Inborn Errors of Metabolism - The Basis of Single Gene Disorders
Tufts' Canine and Feline Breeding and Genetics Conference, 2005
Urs Giger, DACVIM, ECVIM, ECVCP; Charlotte Newton Sheppard Professor of Medicine
Section of Medical Genetics, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania

Genetic defects include anatomical malformations, inborn errors of metabolism, as well as increased susceptibility to disease. There is a considerable degree of overlap and all can have a metabolic basis:

 Congenital skeletal malformations due to genetic defects have been clinically recognized for decades, but there are also many soft tissue and internal organ anomalies. Many are recognized at birth (congenital), but not all birth defects have a hereditary basis. Furthermore, some of the defects are associated with a single malformation, whereas others are part of a syndrome (chondrodysplastic Alaska malamute dwarf with stomatocytosis, group of mucopolysaccharidoses and other storage diseases in various animal species).

 "Inborn errors of metabolism" (Sir Archibald Garrod, 1901) includes today all biochemical disorders due to a genetically determined, specific defect in the structure and/or function of a protein molecule. Aside from the classical enzyme deficiencies, genetic defects in structural proteins, receptors, plasma and membrane transport proteins, and other proteins, covered by this definition, will result in biochemical/metabolic disturbances. Currently inborn errors refer to single gene defects. With the better characterization of hereditary disorders practically all genetic defects could be considered to be inborn errors of metabolism including malformations and susceptibility to disease.

 Increased susceptibility to disease has been recognized more recently to have a genetic basis. Single gene defects for a variety of genetic predispositions have been identified, whereas predispositions caused by complex/polygenic traits are being characterized and include predispositions to infections, inflammation, immune disorders, drug reactions, and neoplasia.

Certainly not all proteins are enzymes with catalytic function. Many proteins are structural or contractile in function, some represent receptors that mediate inter- or intracellular signals or endocytosis of nutrients, some are membrane ion channels or molecular transporters, others are adhesion proteins that mediate cell migration during embryologic development or inflammation. Proper function of any protein may depend not only on tissue specific or developmentally regulated transcription and translation, but also on posttranslational modifications, proper subcellular localization or secretion of a protein, protein stability, substrate or ligand affinity, cofactor binding, and homeostatic regulation. Most of these characteristics of an expressed protein are determined by the amino acid sequence and the inherent secondary and tertiary folding of the protein in question. Thus, any mutation affecting the coding sequence in some way can produce any of a variety of malfunctions of the mature protein.

Furthermore, many enzyme functions depend on the availability of a vitamin or high energy intermediate compound (cofactors). Therefore, in addition to those in which a mutation has altered the affinity of an enzyme for the cofactor, the class of cofactor-responsive metabolic diseases includes some in which mutations in loci distinct from that of the enzyme affect the normal absorption or conversion to the active form of the required cofactors. Of the metabolic disorders, these are the most amendable to traditional forms of therapy (parenteral or megadose vitamin supplementation).

Inborn errors of metabolism may lead to the dysfunction of a biological system or pathway either under normal conditions or during more demanding situations such as the presence of concurrent disease, since many are in the catabolic pathways. Screening tests should lead to the detection of the failing system. Routine tests such as a complete blood count and a chemistry screen may reveal a specific metabolic problem such as inclusions in white blood cells or hyperlipidemia, respectively. Imaging techniques, gastrointestinal and liver function investigations as well as renal clearance function studies may more clearly define an organ failure, while for others the first clue is found only after pathologic examination of tissues.


When a metabolic pathway is blocked by an enzyme deficiency, the substrate of that enzyme and other proximal metabolites either accumulate or divert into an alternate pathway. In contrast the products or distal metabolites subsequent to the enzyme deficiency will be reduced in their amount. In some cases, abnormal metabolites or excessive amounts of normal metabolites affect other metabolic pathways by acting as competitive substrates of another enzyme. Depending on the gene mutation the deficiency can be partial to complete and can lead to more or less severe clinical signs. One of the best examples of a common diagnostic work up based on metabolic pathways is clinical done when suspecting a coagulopathy localizing the defect to the intrinsic, extrinsic, or common pathway.

In the affected animal, the diagnosis of a metabolic enzyme deficiency can often be accomplished by detecting abnormal metabolites or metabolite concentrations in urine, serum, or cerebrospinal fluid. This was the basis of Garrod's initial studies on alkaptonuria (homogentisic acid, a defect in the catabolism of phenylalanine and tyrosine), cystinuria, and pentosuria. Although more sophisticated techniques of analysis can now be used, examination of proximal and distal metabolites is still the mainstay of the efforts to diagnose and characterize metabolic diseases. One proceeds from metabolite identification to the demonstration of an enzyme or other gene product defect by assays of the functions of candidate proteins chosen based upon the knowledge of metabolic pathways and the previous description of metabolic diseases in the same or other species. Metabolic disorders are therefore often named according to the aberrant substrate associated with the pathologic condition rather than the truly defective (deficient) enzyme or cofactor (e.g. cystinuria, lactic aciduria, methylmalonic aciduria, porphyria, mucopolysaccharidosis and other storage diseases).

