood cells (hypersplenism). Initial attempts to treat adults by enzyme replacement therapy met with little success because of difficulty in obtaining sufficient quantities of enzyme and in targeting the appropriate sites. However, modification of β-glucosidase by the addition of mannose 6-phosphate, which targets the enzyme to macrophage lysosomes, has led to dramatic alleviation of symptoms and regression of organomegaly. The treatment is expensive, and regimens using lower doses and alternative methods to target the enzyme may be more rational.

Niemann-Pick Disease

Infants with Niemann-Pick disease present with failure to thrive and hepatomegaly, and a cherry-red spot may be found on their macula. Developmental regression progresses rapidly by the end of the first year, with death by 4 years of age. A characteristic finding is the presence of what are called foam cells in the bone marrow from sphingomyelin accumulation. Confirmation of the diagnosis is by demonstration of deficiency of the enzyme sphingomyelinase. A less severe form without neurological involvement has been reported. As with Tay-Sachs and Gaucher diseases, it is more common in Ashkenazi Jews from eastern Europe.

Disorders of Purine/Pyrimidine Metabolism

Gout is the classic disorder of abnormal purine metabolism. Joint pain, swelling, and tenderness are a result of the inflammatory response of the body to deposits of crystals of a salt of uric acid. In fact, only a minority of persons with gout have an IEM. The cause in most instances results from a combination of genetic and environmental factors; however, it is always important to consider disorders that can result in an increased turnover of purines (e.g., a malignancy such as leukemia) or reduced secretion of the metabolites (e.g., renal impairment) as a possible underlying precipitating cause.

Lesch-Nyhan Syndrome

This is a particularly disabling disorder of purine metabolism, follows X-linked inheritance, and is due to the deficiency of the enzyme hypoxanthine guanine phosphoribosyltransferase, that results in increased levels of phosphoribosylpyrophosphate. The latter is normally a rate-limiting chemical in the synthesis of purines. Excess amounts lead to an increased rate of purine synthesis and accumulation of uric acid and some of its metabolic precursors. The main effect is neurological, with uncontrolled movements, spasticity, mental retardation, and compulsive self-mutilation. Although drugs such as allopurinol, which inhibit uric acid formation, can lower uric acid levels, none is highly satisfactory.

Immunodeficiency Diseases Caused By Defects in Purine Metabolism

Two inherited immunodeficiency disorders (pp. 203–204), perhaps surprisingly, are inborn errors of purine metabolism.

Adenosine Deaminase Deficiency

About half of all children with the autosomal recessive form of severe combined immunodeficiency with impaired B- and T-cell function (p. 203) have deficiency of the enzyme adenosine deaminase. Presentation is in infancy with recurrent viral and bacterial infections which, if untreated, soon cause death from overwhelming infection. The diagnosis is confirmed by deficient red blood cell adenosine deaminase activity. Bone marrow transplantation has been successful—even for the fetus in utero.

Purine Nucleoside Phosphorylase Deficiency

A proportion of children susceptible to severe, recurrent, and potentially fatal viral infections with isolated impaired T-cell function have been shown to have a deficiency of the enzyme purine nucleoside phosphorylase. Treatment with irradiated red blood cells has been reported to result in a temporary improvement in immune function.

Disorders of Porphyrin Metabolism

There are several different disorders of porphyrin metabolism that are from a deficiency of enzymes in the biosynthetic pathway of the iron-containing group in hemoglobin—heme (p. 155). They all follow autosomal dominant inheritance, with the exception of autosomal recessive congenital erythropoietic porphyria. This is because the enzymes are rate limiting (p. 26), so that haploinsufficiency results in clinical disease.

The different types of porphyria are variably associated with neurological or visceral involvement and cutaneous photosensitivity from an accumulation of the different porphyrin precursors in those organs. The porphyrias are divided into two types depending on whether the excess production of porphyrins occurs predominantly in the liver or in the erythropoietic system.

Hepatic Porphyrias

These include acute intermittent porphyria, hereditary co-proporphyria, and porphyria variegata.

Acute Intermittent Porphyria

Acute intermittent porphyria is characterized by attacks of abdominal pain, weakness, vomiting and mental disturbance in the form of confusion, emotional upset, or hallucinations. Even coma may occur, and women are more severely affected than men, with symptoms sometimes associated with the menstrual cycle. Attacks can also be precipitated by the administration of certain drugs, such as exogenous steroids, anticonvulsants, and barbiturates. It is caused by a partial deficiency of the enzyme uroporphyrinogen I synthase leading to increased excretion of the porphyrin precursors porphobilinogen and δ-aminolevulinic acid in urine. Direct gene testing is available.

Hereditary Coproporphyria

In hereditary coproporphyria, a related condition also inherited as a dominant trait, there is partial deficiency of the enzyme coproporphyrinogen oxidase. The disorder is clinically indistinguishable from acute intermittent porphyria, although approximately one-third of affected persons also have photosensitivity of the skin.

