Inborn Errors of Metabolism

Principles of Screening Programs

A few principles apply to all screening programs. First, all screening tests are subject to false-positive results because of normal biologic variation, genetic heterogeneity, and human error. Accordingly, all positive screening results must be confirmed by definitive analysis. It is important that all patients who require therapy receive it and, conversely, that patients who do not require therapy not be treated.

Second, all positive results must be considered medical emergencies. Many positive results turn out to be falsely positive, but the concept underlying newborn screening is that identification of the few affected patients is crucial. In addition to the potential tragedy of a missed diagnosis of the individual neonate, lack of attention that permits delayed care of a single affected patient can seriously jeopardize the public's confidence in and the cost-benefit structure of an entire statewide screening program and can compromise the continuation of such programs.

Third, the disorders that are part of newborn screening may exhibit variable clinical expression even within families. Thus, the siblings of a patient identified by a screening program should be biochemically evaluated for the same disorder, because they could be affected, although they appear free of symptoms.

Fourth, all patients should be referred to an experienced specialist for definitive diagnosis, because these disorders are characterized by clinical and genetic heterogeneity, which can significantly affect care of the patient and genetic counseling for the family.

There is considerable variation in the screening programs of different states in the United States and in various nations. All states and US territories screen for PKU. Until relatively recently, most states performed newborn screening for three to six metabolic disorders (including PKU, homocystinuria, MSUD, and galactosemia), one endocrine disorder (congenital hypothyroidism), and the hemoglobinopathies. The requirements and procedures for the screening programs for congenital hypothyroidism and the hemoglobinopathies are discussed in Chapters 88 and 79 , respectively.

Screening Techniques

Most state screening programs originally focused primarily on the classic inborn errors of metabolism. The testing programs employed separate tests for each disease of interest, which limited the scope of screening. These assays could not be easily adapted to screen for other groups of disorders, such as disorders of organic acid metabolism and fatty acid oxidation. The other limitation in expanding newborn screening was that the standard methods being used to diagnose the organic acidemias and fatty acid oxidation disorders—gas chromatography, or combined gas chromatography and mass spectrometry (GC/MS)—could not be upgraded to large-scale newborn screening programs, because they require tedious sample preparation and long analysis times.

Since the 1990s, intensive efforts have been made to expand the scope of newborn screening using tandem mass spectrometry (MS/MS), which circumvents the limitations of the bacterial inhibition assay, gas chromatography, and GC/MS methods. In brief, MS/MS permits analysis of a broad range of metabolites in hundreds of blood samples per day. Most states have adopted the MS/MS approach to newborn screening as part of their program. The process of expanding the scope of newborn screening programs is still continuing.

Newborn screening by MS/MS starts, as did the traditional screening programs, by collecting by heel stick a small blood sample and applying it to a standardized paper card. The period for appropriate postpartum collection is 24-72 hours in the state of Ohio and is similar in other states. Samples collected from either premature infants or sick newborns are potentially more difficult to interpret and are subject to greater false-positive and false-negative rates. The blood samples are shipped to a centralized laboratory where a standardized amount of the specimen card is punched out, following which the metabolites of interest are extracted from the punch, subjected to specific chemical modifications to make them compatible for subsequent MS/MS analysis, and automatically introduced into and analyzed by the MS/MS system.

As opposed to traditional screening protocols that required different analytic approaches for each disorder, the current MS/MS techniques permit analysis of a large number of metabolites belonging to a particular category of disease—hence, many disorders—in each sample. Hundreds of samples can be prepared for analysis, analyzed, and interpreted each day. The analysis is performed by state-of-the-art mass spectrometers that permit highly sensitive, accurate, and concurrent identification of multiple metabolites. Computer software permits pattern recognition using several related metabolites, thereby improving the reliability of the testing. In summary, the MS/MS technology is ideally suited for newborn screening of many samples for many possible disorders.

As with traditional newborn screening programs, the current MS/MS screening programs must determine the normal range for the different metabolites they analyze in their system. More importantly, the programs must set cutoffs above or below, which they identify a case as at-risk. This is a difficult, ongoing task. Programs that set their cutoffs too high have an unacceptable false-negative rate, and programs that set their cutoffs too low have an unacceptable false-positive rate.

Experience has now demonstrated that MS/MS programs can detect PKU, MSUD, and homocystinuria as well as or better than the traditional screening approaches. Nevertheless, the practitioner must still be aware that the MS/MS-based screening programs have similar problems with false-positive and false-negative results as their older counterparts, although they appear to have lower false-positive rates. The practitioner must still determine whether a particular result is truly positive or falsely positive as expeditiously as possible. The expanded newborn screening programs have found that approximately 1 in 4000 newborns have an identifiable inborn error of metabolism.

