Inborn errors of metabolism

Inborn errors of metabolism BERT New

N. York,

LA New

DU,

M.D.,

PH.D.

York

When Garrod wrote his book, in 1909, on inborn errors of metabolism,2 he was able to cite only four examples of metabolic diseases which fitted this classification; albinism, alcaptonuria, cystinuria, and pentosuria. Today, the list of inborn errors is nearly ten times as long. In approximately half of these, it has been possible to demonstrate a specific enzymatic deficiency in vitro which offers a plausible explanation for the clinical features and the metabolic disturbance observed. The present list of metabolic errors range from defects in amino acid metabolism and amino acid transport mechanisms to defects in the metabolism of sugars, glycogen, steroids, and even the metabolism of several drugs. Furthermore, the family of glycogen storage diseases has been divided into subtypes according to the particular enzymatic step in the metabolism of glycogen which is deficient. In addition, an enzyme or transport catalyst may be altered by more than one type of mutation. Two types of cystinuria,3, 4 and four types of glucose-6phosphate deficiencies have been described.5 Table I lists some of the inborn errors known today selected to illustrate the wide variety of metabolic disorders and clinical findings associated with them. Several excellent books62 Z6, 79-81 have appeared in recent years which discuss exhaustively the individual metabolic disorders and describe the biochemical and clinical features of these conditions in fine detail. Of particular value is the recent reprinting of Garrod’s 1909 edition of his Inborn Errors of Metabolism, with an excellent additional section by Professor H. Harris on the inborn

errors of metabolism are primarily disturbances in enzymatically catalyzed functions and, at first glance, might appear to be quite unrelated to congenital structural anomalies. However, Garrodl recognized over 60 years ago the close analogy in inherited structural and chemical variations in man. In 1902, he wrote, “If it be a correct inference from the available facts that the individuals of a species do not conform to an absolutely rigid standard of metabolism, but differ slightly in their chemistry as they do in structure, it is no more surprising that they should occasionally exhibit conspicuous deviations from the specific type of metabolism than that we should meet with such wide departures from the structural uniformity of the species as the presence of supernumerary digits or transposition of the viscera.” Garrod considered hereditary metabolic disorders such as alcaptonuria and cystinuria to be examples of the extreme limits of the chemical variations which appear in our nonhomogeneous population. Recent advances in biochemistry, molecular biology, and genetics give eloquent support to the concept that many inherited structural anomalies, the “molecular diseases” (such as the hemoglobin variants), and the hereditary metabolic disorders (due to specific enzymatic deficiencies) are all expressions of genetic variations which, in turn, result in modifications in the composition and function of specific protein molecules.

HEREDITARY

From New

the Department York University

of Pharmacology, School of Medicine.

1024

inborn

errors today.8 Therefore, we will not discuss the individual disorders in this chapter but deal rather with some of the general concepts and problems which these disorders present, as a group, to the physician.

Genetics Incidence. Most

of

the metabolic disorders associated with specific enzymatic deficiencies are relatively rare conditions inherited as Mendelian recessive characters. Affected individuals are thus homozygous for the abnormal gene. The incidence of these cases depends upon the incidence of the particular abnormal gene in the population and the likelihood that both parents carry the abnormal gene and both transmit it to their offspring. Obviously, the chances that both parents carry the same rare abnormal gene will be greater if they are related. Garrodl pointed out the importance of consanguineous marriages as a factor contributing to the incidence of alcaptonuria in 1902. He also appreciated that consanguinity, in itself, was not the cause of the inherited metabolic disorder but that such marriages favored the expression of the condition if the abnormal genes should be present in the family. However, the importance of consanguinity has probably been overemphasized since the contribution to the incidence of these conditions is relatively small from parents who are related comparecl to that from unrelated parents. Some metabolic disorders are not rare and these genes are found frequently enough to indicate that a significant fraction of the population must be heterozygous carriers of these disorders. Table II lists the incidence of heterozygous and homozygous individuals for several metabolic conditions due to enzymatic deficiencies. The high gene frequency for some of these conditions implies that special mechanisms must operate to account for their being present at such high levels. One explanation favored is that heterozygous individuals may have some selective advantage in survival or fitness, but the only example of this type in man which is reasonably well established is that a defici-