Porphyria Variegata

People with this form of porphyria, particularly prevalent in South African Afrikaans (p. 109), have variable skin photosensitivity with neurological and visceral features that can also be triggered by drugs. Increased fecal excretion of the porphyrin precursors protoporphyrin and co-proporphyrin can be demonstrated and the disorder has been shown to be due to deficiency of the enzyme protoporphyrinogen oxidase.

Erythropoietic Porphyrias

The erythropoietic porphyrias include congenital erythropoietic porphyria and erythropoietic protoporphyria.

Congenital Erythropoietic Porphyria

The main feature of congenital erythropoietic porphyria is an extreme photosensitivity with blistering of the skin leading to extensive scarring, to the extent that most affected people are unable to go out in normal daylight. In addition, many have a hemolytic anemia requiring regular blood transfusion and frequently splenectomy. Affected individuals have red–brown discoloration of the teeth, which show red fluorescence under ultraviolet light. Congenital erythropoietic porphyria is due to deficiency of the enzyme uroporphyrinogen III synthase.

Erythropoietic Protoporphyria

Erythropoietic protoporphyria is from a deficiency of the enzyme ferrochelatase, which is responsible for the insertion of ferrous iron into the porphyrin precursor to form heme. Affected persons have photosensitivity and sometimes develop chronic liver disease. Successful treatment of the photosensitivity has been reported with β-carotene.

Organic-Acid Disorders

Children with one of the organic-acid disorders present with periodic episodes of poor feeding, vomiting, and lethargy in association with a severe metabolic acidosis, low white cell (neutropenia) and platelet (thrombocytopenia) counts, low blood sugar (hypoglycemia), and high blood ammonia levels (hyperammonemia). These episodes are often precipitated by intercurrent illness or increased protein intake, and after such an episode affected children can lose developmental skills. Analysis of blood from children at the time of these episodes reveals high levels of glycine (hyperglycinemia). It was subsequently found that the acidosis in these episodes was due to increased levels of the organic acids, either propionic or methylmalonic acid.

The two autosomal recessive organic-acid disorders methylmalonic acidemia and propionic acidemia are caused by deficiency of the enzymes methylmalonyl-CoA mutase and propionyl-CoA carboxylase, respectively. The enzyme deficiency results in accumulation of the toxic organic-acid metabolites derived from deamination of certain amino acids, specific long-chain fatty acids, and cholesterol side chains. Therapy for the acute episode involves the treatment of any infection, fluid replacement, correction of the metabolic acidosis, and cessation of protein intake. Long-term prophylactic treatment involves restriction of protein intake and rapid recognition and management of any intercurrent illness. A proportion of individuals affected with propionic acidemia are responsive to biotin, whereas some with methylmalonic acidemia are sensitive to vitamin B₁₂.

Disorders of Copper Metabolism

There are two inborn errors of copper metabolism: Menkes disease and Wilson disease.

Menkes Disease

Menkes disease is an X-linked recessive disorder in which affected males present in the first few months of life with feeding difficulties, vomiting, and poor weight gain. Subsequently, hypotonia, seizures, and progressive neurological deterioration ensue, with death from recurrent respiratory infection usually occurring by the age of 3 years. A characteristic feature is the hair, which lacks pigment, is kinky, and is brittle. This was noted to resemble the wool of sheep suffering from copper deficiency. Serum copper and ceruloplasmin levels are very low. Cloning of the gene for Menkes disease was facilitated through an affected female with an X-autosome translocation (p. 116) and revealed it to code for an ATPase cation transport protein for copper. Treatment regimens with different exogenous copper sources have had limited benefit to date.

Wilson Disease

Autosomal recessive Wilson disease commonly presents in childhood or the early adolescence with fits and abnormal neurological findings, including deteriorating coordination, involuntary movements, abnormal tone, dysarthria, dysphagia, and changes in behavior or frank psychiatric disturbance. Clinical examination may reveal ‘Kayser-Fleischer rings’, which are golden brown or greenish collarettes at the corneal margins. Investigation can reveal the presence of abnormal liver function, progressing to cirrhosis.

High copper levels in the liver, decreased serum concentrations of the copper transport protein ceruloplasmin, and abnormal copper loading test results are suggestive of the diagnosis. The gene for Wilson disease was cloned on the basis of anticipated homology to the Menkes gene, and the gene product has been shown to be an ATPase cation transport protein involved in copper transfer from the hepatocytes to the biliary collecting system.

There are dramatic reports of striking improvement of the neurological features using the chelating agents d-penicillamine and trientine.

Peroxisomal Disorders

The peroxisomes are subcellular organelles bound by a single trilayer lipid membrane present in all cells; they are especially abundant in liver and renal parenchymal cells. The organelle matrix contains more than 40 enzymes that carry out a number of reactions involved in fatty-acid oxidation and cholesterol biosynthesis interacting with metabolic pathways outside the peroxisomes. The enzymes of the peroxisomal matrix are synthesized on the polyribosomes, enter the cytosol and are transferred into the peroxisomes.