One of the most useful specimen in testing for a suspected inherited metabolic disease is a urine sample. Urine is the preferred, because abnormal metabolites in the blood will be filtered through the glomerulus, but they fail to be reabsorbed, as no specific renal transport systems exist for most abnormal metabolites. In those cases in which normal metabolites accumulate, their quantities usually exceed the renal threshold. As a consequence the amount of such compounds in a given volume of urine is often several folds greater than in blood. Defects in renal tubular transport will also produce a drain of metabolites from the blood in the urine. The renal tubules do not have the capacity to reabsorb abnormal metabolites or excess normal metabolites, and they become concentrated as water is conserved. Samples from normal littermates can be helpful for comparison and can account for differences caused by diet and environment. The Metabolic Genetic Screening Laboratory at the University of Pennsylvania offers metabolic screening for animals with suspected hereditary diseases not previously described in a breed or species. This NIH-supported laboratory has discovered several dozen disorders over the past three decades and information and instructions on sample submission are at

The generally used Clinitest screens for reducing agents in the urine such as glucose. Glucosuria either due to diabetes mellitus or a tubular defect can be confirmed by the Clinistix reaction or paper carbohydrate chromatography. Although glucosuria is frequently encountered, the Acetest for ketone bodies is rarely positive in small animals.

An important group of inherited metabolic defects are lysosomal storage disorders which involve housekeeping enzymes in the continual turnover of intracellular or phagocytized waste products. Degradation of glycosaminoglycans (GAG), glycoproteins and neurolipids requires the sequential action of a number of enzymes, and deficiency of any one of them leads to the accumulation of the undegraded substrate. In some lysosomal storage diseases the undegraded substrate is excreted in the urine (in mucopolysaccharidoses [MPS]), but in others, detection of the accumulated substance requires histopathology and -chemistry (glycogenoses). In the simple spot test used to screen for mucopolysaccharidosis, toluidine blue produces a metachromatic purple reaction with urinary GAGs. The stain can also be used to detect GAG storage in white blood cells or other tissues. To further characterize the type of mucopolysaccharidosis, urinary GAGs are separated by cellulose acetate electrophoresis into dermatan, heparin, and chondroitin sulfate. It should be noted that very young and growing animals normally excrete chondroitin sulfate in the urine. Storage diseases other than mucopolysaccharidoses can be detected by other screening methods. A large number of storage diseases have been recognized in small animals.

The first canine MPS disorder identified was MPS VII in a mixed breed dog at the University of Pennsylvania 25 years ago. The study of this and other MPS types led to better understanding of the disease process and also to the first successful attempt of treating a multisystemic disorder with gene therapy. The same mutation causing MPS VII in the original mixed breed dog has now been identified to cause MPS VII in German Shepherds. MPS I was first recognized in the Plott hound, and has since also been identified in an isolated case of a Rottweiler. All MPS disorders are autosomal recessively inherited, except for MPS II seen in Labrador retrievers, which is inherited by an x-linked recessive mode. MPS III also known as Sanfilippo syndrome is unique in that this is the only MPS disorder with mostly neurologic signs such as ataxia and tremors. MPS IIIA occurs in the Dachshund and the New Zealand Huntaway dog. Most recently MPS IIIB has been found in Schipperkes. Typically they do not exhibit clinical signs until they are two years of age and their condition is slowly progressing until they deteriorate to be euthanized by 5 years of age. The disease-causing mutation has recently been identified, and the results from screening 1000 Schipperkes would indicate that the mutant allele seems to be very prevalent in the breed. MPS VI was first seen in Miniature Pinschers with stunted growth and skeletal abnormalities mostly involving the hips, hence they were misdiagnosed to have hip dysplasia or femur head necrosis (Legge-Calves-Perthes, LCP disease). The molecular defects in this breed as well as in Miniature Schnauzers have just been identified and screening of Miniature Pinschers for carriers and affecteds is now possible. The DNA tests for MPS disorders require cheek swabs or an EDTA blood sample and are being offered through the Josephine Deubler Genetic Disease Testing Laboratory at the University of Pennsylvania (