errors

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‘iO25

ency of glucose-6-phosphate dehydrogenase, gll 92 like the sickle celI trait,g3 may offer some protection against malaria. Almost all of the metabolic disorders due to enzymatic deficiencies are inherited as Mendelian recessive traits, and one might inquire why dominant transmission is so rarely encountered. Since dominance and recessivity refer to characters and not to genes, our question is really why heterozygous individuals, having both a normal gene and an abnormal gene, generally show no appreciable metabolic disturbances. A reasonable explanation is that most enzymes are present in considerable excess of the amounts required. For example, it can be calculated,l” that under optimal conditions human adult liver could oxidize up to 1,600 grams of homogentisic acid per day, over 200 times the daily load of this substrate arising from tyrosine and phenylalanine. A reduction even to approximateIy haIf of the normal Ievel of most enzymes does not produce a significant block in the intermediary metabolic pathway. Some allelic variants may produce an enzyme of reduced but significant activity so that in the heterozygous state the activity is still above half the normal level. In general, it appears that only when the abnormal allele is present in double dose are enzymatic activity and metabolic function altered sufficiently to be evident clinically.

Mechanisms leading to enzymatic defects. Before discussing several mechanisms by which genetic mutations can result in enzymatic deficiencies, let us briefly rev.iew the current theory of how genes produce their effects. Recent workg*-g6 indicates that the primary hereditary material is desoxyribonucleic acid (DNA) and that the precise structure of the DNA dictates, through specific mediators, the molecular structure and the rate of synthesis of proteins. Some genes, called structural genes, presumably represented in certain portions of the DNA molecule, determine the exact amino acid sequence in the proteins; other genes, comprising other portions of DNA, are. called control genes and regulate the rate of protein synthesis. Evidence from microbial ge-

1026

La Du

Table I. Enzymatic

defect,

hereditary

disorders*

metabolic

clinical

features,

and treatment

of a selected

group

of

-Reference

Condition

Enzyme

deficient

Main

clinical

features

Treatment

Acatalasia

Catalase

Oral lesions in Japanese; abnormalities reported Swiss group

Adrenogenital syndrome

C-2 1 Hydroxylase

Virilization in males; pseudohermaphmditism in females

Cortisone

Albinism

Tyrosinase

Lack

None

available

Alcaptonuria

Homogentisic oxidase

Atypical pseudocholinesterase

acid

of melanin

pigment

None

required

and

arthritis

None

available

Pseudocholinesterase

Prolonged apnea succinylcholine

from

Avoid

drug

Galactosemia

Galactose-l-phosphate uridyl transferase

Mental retardation, cirrhosis, poor condition

cataracts, general

Dietary; remove galactose

Glycogen deposition disease, type 1 (von Gierke’s)

Glucose-6-phosphatase

Enlarged liver, hypoglycemra

Glycogen deposition disease,

Muscle

Pain and weakness exercise

phosphorylase

Ochronosis

no in

no.

ketosis,

upon

Prevent hypoglycemia by diet, prevent acidosis and infections No

specific treatment available

we 5, ( McArdle’s

syndrome) Histidinemia

Histidase

Some have mental retardation; ?speech defects; some without clinical symptoms

None

reported

Homocystinuria

Cystathionine synthetase

Mental deficiency, epileptic seizures, iridodonesis, fine hair

None

reported

Hypophosphatasia

Alkaline

Generalized bone disease; rickets in childhood, fractures, ossification defects

No

Branched chain-keto acid decarboxylase

Mental retardation, rapid deterioration, feeding problems

Dietary; low in branched chain amino acids

Methemoglobinemia

Erythrocyte diaphorase

Methemoglobin

Ascorbic acid, methylene blue; avoid methemoglobin producing drugs

Phenylketonuria

Phenylalanine hydroxylase

Mental retardation, EEG normalities, muscular hypertonicity, agitated behavior

Diet low in phenylalanine

Primaquine sensitivity

Glucose-6-phosphate

Hemolysis from various drugs; primaquine, acetanilid, antipyrine, probenecid, and others