Screening for Disorders

Most states in the United States have adopted MS/MS screening to analyze disorders of amino acid metabolism (including several urea cycle disorders), organic acid metabolism, and fatty acid oxidation while still screening for several other disorders using test-specific methods (e.g., biotinidase deficiency or galactosemia). The amino acid disorders and urea cycle disorders are detected by analyzing for increased blood concentrations of specific amino acids or combinations of amino acids. Most programs do not screen for disorders that are associated with reduced concentrations of specific amino acids, such as using a low serine concentration to screen for serine biosynthesis disorders. Similarly, the organic acidemias and fatty acid oxidation disorders are detected by analyzing for increased blood concentrations of specific acylcarnitines, which are the esters formed between carnitine and the acid(s) that characteristically accumulate in the various organic acidemias and fatty acid oxidation disorders. Many programs evaluate samples for combinations of particular acylcarnitines to increase the reliability of their results. Screening for the plasma membrane carnitine uptake defect (also known as carnitine uptake deficiency) is an exception to the rule of looking for increased concentrations of characteristic metabolites, because it is based on identification of a reduced (rather than increased) concentration of free carnitine.

More recently, significant progress has been achieved in developing and introducing newborn screening for two additional categories of inborn errors of metabolism: lysosomal storage disorders and peroxisomal disorders. Two lysosomal disorders, Hurler syndrome (aka mucopolysaccharidosis type I) and Pompe disease (aka glycogen storage disease type II), and one peroxisomal disorder, X-linked adrenoleukodystrophy (X-ALD), have been added to the federally recommended newborn screening panel. Screening for each of these disorders is currently available or approved for implementation in several states. The infantile form of Hurler disease typically manifests in the first year of life, whereas Pompe disease typically manifests in the first 6 months of life. Effective treatment is now available for Hurler syndrome (hematopoietic stem cell transplantation, HSCT) and Pompe disease (enzyme replacement therapy) if it is started prior to the onset of irreversible clinical manifestations. Similarly, hematopoietic stem cell transplantation significantly improves the prognosis for X-ALD, a peroxisomal disorder that does not manifest clinically until 2 years or older. Here too, follow-up studies have shown that the outcome for patients who have received HSCT are best when it is performed prior to onset of subclinical evidence of neurologic damage. The screening protocol for X-ALD also identifies patients who have Zellweger syndrome, a generalized defect in peroxisomal assembly and function, or one of several single-gene defects in peroxisomal very long-chain fatty acid oxidation. Unfortunately, there is no effective therapy for these peroxisomal disorders.

It should be noted that there is ongoing concern regarding screening for these lysosomal and peroxisomal disorders, since it is possible to detect later-onset forms of these diseases with the screening tests, for which there are limitations in our ability to predict if, or when, the patient will develop significant clinical manifestations. For example, in the case of X-linked adrenoleukodystrophy, some patients identified by newborn screening and then found to have an ABCD1 mutation might not develop clinically significant disease but might nevertheless undergo HSCT transplantation because of the requirement to initiate treatment for the childhood form of the disease as early as possible. This concern may also apply to other disorders detected by newborn screening, but the concern for the lysosomal storage disorders and peroxisomal disorders is heightened by the fact that the recommended treatment for these disorders is more invasive (and expensive) than that required for most other disorders detected by newborn screening. It is anticipated that newborn screening for the lysosomal storage disorders and peroxisomal disorders will become available in more states in the near future. Similarly, it is expected that additional inborn errors of metabolism will be incorporated into newborn screening programs as screening technology improves, more therapeutic options are developed and assessed, and better genotype/phenotype correlations are established.

Table 90.1 lists abnormal laboratory findings, along with the disorders associated with those findings and the additional testing recommended to evaluate the significance of the findings, for the currently recommended newborn screening panel. In many cases, a particular abnormal laboratory finding can be associated with more than one disorder, because different enzymatic defects can lead to excessive accumulation of that metabolite. For example, acylcarnitine analysis can yield ambiguity in identifying several of the acylcarnitines evaluated in the newborn screening program, such as C4-acylcarnitine. C4-acylcarnitine is composed of an acid group with four carbons and carnitine; the acid group can be either butyrylcarnitine (wherein the four carbons are arranged in a linear pattern) or isobutyrylcarnitine (wherein the four carbons are arranged in a branched pattern). The diseases associated with these two acylcarnitines are quite different, and further studies are required to determine which metabolite, or which disorder, is present.

The confirmatory studies listed in Table 90.1 are readily available or orderable in most clinical settings and are discussed in detail later (see Specialized Biochemical Testing ). The confirmatory studies cited include tests that have a relatively rapid turnaround time, generally 1-2 weeks, hopefully leading to rapid confirmation or elimination of a possible diagnosis. Additional, more refined studies, including specific enzyme analysis, whole cell studies, or genetic mutational analysis, are often required to definitively establish a specific diagnosis, but these generally have a longer turnaround time.