There are two main categories of peroxisomal disorder: disorders of peroxisome biogenesis, such as Zellweger syndrome, in which there are severely reduced numbers of peroxisomes in all cells, and single isolated peroxisomal enzyme deficiencies, such as X-linked adrenoleukodystrophy.

Zellweger Syndrome

Newborn infants with Zellweger syndrome present with hypotonia and weakness and have mildly dysmorphic facial features (Figure 11.10), consisting of a prominent forehead and a large anterior fontanelle (‘soft spot’). They may also have cataracts and an enlarged liver. They generally go on to have fits with developmental regression and usually die by 1 year of age. Investigations can reveal renal cysts and abnormal calcification in the cartilaginous growing ends of the long bones (Figure 11.11). There is a range of severity of this disorder, with different clinical diagnoses being given to the less severe types. The diagnosis can be confirmed by raised levels of plasma long-chain fatty acids. It is genetically heterogeneous, due to any one of several genes crucial to peroxisome biogenesis.

FIGURE 11.10 Facies of an infant with Zellweger syndrome showing a prominent forehead.

FIGURE 11.11 Radiograph of the knee of a newborn infant with Zellweger syndrome showing abnormal punctate calcification of the distal femoral epiphyses.

It is unusual for IEMs to give rise to a dysmorphic syndrome (p. 249), but another is Smith-Lemli-Opitz syndrome, an inborn error of cholesterol biosynthesis from a mutation in the sterol delta-7-reductase (DHCR7) gene.

Adrenoleukodystrophy

Males with the X-linked disorder adrenoleukodystrophy (ALD) classically present in late childhood with deteriorating school performance, though presentation may occur at any age; occasionally, there are no symptoms. Some males present in adult life with less severe neurological features and adrenal insufficiency, so-called adrenomyeloneuropathy. ALD has been shown to be associated with a deficiency of the enzyme very long-chain fatty acid CoA synthase, but is secondary to deficiency of a peroxisomal membrane protein, due to mutation in the ABCD1 gene.

Treatment of ALD with a diet that uses an oil with low levels of very long-chain fatty acids—‘Lorenzo’s’ oil—has proved disappointing.

Disorders Affecting Mitochondrial Function

Mitochondrial disease was first identified in 1962 in a patient whose mitochondria showed structural abnormalities and loss of coupling between oxidation and phosphorylation, although it was not until 20 years later that the relevance of mutated mitochondrial DNA (mtDNA) to human disease began to be appreciated. The small circular double-stranded mtDNA (see Figure 2.7, p. 18) contains genes coding for ribosomal RNA (rRNA) production and various transfer RNAs (tRNA) required for mitochondrial protein biosynthesis, as well as some of the proteins involved in electron transport. There are 5523 codons and a total of 37 gene products. Guanine and cytosine nucleotides are asymmetrically distributed between the two mtDNA strands—the guanine-rich strand being called the heavy (H) strand and the cytosine-rich the light (L) strand. Replication and transcription is controlled by a 1122-bp sequence of mtDNA known as the displacement loop (D-loop). Oxidative phosphorylation (OXPHOS) is the biochemical process responsible for generating much of the ATP required for cellular energy. The process is mediated by five intramitochondrial enzyme complexes, referred to as complexes I–V, and the mtDNA encodes 13 OXPHOS subunits, 22 tRNAs, and 2 rRNAs.

The ‘complexes’ are aptly named. Analysis of complex I, for example, has revealed approximately 41 different subunits, of which 7 are polypeptides encoded by mtDNA genes known as ND1, ND2, ND3, NDL4, ND4, ND4L, ND5, and ND6, with the remaining 34 subunits encoded by nuclear DNA genes. Complex V comprises 12 or 13 subunits, of which two, ATPase 6 and 8, are encoded by mtDNA. Maximal activity of complex V appears to require tight linking with cardiolipin (see Barth syndrome, p. 182), encoded by nuclear DNA.

Because most mitochondrial proteins, including subunits involved in electron transport, are encoded by nuclear genes, these most often follow autosomal recessive inheritance. As with other metabolic autosomal recessive diseases, disorders resulting from mutations in these genes tend to breed true. However, the disorders resulting from mutations in mtDNA are extremely variable owing to the phenomenon of heteroplasmy (see Figure 7.30, p. 126). The clinical features are mainly a combination of neurological signs—encephalopathy, dementia, ataxia, dystonia, neuropathy, and seizures—and myopathic signs—hypotonia, weakness, and cardiomyopathy with conduction defects. Other symptoms and signs may include deafness, diabetes mellitus, retinal pigmentation, and acidosis may occur. The clinical manifestations are so variable that a mitochondrial cytopathy should be considered as a possibility at any age when the presenting illness has a neurological or myopathic component. Several distinct clinical entities have been determined and, although some of them overlap considerably, there is a degree of genotype–phenotype correlation.