Avoid offending drugs

Xanthinuria

Xanthine

Renal

Prevent stones by fluid intake and alkali

Maple syrup disease

*See

Harris6

urine

for a more

comdete

phosphatase

oxidase

fist.

stones,

formation

renal

dysfunction

specific ment

treat-

Volume Number

90 7, part

Inborn

2

Table II. Frequency enzymatic

of affected conditions

deficiency

Condition

Atypical pseudocholinesterase Isoniazid-slow inactivators*

-Inability *Deficiency tAssuming

area

Northern United

(generalized)

to synthesize of liver enzymatic

vitamin

C

and carrier

heterozygotes

and

reference

Ireland82 State@ General population66 Ireland and West Scotlands’ Southeast England** Northern Ireland83 San Blas Indians of Panamaso Canada07 Eskimo Latin America United States and Canada General populations8

acetylase. 89 Data from defect in L-gulonolactone

several sources, -+ L-ascorbic

netic experiments enabled Jacob and Monod,97 to propose an elaborate hypothesis on how regulation of protein synthesis may be controlled, A “repressor,” which combines with the “operator” gene (located adjacent to the structural gene) controls the rate of protein synthesis through its suppressive action. Whether or not the model proposed by Jacob and Monod is applicable to buman genetics, it does suggest many possibilities which must be considered in investigations on mechanisms responsible for hereditary metabolic disorders in man. The simplest genetic mechanism to consider which may result in an enzymatic deficiency would be a structural gene mutation. This could result in the substitution of a single amino acid in the protein polypeptide chain, or more extensive structural modification of the enzyme protein. Possibly, no protein would be formed. So far, it has not been possible to identify single amino acid replacements in variant enzyme proteins, analogous to those in some of the hemoglobin molecules 98 but it seems reasonable to expect that iome example of this type will be found. One of the most likely prospects is the atypical pseudocholinesterase in plasma.26 Individuals homozygous for this variant have prolonged apnea following the administration of succinylcholine. This is due to a de-

1027

in several

-

Geograghic

Alcaptonuria Pentosuria Phenylketonuria

Albinism

homozygotes

errors of metabolism

---... Frequency of affected homozygotes (per 1,000) 0.004

to 0.01 0.02 0.04 0.25 0.015 0.131 4.70 0.354 46 328 551 1,ooot

compiled by K&w.” acidDO convwsion in inheritrd

as rcressiw

~.

Frequency of heterozygote carriers (per 1,000) 4.0 to 6.4 8.9 12.8 31.2 7.8 22.; 1’28 S7.fj 33 8 49” 3Si .. trait

creased rate of hydrolysis of the ester. Kalow and co-workers made a careful study of the properties of the atypical esterase and found that it has a much lower affinity for various choline ester substratesQg and a number of inhibitors, such as dibucaine and decamethonium.lOO Kalow?@ suggests that the amount of atypical esterase may not be reduced in plasma but that it is a less efficient catalyst (lower turnover number). Liddell and associates’o1 were able to separate the atypical and normal esterase by column chromatography because of a difference in charge. The atypical enzyme has not been highly purified nor subjected to “fingerprint” analysis, but it seems likely that this will show an alteration of the amino acid sequence in the anionic site of the enzyme. Another enzymatic protein which may represent a structural alteration is the glucose-6-phosphate dehydrogenase variant observed by Kirkman, Riley, and Crowell,l” in a patient with congenital nonspherocytic anemia, The variant enzyme differed from the normal in its physical properties and had different affinity constants for both substrate atid TPN. Mutations altering the structure of enzyme proteins may change the substrate specificity and confer new catalytic activities to the mutant enzyme. We cannot cite 3 specific

1028 La Du

Table activity normal

III. Ratio of erythrocyte in heterozygous individuals homozygous individuals*

enzymatic to

Heterorygote activity

Normal homorygote mean activitv

Condition Galactosemia Methemoglobinemia Acatalasia Glucose-6-phosphate dehydrogenase deficiency Caucasian Negro Pvruvate kinase deficiencv *Data J. Med.

taken 34:633,

from Childs, 1963.6

mean

0.49 0.48 0.41

0.43 0.56 0.45 B.,

and

Young,

W.