The abnormal laboratory findings listed in Table 90.1 permit the diagnosis of more than 30 genetic disorders, including amino acid disorders, fatty acid oxidation disorders, organic acidemias, urea cycle disorders, and several unrelated enzymatic defects. The list of metabolites provided in Table 90.1 is not comprehensive. Many other metabolites have been identified or can theoretically be identified, but they are not listed because of the rarity or uncertain clinical phenotype of the associated disorder. Not all states test for this particular list of metabolites; some test for fewer and others for more. Practitioners should be familiar with the scope of their local newborn screening program. In any event, the laboratory findings listed in Table 90.1 should provide all practitioners with a foundation for interacting with their local program.

Table 90.2 provides basic information about the disorders cited in Table 90.1 , including the name of each disorder along with its common abbreviation (if one is available), the underlying enzymatic defect, the clinical features and natural history, the general approach to treatment, and the prognosis. The frequency of these disorders ranges between approximately 1 in 10,000 for PKU to 1 in 200,000 for MSUD. However, several disorders have been reported in only a small number of patients. In addition to their rarity, most of these disorders are characterized by a high degree of clinical variability, making it difficult to provide a succinct but accurate summary. Hopefully, the information will provide the practitioner with a reasonable place to start when confronted with a patient who has an abnormal newborn screening result, following which he or she can turn to other resources after a diagnosis is established.

Handling Test Results

The first obligation of the practitioner who receives an abnormal newborn screening report is to inform the parents of the result. The practitioner should explain that the results are provisional and that confirmation is required. The physician must aim for an appropriate balance between his or her own natural desire to reassure the parents that the result might be falsely positive and the desire to instill a sense of appropriate concern in the parents so that they can carry through with appropriate follow-up evaluation. The practitioner's burden is generally more straightforward when the abnormal metabolite is associated with only a single disorder, but the principles of reassurance and follow-up are the same for metabolites that can be found in more than one disorder (see Table 90.1 ). The physician should then assess the newborn's clinical status and arrange to see the family as expeditiously as possible.

The primary physician should either see the patient or refer the patient to a metabolic disorders specialist for further evaluation and care. It is often best that the primary physician see the patient as soon as possible to assess the patient's status, discuss the newborn screening results with the family, and then work with the metabolic disorders specialist to develop an expeditious plan for evaluation. Confirmatory testing should be initiated as soon as possible.

The decisions about when to initiate treatment and how to treat are based on the nature of the laboratory abnormality found, the quantitative degree of the abnormality, the program's prior experience with false-positives for that metabolite, and the patient's clinical status. In general, starting a treatment immediately after the initial confirmatory studies are initiated is both safe and unlikely to compromise the ability to establish a diagnosis. However, this option is predicated on the ability of the physician to make certain that the diagnostic samples are collected properly, sent to the appropriate laboratory, and received in satisfactory condition by the laboratory. Failure to do this before starting treatment might significantly delay the time required to establish a diagnosis and initiate appropriate treatment.

Many of the disorders identified are treated, at least in part, by some form of dietary restriction. A family's desire to continue breastfeeding while the diagnostic studies are in progress should be carefully considered in all cases. However, depending on the disorder under consideration and the patient's clinical status, the default position should be in favor of pausing breastfeeding until a provisional diagnosis has been established. During that time, the mother can continue to express breastmilk via a breast pump and store it for later use. It is generally a matter of days to a week before the results of the initial confirmatory studies are available, when a more definitive decision can be made about the advisability of breastfeeding. Similar reasoning should be exercised about starting vitamin or cofactor supplementation.

Biotinidase Deficiency

Biotinidase is an enzyme necessary for recycling biotin, a vitamin cofactor required for four critical intracellular carboxylation reactions: acetyl-coenzyme A (acetyl-CoA) carboxylase, 3-methylcrotonyl-CoA carboxylase, propionyl-CoA carboxylase, and pyruvate carboxylase. Hence biotinidase deficiency is one cause of multiple carboxylase deficiency (the other cause is holocarboxylase synthetase deficiency). These carboxylase reactions are involved in fatty acid biosynthesis, branched-chain amino acid metabolism, and gluconeogenesis.

Biotinidase deficiency is characterized by a variable clinical presentation but can lead to severe metabolic decompensation in the newborn period; features include ketoacidosis, hypotonia, seizures, and coma. Some infants also have significant dermatologic findings (including rash and alopecia) and immunodeficiency. If untreated, older children could have visual problems, hearing loss, and developmental delay. This disorder can be treated successfully with biotin supplementation (5-10 mg/day PO). Some residual neurologic deficits could persist if treatment does not begin before the onset of symptoms. Newborn screening allows for early diagnosis and initiation of treatment, which may reduce morbidity.