The cyanide nitroprusside reaction which detects any compound containing a sulfhydryl group is used to screen for cystinuria (and homocystinuria). Cystinuria and other amino acidurias can be detected by paper chromatography using butanol/acetic acid/water as a solvent and ninhydrin stain and High Pressure Liquid Chromatography. Interpreting these metabolic patters can be difficult as they are affected by diet. Cystinuria is caused by renal basic amino acid transporters which can also affect the reabsorption of other amino acids, and thus these animals have normal to low serum cystine concentrations. Type I cystinuria is most severe and the molecular defects have been identified in Newfoundlands and Labrador retrievers by researchers at Penn. The most severe generalized renal tubular defect involves glucosuria, lactic aciduria and generalized amino aciduria and is known as Fanconi syndrome. Interestingly a large amount of felinine excretion is normal for healthy cats, and lack of felinine is a marker for a sick or poorly nourished cat. Taurine deficiency, sarcosinemia, and hyperornithinemia are other examples of amino acid abnormalities.

Methylmalonic aciduria is a prime example of a metabolic defect. Methylmalonic acid is a metabolite of an alternative pathway that only accumulates, when there is a block in the catabolism of various amino acids, fatty acids, and cholesterol to the tricarboxic cycle. It may be caused by either an intermediary enzyme deficiency of cobalamin deficiency as cobalamin is a cofactor of a mutase. In several breeds of dogs, including Giant Schnauzers, Border Collies and Beagles, a selective malabsorption of cobalamin has been identified due to a lack of expression of the intrinsic-cobalamin receptor on the surface of the brush border of the enterocytes in the ileum. Cobalamin is also involved in transmethylation reactions along with folate and thus its deficiency affects cell growth and thereby hematopoiesis. Fortunately this serious metabolic disorder can be readily treated by regular parenteral supplementation of cobalamin. There are several other organic acidurias described in animals including lactic acidurias causing myopathies and primary hyperoxaluria.

Usually the biochemical phenotype of an enzyme deficiency is associated with the disease phenotype, and, as traits, they are inherited recessively, because most enzyme activities are present in excessive amounts than what is minimally necessary to process sufficient substrate for normal development and health. Enzyme activity is expressed in units or as percent of control which is 100% and generally activity levels of 30% are still sufficient for most biological functions. The most immediate effects are seen with key regulatory enzymes. One can often demonstrate the coexpression, or gene dosage effect, of the normal and mutant allele by measuring the activity or quantity of the protein (enzyme) in question in tissues of the affected individual, parent and littermates compared to normal unrelated control animals. In autosomal recessively inherited diseases, the affected individuals will have enzyme activities of < 20% and often 0-5% of the normal, unrelated animals, and the parents of the affected animal will have an enzyme activity somewhere around half of the normal (30-75% of normal control). This forms the basis of many carrier detection programs for hereditary diseases, but the efficiency of carrier detection by protein quantification or function can be seriously affected by various parameters. It is important to recognize that the Mendelian concepts of dominant and recessive modes of inheritance refer to the phenotypic (clinical) presentation of heterozygote and homozygote animals for a particular trait and does not refer to the protein level or gene (genotype).

Unfortunately, enzyme assays require generally a particular sample due to the tissue specific expression and control sample from an age matched animal of the same species. In addition, the collection, handling, shipping has to be carefully followed according to the instructions. Moreover, the enzyme activity is label and thus samples have to be cooled and analyzed shortly after collection unless frozen. Finally in vitro enzyme activity may not really reflect the in vivo expression and function, but may depend on substrate and cofactor availability and affinity. Despite the lack of functional activity of a protein in a disorder, the dysfunctional protein may or may not be present and can be detected be through immunological techniques. Thereby animals with and without a protein can be differentiated into cross-reacting material (CRM) positive and negative. More recently many metabolic diseases can be most accurately diagnosed by DNA testing for the disease-causing gene mutation. These tests are based on amplifying the DNA around the disease-causing mutation and differentiating the mutant from the normal sequence by sequencing, identifying fragment size differences with or without restriction enzyme digest. Such molecular readily permit the identification of normal, carrier and affected animals for recessive traits. However, mutation-specific DNA tests are species and breed specific.

In conclusion, the study of inherited metabolic diseases has come to encompass all biochemical disorders, structural or homeostatic, which result from a specific, genetically determined defect in the structure and/or function of a protein molecule. Since genetic alterations are possible at any gene locus, inborn errors of metabolism compromise a large heterogeneous group of monogenic disorders. Inborn errors of metabolism represent a major group of disorders in small animals that present with a large variety of clinical signs. Simple and inexpensive screening tests can readily recognize the failing biological system. Special laboratory tests, including biochemical and molecular genetic techniques, are usually required to make a definitive diagnosis and to detect carriers.