J.: Am.

example of this occurring in man, but it has been suggested that the unusual esterase in some rabbits which catalyzes the hydrolysis of atropine has arisen by a slight mutational change in structure of some other protein.26 Another mechanism by which genetic mutations might lead to an enzymatic deficiency would be by mutation of a control gene. Here we would expect the enzyme formed to be qualitatively normal but quantitatively different. It is obvious that in hereditary conditions in which there is no detectable enzyme activity, we cannot tell whether the primary defect has been a structural or control gene mutation. In some instances, it may be possible to identify the defect as a structural gene mutation by showing the presence of a protein analogous to the enzyme, but lacking enzymatic activity. If purified normal enzyme is available as an antigen, immunochemical methods may show a cross reaction with the structurally altered protein. Little progress has been made applying this technique to the enzyme deficiency metabolic disorders. However, Nishimura and associateP3 found no cross reacting protein with antiserum prepared against purified catalase in acatalasic individuals. The level of enzyme in heterozygous individuals may be helpful in determining whether we are dealing with a structural Or control autosomal gene mutation. Very little is known about the control mechanisms

which regulate the rate of enzyme synthesis in mammalian cells, but if the mechanisms are similar to those proposed in bacteria,!” we would expect that control gene mutations in single dose could result in enzyme levels anywhere from zero to well above the normal values. This wide spectrum of activity levels would be possible because control genes are believed to regulate both allelic operator lo4 On the other hand. genes in diploid cells. structural genes show little interaction and generally there seems to be a quantitative relationship between the level of enzyme and the number of normal structural genes present. When we examine the level of enzyme in heterozygotes in five conditions which are probably structural gene mutations (Table III), we find that the level is close to half that found in normal cells in each instance. This quantitative relationship is more complicated when we consider the sex-linked conditions, the glucose-6-phosphate dehydrogenase deficiencies. Female heterozygotes show a rather wide range of enzyme levels, though the mean values for the group is approximately half of that found in normal females.los Several hypotheses have been proposed to explain the relationship between dosage level of the X-linked genes and the enzyme levels (see Harri?) . One attractive hypothesis, proposed by Lyon:loFs lo7 is that only one X chromosome is functionally active in female (XX) somatic cells, but in male somatic cells (XY) , the X chromosome is always functional. This hypothesis would explain why the enzyme level of glucose-6phosphate dehydrogenase in normal females is about the same as it is in normal males (instead of twice as high). If the inactivation of the X chromosome in female heterozygous for the deficiency were predominantly either the normal or the mutant gene-containing X chromosome, the resulting enzyme activity could range from nearly fully deficient to normal values. The Lyon hypothesis further implies that the individual cells in heterozygous females should be either deficient or normal in their dehydrogenase activity. Recently, two laboratories, using different methods 21o8p*OS have demonstrated that there are

Inborn

two distinct cell populations, normal and deficient, in females heterozygous for this dehydrogenase deficiency. Four types of glucose-6-phosphate deficiency conditions have been described; in two of them the enzyme properties differ from the normal and can be assumed to be structural mutation variants, in the other two types the enzyme appears to be normal qualitatively but reduced in the amount formed.5 The latter may be examples of control gene mutations. It is difficult to cite any enzymatic metabolic deficiency in man as an example of a clearly established regulatory gene mutation. Parker and BearnIl have recently discussed the problem of regulatory mechanisms in human genetics and the difficulty in classifying metabolic disorders as control or structural gtbne mutations. They consider most of the metabolic disorders listed in Table I (with the exception of atypical pseudocholinestcrase) to be control gene mutations, but many investigators would not agree with this interpretation. We would like to have an example as unequivocal as the hereditary persistence of fetal hemoglobin, the so-called high-fetal hemoglobin condition.lll It is of interest that Granick and Urata havelIz recently found that a number of drugs induce a marked increase in the liver S-aminolevulinic acid synthetase, the rate limiting enzyme in porphyrin synthesis, to forty times its normal level. He proposes that the mechanism of this induction is by activation of the gene for S-aminolevulinic acid synthetase by combining with and thus inhibiting a repressor.“” This would be an example of enzyme induction by “depression,” and the enzyme system seems particularly useful for further studies on the control mechanisms in mammalian cells. Granick and LeverelI* have recently suggested that the inborn error in the congenital hepatic type of porphyria may be a defect in the repressor of the synthetase which allows several drugs to induce the enzyme and lead to an excessive formation of porphyrins. Other hereditary mechanisms should be mentioned which could lead to enzymatic