Serum biotinidase enzyme activity is the gold standard for newborn screening of biotinidase deficiency. The disorder can also be detected using MS/MS to measure the blood concentration of C5-OH (3-hydroxyisovalerylcarnitine), the acylcarnitine that is formed due to the secondary deficiency of 3-methylcrotonyl-CoA carboxylase. However, the sensitivity of the MS/MS approach is unknown, and it might not provide a reliable method for newborn screening. A positive screening result should be confirmed by quantitative serum biotinidase analysis and by performing plasma carnitine analysis and urine organic acid analysis, looking for the characteristic plasma acylcarnitine pattern and organic aciduria that is present in a small percentage of affected patients.

Care must be exercised in collecting and processing the serum specimen used for biotinidase enzyme analysis. It is best to obtain a concurrent control from an unrelated individual to establish that the sample has been processed properly (i.e., eliminate the chance of a false-positive result) and was not exposed to environmental factors during transport that could affect the results (such as excess heat or cold).

Galactosemia

Classic galactosemia is the consequence of galactose-1-phosphate uridyltransferase ( GALT ) deficiency. Classic galactosemia can manifest in the newborn period with lethargy, poor feeding, jaundice, cataracts, and in some cases, Escherichia coli sepsis. If unrecognized, this disorder can lead to early death or a chronic course characterized by cirrhosis, cataracts, seizures, and mental retardation. The mainstay of therapy for classic galactosemia is strict dietary galactose restriction. Diet therapy is difficult to sustain because lactose (a disaccharide formed from galactose and glucose) is a ubiquitous food additive. Dietary galactose restriction should be started as early as possible (preferably within the first few days after birth) to have the best chance of precluding the development of the severe illness described above as well as long-term speech and learning problems. However, even children treated early often have mild growth failure, learning disabilities, and verbal dyspraxia. Affected girls almost invariably develop premature ovarian failure (~80%). This observation serves as a caution to those caring for children with galactosemia that long-term follow-up is mandatory and further improvements in treatment are required over time as new recommendations and information becomes available.

There are two other forms of galactosemia: uridine diphosphate galactose-4′-epimerase deficiency and galactokinase deficiency. In most cases, epimerase deficiency is a benign condition that does not require treatment. The rarer, systemic form of epimerase deficiency produces a clinical picture similar to classic galactosemia. Galactokinase deficiency is also rare and produces nuclear cataracts but none of the other manifestations of classic galactosemia. Early recognition and treatment of this disorder via dietary galactose restriction is generally successful.

One approach to newborn screening for galactosemia measures GALT enzyme activity in red blood cells. This assay can detect GALT deficiency without regard to prior dietary intake of galactose. It does not evaluate for either epimerase activity or galactokinase activity. Therefore, newborns with either epimerase deficiency or galactokinase deficiency are not ascertained by this approach to newborn screening. Another approach to newborn screening for galactosemia is to measure galactose and galactose-1-phosphate (the substrate for GALT ), which evaluates for all three enzyme deficiencies. However, recent galactose intake (in the form of breastmilk or formula) may affect these results; patients not receiving a lactose-containing diet may have false-negative results. Because of the rapid onset of symptoms in classic galactosemia and the presence of galactose in breast milk and many cow milk–based formulas, screening programs for galactosemia must provide rapid results. However, the screening results are not always available before the affected neonate becomes ill; initial evaluation of a sick newborn should, therefore, include testing for the presence of urinary-reducing substances (see Specialized Biochemical Testing ). A newborn identified by newborn screen as possibly having classic galactosemia should have definitive biochemical testing by measuring whole blood or erythrocyte GALT activity and erythrocyte galactose-1-phosphate. In addition, genetic analysis for the common GALT mutations is often helpful in interpreting the results of the GALT activity measurements and making treatment decisions. Following initiation of these studies, galactose should be withdrawn from the diet, pending results of the laboratory investigations, by replacing breastmilk or cow's milk–based formula with a soy-based formula).

Widespread neonatal testing of erythrocyte GALT activity in various populations has revealed considerable genetic heterogeneity of this enzyme deficiency. Some individuals have a partial enzyme deficiency that does not result in significant impairment of galactose metabolism or any discernible clinical disorder; there is no evidence of a need for dietary treatment of these cases. In other cases with partial enzyme activity, erythrocyte galactose-1-phosphate concentrations are increased, and minimal symptoms can develop. These cases can be managed with less severe restriction of dietary galactose intake.

Homocystinuria

Several inborn errors of metabolism produce homocystinuria. The most common of these disorders is caused by cystathionine β-synthase deficiency, an autosomal recessive disorder. Cystathionine β-synthase is a pyridoxine (vitamin B ₆ )-dependent enzyme. Rare disorders that also lead to homocystinuria include defects in folate or cobalamin metabolism. Screening programs for homocystinuria are based on detection of elevated blood levels of methionine, a precursor of cystathionine. Hypermethioninemia is characteristic of cystathionine β-synthase deficiency but may not be associated with other causes of homocystinuria. In fact, patients who have homocystinuria due to a defect in cobalamin metabolism may actually have low methionine levels. Therefore, only some cobalamin disorders are detected by newborn screening for hypermethioninemia, while others are detected by their abnormal acylcarnitine profile. For example, cobalamin C disease will exhibit both methylmalonic acidemia and homocystinuria. These patients are ascertained by an elevated propionylcarnitine (C3), which is related to the methylmalonic acidemia associated with their disease. In the case of a related disorder, patients with cobalamin E disease exhibit homocystinuria/homocystinemia, megaloblastic anemia, and low methionine but are missed by newborn screening.