Intermediate metabolites detected in small animals with metabolic diseases

Urinary metabolite


Species, breed



Congenital diabetes mellitus

Many dog breeds

Renal glucosuria

Norwegian Elkhound

Fanconi syndrome

Basenji, others


Nursing animals


Hepatic disorders

Dogs with liver disease

Organic aciduria

Lactic acid

Lactic acidosis

Old English Sheep dog, Clumber spaniel, others

Fanconi syndrome


Methyl malonic acid

Cobalamin malabsorption

Giant Schnauzer, Beagle, Border Collie, Australian S.

Methyl malonic aciduria

Shar Pei

Isovaleric acid

Isovaleric aciduria


Oxalate, L-glycerate

Primary hyperoxaluria


Amino aciduria


Fanconi syndrome

Basenji, others

Cystine and dibasics

Cystinuria type I

Newfoundland, Labrador

Cystinuria non-type I

Many breeds, cats


Gyrate atrophy

Domestic shorthair cats


Tyrosinemia type II

German Shepherd



Portugese water dog

Alanine, glutamine

Hepatic diseases, shunts

Many dogs

Felinine deficiency

Sick, malnourished cats

Taurine deficiency

Retinal atrophy or cardiomyopathy


Lysosomal Storage Diseases in Animals


Deficient Enzyme


Mucopolysaccharidosis (MPS)

(Hurler & Scheie Syndrome)


DSH cat, Plott hound*, Rottweiler

(Hunter Syndrome)

Iduronate-2-sulfate sulfatase

Labrador retriever dog

(Sanfilippo A Syndrome)

Heparan N-sulfatase, Sulfamidase

Wirehaired dachshund
New Zealand Huntaway dog

(Sanfilippo B Syndrome)


Schipperke dog

(Sanfilippo D Syndrome)


Nubian goat

(Maroteaux-Lamy Syndrome)

(Arylsulfatase B)

Siamese & DSH cats, rat,
Miniature pinscher, Welsh corgi, Miniature Schnauzer,
Chesapeake Bay Retriever

MPS VII (Sly disease)


Mixed breed, German shepherd
DSH cat, Gus mouse





English springer spaniel dog

Glycogenosis II
(Pompe Disease)


Lapland dog
DSH cat, Corriedale sheep, Shorthorn cattle & Brahman cattle, Japanese quail



Persian, DSH & DLH cats Angus, Murray gray, & Galloway cattle
Guinea pig



Anglo-Nubian goat, Saler cattle


Galactosylceramide lipidosis
(globoid cell leukodystrophy, Krabbe disease)


Cairn & West Highland white terrier Miniature poodle, Beagle, Irish setter & Blue tick hound
Dorset sheep
Twitcher mouse
DSH cat
Rhesus monkey

(Gaucher disease)

Acid β-glucosidase

Sydney silky terrier dog



Siamese, Korat & DSH cat Beagle mix, Springer spaniel, Portuguese water dogs, Friesian cattle,
Suffolk sheep, sheep

(Sanhoff disease)

β-hexosaminidase A and B

DSH & Korat cats
Japanese spaniel, German short-haired pointer
Yorkshire pig

Mucolipidosis II
(I-cell disease)


DSH cat

Sphingomyelinosis A and B (Niemann-Pick A and B)

Acid sphingomyelinase, Cholesterol esterification deficiency (type C)

Balinese, Siamese & DSH cat
Boxer, Miniature poodle

Ceroid lipofuscinosis

Several enzymes

Border collies and many breeds


1.  Scriver CR, et al. (eds): The Metabolic and Molecular Bases of Inherited Disease. 8th edition. McGraw-Hill, New York, 2001.

2.  Desnick RJ, Patterson DF, Scarpelli DG (eds): Animal Models of Inherited Metabolic Diseases. Progress in Clinical and Biological Research, 94, 1982.

3.  Kaneko JJ, Harvey JW, Bruss ML (eds): Clinical Biochemistry of Domestic Animals, 5th edition, Academic Press, San Diego, 1997.

4.  Ostrander E, Giger U, Lindblad-Toh K (eds): The Dog and its Genome. Cold Spring Harbor Press, October 2005.

5.  Giger U: Clinical Genetics. In Textbook of Veterinary Internal Medicine. Ettinger S, Feldman E. Saunders, Philadelphia, 2005.

6.  Giger U and Jezyk PF: Diagnosis of inborn errors of metabolism in small animals. In Current Veterinary Therapy XI. Ed. RW Kirk, WB Saunders, Philadelphia, pp 18-22, 1992.

7.  Countless original articles on specific disorders.

Speaker Information
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Charlotte Newton Sheppard, Professor of Medicine
Section of Medical Genetics
School of Veterinary Medicine, University of Pennsylvania
Philadelphia, Pennsylvania

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