errors

of metabolism

7O29

deficiencies in more indirect ways. One passibility is through the formation of an inhibitor which would decrease the effectiveness of the enzyme in vivo. The inhibition of melanin synthesis in phenylketonuria b? the accumulation of phenylalanine metabolites115 would be an example. Another indirect mechanism is an enzymatic deficiency due to the failure in development or ditherentiation of particular cells in which the r’nzyme is synthesized. Such a failure would bc analogous to the lack of gamma glob111in when plasma cells are not formed or nlaintained.l16 There are practical reasons why it is KIIportant to investigate and determine more precisely the types of mechanisms operating in the various metabolic disorders associated with altered enzymatic activity. Whether the patient is likely to have a hemolytic episode from fava beans or from primaquine depends upon the type of glucose-6-phosphate dehydrogenase deficiency he carries,5 and the Iikelihood of reacting to succinylcholine \vith prolonged apnea depends upon the typ(x of pseudocholinesterase variant he has illllerited.l” In predictions of the affected offspring from information on the genotypes of the parents, accurate data are needed. For example, we might expect that if both parents are albino, they would not have a child with normal pigmentation. However, such an occurrence has been reported,l*” and may be due to the fact that the primary defect in both parents is not exactly the same. Kecently, evidence has been presented which suggests that albinism in man may be dur: to more than one type of mutation.‘* Detection of heterozygotes. Even thong!1 heterozygote carriers of abnormal genes generally show very little evidence of a metabolic deficiency under ordinary circumstances, many attempts have been made to develop methods to distinguish these individuals from normal homozygotes and to determine quantitatively the extent of the enzymatic cleficiency they carry. In general, two experimental approaches have been used; by direct enzvmatic assay, providing the enzyme ill yuestion occurs in tissues accessible and suitable

for enzymatic analysis, and by tolerance tests using loading doses of the particular metabolite which best taxes the capacity of the enzyme system in expectation that the heterozygous carrier will show a reduced ability to metabolize the compound. The instances in which a direct enzymatic assay has been possible (practically, now limited to either erythrocytes or white blood cells) have given a clearer distinction between the normal and heterozygous individuals than the tolerance tests, and the enzymatic assays have shown values in the heterozygotes to be approximately half the normal levels (Table III). In metabolic disorders such as phenylketonuria, in which the defective enzyme is found only in liver, the loading test with phenylalanine does not give as high a degree of discrimination as desired. There is adequate evidence that the mean values obtained for groups of normal and heterozygous carriers differ significantly, but the classification of an individual with any confidence into one group or the other is much less secure.llg* l*” It is not unexpected that tests of heterozygosity based upon the response to oral loading doses are less satisfactory, since the measurement of enzymatic activity is more indirect and subject to additional variables, such as the rate of absorption of the test compound. Although we can expect that improvements in tolerance tests will reduce the chances of misclassification, it is apparent that new methods based upon a direct assay of enzyme activity will offer the most hopeful means of improving our detection methods. Under special circumstances, other methods have been useful in detecting heterozygotes. For example, in one type of cystinuria, heterozygotes excrete cystine and lysine in amounts well above the normal, though considerably below the levels found in affected homozygotes. This type of cystinuria has been called “incompletely recessive” cystinuria, to distinguish it from the “recessive” type, in which no elevated cystine or lysine excretion is found in heterozygotes.3, 4 The individual heterozygous for the atypical pseudocholinesterase in plasma can be detected by a determination of the sensitivity of the plasma