Other false-positive and false-negative results occur in screening programs for homocystinuria. False-positive results are generally the consequence of artifacts (e.g., poor quality of sample), but they may also result from nongenetic causes of hypermethioninemia, such as parenteral administration of amino acids, generalized liver disease, hepatic immaturity, or rarely, from a genetic cause such as hereditary tyrosinemia, galactosemia, or citrin deficiency (see Hepatic Dysfunction ). False-negative results are produced by the milder variants of cystathionine β-synthase deficiency, especially the pyridoxine-responsive form, which might not exhibit hypermethioninemia in the neonatal period.

The diagnosis of homocystinuria should be confirmed in patients with a positive newborn screening test by measuring total plasma homocysteine, plasma amino acids, and plasma methylmalonic acid. The diagnosis of cystathionine β-synthase deficiency can be confirmed by measuring the enzyme activity in cultured skin fibroblasts or by genetic testing.

Cystathionine β-synthase deficiency rarely manifests in the neonatal period, but it can cause lethargy, poor feeding, and thromboembolic phenomena when it does. If untreated, the disorder can lead to musculoskeletal anomalies suggestive of a marfanoid habitus, ectopia lentis, thromboembolic vascular disease, behavioral or psychiatric problems, and mental retardation. Patients with pyridoxine-responsive defects tend to have milder disease than do patients with pyridoxine-nonresponsive defects. All patients should undergo a pyridoxine challenge test to determine whether they are pyridoxine responsive.

Patients with pyridoxine-responsive forms of homocystinuria might require only daily vitamin B ₆ supplementation and mild methionine restriction, whereas nonresponders may require a stricter, low-methionine diet with cysteine supplementation (cysteine is the product of the cystathionine β-synthase reaction, and may, therefore, be deficient in patients with classic homocystinuria). Patients might also benefit from folate, vitamin B ₁₂ , and betaine supplementation, which augment the remethylation of homocysteine to methionine, thereby reducing the toxic effects of excessive homocysteine. The outcome for vitamin B ₆ –responsive patients appears to be good, but the outcome for nonresponders has often been less satisfactory because of the irreversible damage caused by the presenting episode, the therapeutic inadequacy of the present dietary management, or both.

Medium-Chain Acyl-CoA Dehydrogenase Deficiency

Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is the most common inherited disorder of fatty acid oxidation, with an incidence of approximately 1 in 15,000. It has a highly variable clinical presentation, even within families. Patients are rarely symptomatic in the newborn period. Most patients present between 3 and 24 months of age, but others remain asymptomatic until they are much older, even into adulthood. The initial retrospective studies on MCAD deficiency found that in almost one-fourth of patients, the diagnosis followed sudden, unexplained death.

The typical presentation is that of an infant who has unexplained progressive vomiting, hepatomegaly, lethargy leading to coma, and seizures associated with an infectious illness or a period of prolonged fasting. The characteristic laboratory finding is nonketotic or hypoketotic hypoglycemia; signs of liver dysfunction may also be present. Older patients could have a more indolent course that is associated with failure to thrive, developmental delay, or chronic muscle weakness. A significant proportion of patients are asymptomatic. MCAD deficiency may be diagnosed in an “unaffected” older sibling after his or her younger sibling comes to medical attention for an acute metabolic crisis or a positive newborn screening result for MCAD deficiency.

The diagnosis of MCAD deficiency is confirmed by plasma carnitine analysis with an acylcarnitine profile, urine organic acid analysis, and urine acylglycine analysis. The plasma total and free carnitine concentration can vary with the phase of the illness. The plasma acylcarnitine profile typically shows increased amounts of C6, C8, and C10 acylcarnitines. However, patients with secondary carnitine deficiency could have a relative increase, but not an absolute increase, in these acylcarnitines. Urine organic acid analysis generally shows a typical medium-chain dicarboxylic aciduria (C6-C10) in symptomatic patients, but it could be normal in asymptomatic patients. Urinary acylglycine analysis can be useful in detecting the characteristic metabolite (suberylglycine) in asymptomatic patients.

Genetic testing is useful for confirming the diagnosis. It used to be thought that at least 90% of patients carried at least one copy of the same abnormal allele, the A985G mutation, but the frequency of this particular allele is lower in patients identified by newborn screening programs compared with those who present symptomatically.

Patients with MCAD deficiency should receive frequent feedings and avoid fasting. Infants younger than 6 months require feeding every 3-4 hours. The older infant or child may be allowed to fast for progressively longer periods of time as they get older but not for more than 12 hours. The potential metabolic stress of nocturnal fasting might be reduced for toddlers by providing them with uncooked cornstarch as a source of slow-release glucose immediately before going to bed for the night. Dietary fat restriction is no longer recommended, but oral carnitine supplementation may be indicated for patients with secondary carnitine deficiency. Carnitine supplementation should be monitored with periodic plasma carnitine analysis. Patients should be evaluated and admission to the hospital considered for patients with anorexia or vomiting associated with an acute infectious illness or other potential metabolic stressor. Acute metabolic crises should be treated with intravenous glucose and carnitine. The prognosis for patients who are identified and treated before the onset of irreversible neurologic damage due to severe or recurrent hypoglycemia and its sequelae is generally excellent.

Phenylketonuria

Several genetic disorders produce hyperphenylalaninemia. Classic PKU and non-PKU hyperphenylalaninemia are caused by defects in phenylalanine hydroxylase, and variant PKU is caused by one of several defects in tetrahydrobiopterin (BH ₄ ) metabolism. Several nongenetic factors also produce hyperphenylalaninemia in the newborn, but this hyperphenylalaninemia disappears in the first year of life. The most common causes of transient hyperphenylalaninemia are prematurity and high protein intake just prior to the blood draw. Newborn screening programs identify infants with genetic and nongenetic hyperphenylalaninemia. It is imperative that patients identified by the screening program receive a rapid, accurate, and definitive diagnosis, because the clinical implications and therapies for the various forms of hyperphenylalaninemia are different and early initiation of appropriate treatment minimizes morbidity.

All genetic forms of hyperphenylalaninemia are caused by defects that directly or indirectly affect the activity of the enzyme phenylalanine hydroxylase. This enzyme catalyzes the conversion of phenylalanine to tyrosine and requires BH ₄ as a cofactor. Classic PKU and non-PKU hyperphenylalaninemia are caused by allelic defects of phenylalanine hydroxylase itself, whereas variant PKU is caused by defects of BH ₄ biosynthesis or reutilization. BH ₄ is also a cofactor for tyrosine hydroxylase and tryptophan hydroxylase, which are enzymes involved in neurotransmitter biosynthesis. Approximately 98% of patients with hyperphenyl have a defect in phenylalanine hydroxylase, whereas 2% have a defect in BH ₄ metabolism.

Patients with milder deficiencies of phenylalanine hydroxylase (i.e., patients with transient hyperphenylalaninemia or non-PKU hyperphenylalaninemia) usually do not require dietary treatment. However, patients with more severe deficiencies, that is, patients with classic PKU, require lifelong phenylalanine restriction. Treatment should start within the first month of life to avoid irreversible neurologic damage. Early treatment is generally effective in preventing the long-term neurologic sequelae of this disease. However, standard treatment may not prevent subtle intellectual and behavioral disabilities in some individuals. Dietary restriction of phenylalanine should be lifelong. Women with PKU who fail to maintain the appropriate diet are at risk for having neurologically impaired offspring (maternal PKU syndrome; see Maternal Diseases Affecting the Fetus ).

In recent years, clinical trials have demonstrated that some patients with milder forms of classic PKU respond to oral supplementation with a synthetic form of BH ₄ known as sapropterin. They appear to tolerate a higher dietary protein intake while maintaining acceptable serum phenylalanine concentrations. However, most patients still require some degree of dietary phenylalanine restriction with or without a specialized metabolic formula. The drug is now available for clinical use.

Defects of BH ₄ metabolism cause defective neurotransmitter synthesis as well as hyperphenylalaninemia and lead to a more generalized neurologic syndrome known as variant PKU, characterized by convulsions, abnormal tone and posture, abnormal movements (i.e., dystonia), hyperthermia, hypersalivation and swallowing difficulties, drowsiness, irritability, and developmental delay. Standard dietary management corrects the hyperphenylalaninemia that these patients have but does not improve the neurologic problems related to their neurotransmitter deficiencies. Only a small percentage of patients with hyperphenylalaninemia have BH ₄ -related defects, and these patients must be identified early to initiate appropriate therapy. A variety of approaches are being used to treat these patients with some success, depending on the specific biopterin defect present.

Newborn screening for hyperphenylalaninemia is based on measuring the concentration of phenylalanine in the blood while the newborn infant is receiving breast milk or a standard formula (i.e., a phenylalanine-containing diet). Blood samples are generally obtained for newborn screening between 24 and 48 hours of age. The results for samples collected before 24 hours of age may not be reliable, and a follow-up sample should be obtained for screening.