esterase to inhibition by dibucaine.‘:’ Heterozygotes show dibucaine inhibition “numbers” of about 60, whereas normal individuals have values about 80, and those homozygous for the atypical enzyme have values about 20. Epithelial tissue culture cells grown from skin biopsy may maintain the enzymatic defect of the donor. The level of enzyme in cells from heterozygous donors has been found to be intermediate between that of normal and homozygous affected donors in acatalasia,‘“l galactosemia,‘2S and white cells show intermediate values in the maple syrup urine disease.lZ3 Human cell lines with these genetic “markers” will be of great value in future investigations on biochemical and genetic aspects of hereditary metabolic disorders.

Detection and diagnosis of inborn metabolic disorders. There is nothing in the clinical signs and symptoms of the hereditary metabolic disorders due to enzymatic deficiencies which clearly differentiates this class of diseases from nonhereditary metabolic disorders for the physician. Only by detailed study of the individual patients and their families can the inherited basis of the disease be established. One might expect that hereditary diseases would make their appearance in early life and remain constant throughout the lifetime, but even this generalization does not hold. For example, the onset of symptoms of porphyria or the signs of ochronotic arthritis in alcaptonuria may be delayed until adulthood or middle age. The individual with the atypical plasma pseudocholinesterase will probably remain undetected throughout life unless he is given succinylcholine during an operation. However, in most of the inherited disorders of metabolism, there are abnormal metabolites in the blood and tissues and unusual products in the urine which are characteristic indicators of the disorder, and diagnosis requires the detection and identification of the abnormal metabolites, and, in some conditions, assay of the enzyme system under suspicion. In the past, metabolic disorders have been detected by physicians who observed

Volume Number

90 7, part 2

Inborn

errors

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1031

some unusual sign or symptom of a patient or a peculiar laboratory test response, A

diagnosis in very young babies of conditions in which early dietary treatment is beneti-

false positive urine test for glucose with Fehling’s reagent might uncover a new case of pentosuria, galactosemia, fructosuria, or alcaptonuria. One of the unfortunate consequences of our trend to use more specific methods is that by testing for urinary glucose with glucose oxidase we will miss the other reducing compounds mentioned above. Any disadvantage this represents is more than offset by the increased number of analytical techniques available today, such as paper and coiumn chromatography, amino acid analyzers, starch-gel electrophoresis, spectrophotometric, and enzymatic assays, which can be employed to detect and characterize metabolic variants. The advances in methodology, however, do not replace the need for preception and inquisitiveness on the part of the physician. A peculiar odor of the urine led to investigation of the first cases of both phenylketonuria and the maple syrup urine disease. An unusual reaction of a patient’s urine during analysis for creatinine resulted in finding the first (and only) case of tyrosinosis.lz4 The first case of acatalasia was found by a Japanese doctor who observed that blood turned black when hydrogen peroxide was applied following oral surgery.’ The first case of histidinemia was initially thought to be another case of phenylketonuria because the urinary ferric chloride test ,gave the typical olive-green color.*o Later it was found that the amino acid elevated in the blood was histidine, not phenylalanine, and the urinary metabolite responsible for the color with ferric chloride was the keto acid of histidine rather than phenylpyruvic acid. This illustrates the necessity of confirming the diagnosis with a specific test if a nonspecific reagent has been used to detect the condition. Specific confirmatory tests have been developed for many of the metabolic disorders, such as galactosenlia,‘25 phenylketonuria,71 histidinemia,lzE and the maple syrup urine disease.l”’ There are special problems in making a