The rate of false-negative results is less a problem with MS/MS screening than the traditional approaches, partly because it is possible to measure the concentrations of phenylalanine and tyrosine (the product of the phenylalanine hydroxylase reaction) concurrently. Those who have a positive screening result require prompt attention, including plasma amino acid analysis. If the plasma phenylalanine concentration and the plasma phenylalanine-to-tyrosine ratio are increased, the patient should be placed on a low-phenylalanine diet. Therefore, breastfeeding should be stopped. In all cases of confirmed hyperphenylalaninemia, the patient should be evaluated by a metabolic disorders specialist to rule out a defect in BH ₄ metabolism.

Maternal Diseases Affecting the Fetus

The prototype of a maternal disease affecting the fetus is maternal PKU. A pregnant woman who has poorly controlled PKU is at increased risk for spontaneous abortion. She also has an increased risk of having a child with major birth anomalies, including intrauterine growth restriction, microcephaly, mental retardation, and a congenital heart defect, as well as a broad range of minor anomalies. The risk of birth defects is proportional to the mother's serum phenylalanine concentration. There is no safe level below which the fetus is not at risk. Women with PKU should be placed on a strict low-phenylalanine diet before conception. This has proved difficult to do in practice, however, and maternal PKU syndrome remains a significant problem for women with PKU.

Fetal Diseases Affecting the Mother

As a rule, inborn errors of metabolism of the fetus do not affect the pregnant mother. However, reports began to appear in the 1990s describing mothers who had experienced acute fatty liver of pregnancy (AFLP) while carrying a fetus who subsequently manifested evidence of a long-chain fatty acid oxidation defect after birth. Acute fatty liver of pregnancy is the most extreme end of a clinical spectrum of maternal complications of pregnancy that includes HELLP syndrome (hemolysis, elevated liver enzymes, and a low platelet count) and AFLP. The HELLP syndrome and AFLP are potentially serious complications of pregnancy (see Chapter 17 ).

Clinical, biochemical, and histologic evidence of hepatic dysfunction mark both disorders. Patients with HELLP syndrome commonly develop epigastric pain, nausea, vomiting, headache, proteinuria, low platelets, elevated serum liver enzymes, and occasionally disseminated intravascular coagulation. Acute fatty liver of pregnancy is less common than HELLP syndrome and shows a greater degree of hepatic dysfunction. It often produces a severe coagulopathy, hypoglycemia, and fulminant hepatic failure. Microvesicular fatty deposits in the liver characterize both disorders. Both disorders can have life-threatening consequences for the fetus and mother. Women with either of these disorders improve remarkably after delivery, suggesting that the fetus is causing a toxic effect on the mother that resembles that seen in patients with inborn errors of fatty acid oxidation.

There is convincing evidence that a specific defect of fatty acid oxidation, long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) deficiency in the fetus, is associated with the HELLP syndrome and AFLP spectrum in the mother. Long-chain 3-hydroxyacyl-CoA dehydrogenase is part of a trifunctional multimeric enzyme complex that performs the three terminal steps in the long-chain fatty acid β-oxidation: long-chain 2,3-enoyl-CoA hydratase, LCHAD, and long-chain 3-ketoacyl-CoA thiolase. Long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency can be seen as an isolated deficiency or as part of trifunctional protein deficiency that affects all three enzyme activities. Both disorders are inherited as autosomal recessive traits.

Isolated LCHAD deficiency is marked by relative genetic homogeneity. More than 50% of the mutant alleles found in patients who have this disease carry the same mutation: a G1528C change in the gene that encodes for the α-subunit of LCHAD. Other mutations in the same gene that predispose a heterozygous mother to AFLP have also been identified. A relatively large number of at-risk pregnancies have now been identified and reviewed after the birth of a child with LCHAD deficiency. These studies show that a woman who is heterozygous for LCHAD deficiency is at risk for developing HELLP syndrome or AFLP during a pregnancy in which she is carrying a fetus who is homozygous for the same deficiency. Thus, a heterozygous mother is only at risk if her fetus inherits a second LCHAD mutation from the father. The fetus, in turn, is at risk for significant postnatal problems associated with its enzyme deficiency. The diagnosis and care of a child with LCHAD deficiency is discussed later under Hypoglycemia. Rarely, other inborn errors of long-chain fatty acid β-oxidation can also produce AFLP.

The current recommendation is to evaluate all pregnant women who develop AFLP for LCHAD deficiency and related fatty acid oxidation disorders by biochemical and genetic testing. This recommendation should also be considered for women who develop recurrent and/or severe HELLP syndrome.

Fetal Diseases Affecting the Fetus

The fetus does not generally suffer prenatal consequences of its own inborn error of metabolism, because the abnormal metabolites that it produces are removed by the maternal circulation, or conversely, metabolic deficiencies produced by the inborn error are replenished by the maternal circulation. There are, however, several groups of disorders that include significant exceptions: amino acid disorders, congenital disorders of glycosylation, fatty acid β-oxidation disorders, lysosomal storage disorders, mitochondrial disorders, and peroxisomal disorders (see Dysmorphic Syndromes ).

Although most of the clinical disorders that affect the developing fetus are discussed later, the disorders that can lead to congenital ascites or hydrops fetalis are discussed here. A major group of inborn errors of metabolism that lead to hydrops fetalis are the lysosomal storage disorders, including β-glucuronidase deficiency and Morquio syndrome (which are mucopolysaccharidoses); Farber disease and GM1-gangliosidosis (which are spingolipidoses); galactosialidosis and sialidosis (which are oligosaccharidoses); and free sialic storage disease (a lysosomal transport defect). Other disorders that can also lead to hydrops fetalis are congenital disorders of glycosylation (specifically type Ia), glycogen storage diseases (specifically type IV), Niemann-Pick disease type C, and transaldolase deficiency, as well as several inborn errors of red cell glycolytic enzymes (see Chapter 23 ).

Metabolic Encephalopathy

The inborn errors of metabolism that can produce metabolic encephalopathy in the neonatal period or early infancy are associated with what appears to be a cascade of biochemical effects. A number of specific metabolites are overproduced as a result of a particular enzyme deficiency. In excess, these metabolites serve as endogenous toxins that impair other metabolic or physiologic processes. Disruption of these processes leads, in turn, to production of additional metabolites, which further impair cellular processes. Early interruption of the cascade through such relatively simple measures as fluid and caloric support might abort episodes of metabolic decompensation. Physicians who care for children who present with a metabolic disorder in late infancy, childhood, or even adulthood often receive a retrospective history of “sepsis” in the neonatal period that was never confirmed by culture and that resolved spontaneously; these episodes might have represented an interrupted metabolic intoxication syndrome. Early nonspecific supportive treatment may abort the pathologic cascade or delay the onset of a more fulminant course until a provisional metabolic diagnosis and specific treatment become available.

The symptoms and signs of neonates who develop metabolic encephalopathy generally do not appear on the first day of life but usually begin later in the first week. The initial symptoms are often poor feeding associated with a poor suck and irritability. Muscle tone is decreased, sometimes marked by a fluctuating pattern of decreased and increased tone. Reflexes are sometimes abnormal, and seizures can develop. The poor feeding is sometimes accompanied by vomiting. Diarrhea is uncommon. Disorders accompanied by a metabolic acidosis (e.g., the lactic acidemias or organic acidemias) can lead to a compensatory increase in respiratory rate. In the case of urea cycle defects, hyperammonemia increases the respiratory drive, leading to hyperpnea. The neonate shows lethargy, which can progress to coma and death. These symptoms often progress rapidly, sometimes within a matter of hours, but more often during the course of a few days. It is important to suspect a metabolic disorder as early as possible in its course to interrupt the progression of symptoms because many of these disorders are life threatening.

The diagnostic evaluation of a newborn suspected of having a metabolic encephalopathy should include testing for disorders of amino acid metabolism, organic acid metabolism, the mitochondrial respiratory chain, and the urea cycle (see Table 90.3 ). The differential diagnosis for these patients can often be narrowed by the presence of other clinical or routine laboratory findings, such as acidosis, hyperammonemia, hypoglycemia, ketosis, or lactic acidemia. The diagnosis will be narrowed further by performing more specialized laboratory testing, including plasma amino acid analysis, urine organic analysis, plasma carnitine analysis with acylcarnitine profile, and urine carnitine analysis with acylcarnitine profile. Effort should be made to perform a prompt and vigorous laboratory evaluation of the patient suspected of having a metabolic encephalopathy, because many of them are potentially treatable. The disorders that produce metabolic encephalopathy are discussed later in this chapter (see The Abnormal Newborn Infant: Laboratory Phenotypes ).

Metabolic Seizures

There are an ever-growing number of inborn errors of metabolism that have been recognized as a cause of seizures in the neonatal period or early infancy. These disorders produce a variety of seizure types, including partial, tonic, and myoclonic seizures, and EEG patterns, including burst-suppression (see Chapter 55 ). The seizure type and EEG pattern may evolve over time. As in the case of patients who present with a metabolic encephalopathy, many of the disorders that produce neonatal seizures are potentially treatable. The approach to laboratory testing of these patients should include the same laboratory testing noted in the preceding for the evaluation of patients with suspected metabolic encephalopathy. In addition, the patient with a suspected metabolic seizure disorder should undergo additional, specialized testing for both potentially treatable and untreatable disorders, with emphasis directed toward identification of potentially treatable disorders. As a rule, different specialized testing must be obtained for each disorder. The physician caring for these patients should strongly consider starting therapy after the samples for laboratory testing have been obtained rather than awaiting the receipt of test results.

The clinical features, biochemical basis, diagnostic testing, treatment, and prognosis for several of the disorders that produce metabolic seizures are presented in three groups: treatable disorders, potentially treatable disorders, and untreatable disorders, to allow the attending physician to prioritize the diagnostic evaluation. The listing of disorders is by no means complete.