cial (galactosemia, phenyfketonuria, and the maple syrup urine disease). Abnormal metabolites appear in the blood earlier than in the urine in all three conditions, and by employing specific blood tests the diagnosis can be established within the first week after birth. The diagnosis may be missed if urine tests are made too early, before tire abnormal urinary metabolites appear. New cases of metabolic disorders ma); be found by searching in families with known affected members, or by screening large population groups. The physician in practice is likely to be involved in both methods of case detection. In taking a history, the ohstetrician is in an excellent position to hear of metabolic disorders in the family and to prepare for the special examinations and diagnostic tests which are indicated, particularly if the risk of a serious metabtllic disorder is unusually high. For example. if a sibling of the expected baby is known to have phenylketonuria or galactosernia: the chances that the new infant will be affected are one in four. Some investigators suspect that in galactosemia some damage may occur in utero, and have advocated a restriction of milk in pregnant mothers in this high risk group.“** 33 If further studies should indicate that dietary restrictions are indicated, the obstetrician would have the responsibility for the prenatal treatment. Screening programs to find new cases of specific metabolic disorders requires the examination of large populations if the condition is relatively uncommon. Blatherwickl’* examined over 22,000 urine specimens with a trace of sugar in a search for another case of tyrosinosis, without success. Aebi” found two cases of acatatasia in Switzerland by screening 18,459 blood samples in army recruits and from these individuals he found four additional acatalasic members of their families. General screening programs for cases of phenylketonuria, galactosemia, and, possibly, maple syrup urine disease are certain to be more extensive in the future as methods of

1032

La Du

treatment improve with those associated with mental retardation. In fact, routine testing of all babies for phenylketonuria has become a legal obligation in two states, New

York and Massachusetts. It is obvious that a considerable effort is being made to find all new casesas early as possibleand to prrrvent mental retardation by dietary treatment.

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Garrod, A. E.: Lancet 2: 1616, 1902. Garrod, A. E.: Inborn Errors of Metabolism, London, 1909, Frowde, Hodder & Stoughton. 3. Harris, H., Mittwoch, U., Robson, E. B., and Warren, F. L.: Ann. Human Genet. 19: 196, 1955. H., Mittwoch, U., Robson, E. B., 4. Harris, and Warren, F. L.: Ann. Human Genet. 20: 57, 1955. 5. Childs, B., and Young, W. J.: Am. J. Med. 34: 663, 1963. 6. Harris, H.: Garrod’s Inborn Errors of Metabolism, London, 1963, Oxford University Press. 7. Takahara, S.: Lancet 2: 1101, 1952. a. Kaziro, K., Kikuchi, G., Nakamura, H., and Yoshiya, M.: Chem. 85: 886, 1952. 9. Wyngaarden, J. B., and Howell, R. R.: Acatalasia, in Stanbury, J. B., Wyngaarden, J. B., and Fredrickson, D. S., editors: The metabolic basis of inherited disease, New York, 1960, McGraw-Hill Book Company, Inc. 10. Aebi, H., Heiniger, J. P., Butler, R., and HLssig, A., Experientia 17: 466, 1961. Il. Bongiovanni, A. M., and Eberlein, W. R., Metabolism 10: 917, 1961. 12. Childs, B., Grumbach, M. M., and Van Wyk, J. J.: J. Clin. Invest. 35: 2 13, 1956. 13. Lerner, A. B., and Fitzpatrick, T. B.: The control of melanogenesis of human pigment cells, In Gordon, M., editor: Pigment cell growth, New York, 1953, Academic Press, Inc. 14. Kukita, A., and Fitzpatrick, T. B.: Science 121: 893, 1955. 15. Fitzpatrick, T. B., and Kukita, A.: J. Invest. Derm. 26: 173, 1956. 16. Knox, W. E.: Am. J. Human Genet. 10: 249, 1958. 17. Fitzpatrick, T. B.: Albinism, In Stanbury, J. B., Wyngaarden, J. B., and Fredrickson, D. S., editors: The metabolic basis of inherited disease, New York, 1960, McGrawHill Book Company, Inc. la. Kugelman, T. P., &d Van Scott, E. J.: J. Invest. Dermat. 37: 73. 1961. 19. La Du, B. N., Zannoii, V. G., Laster, L.: and Seegmiller, J. E.: J. Biol. Chem. 230: 251, 1958. 20. Knox, W. E.: Am. J. Human Genet. 10: 95, 1958. 21. La Du, B. N.: Alcaptonuria, In Stanbury, J, B., Wyngaarden, J. B., and Fredrickson, D. S., editors: The metabolic basis of in1. 2.

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Inborn

44.

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