Multiple Carboxylase Deficiency (Biotinidase deficiency)

Are You Confident of the Diagnosis?

Multiple carboxylase deficiency is a rare inborn error of biotin metabolism caused by defects in biotinidase or holocarboxylase synthetase in the biotin cycle. Biotin serves as a cofactor for four carboxylases: 3-methylcrotonyl carboxylase, propionyl-CoA carboxylase, acetyl-CoA carboxylase and pyruvate carboxylase. Holocarboxylase synthetase catalyzyes the covalent addition of biotin to the four carboxylases, thereby activating the enzymes. When the caboxylases are degraded, biotinyl-lysine is produced. Biotinidase catalyzes the recovery of protein-bound biotin by hydrolyzing the biotnyl-lysine to release free biotin which is recycled to activate newly synthesized carboxlyase enzymes.

Characteristic findings on physical examination

Multiple carboxylase deficiency may occur early in infancy as result of holocarboxylase synthetase deficiency. These infants present with poor feeding, vomiting, lethargy, coma and seizures. The urine has the strong odor of a cat’s urine. Metabolic acidosis occurs with an increased anion gap, hyperammonemia and abnormal urine organic acids characteristic of ketosis: increased lactic acid, 3-hydroxy isovaleric acid, 3-methylcrotonylglycine, methylcitrate, hydroxy-propionate and propionylglycine . Skin rash (ichthyosis) and alopecia are more common in later onset cases.

Biotinidase deficiency is associated with a later and more gradual onset characterized by hypotonia, seizures, ichthyosis and alopecia totalis. Untreated patients experience moderate to severe mental retardation/disabilities. Hearing loss is common.

Expected results of diagnostic studies

Babies with multiple carboxylase deficiency are normal at birth and should be ascertained in the first few days or weeks of life by newborn genetic screening. Newborn screening for biotinidase deficiency is mandated in all 50 States. Those not detected early on become lethargic and feed poorly with vomiting and poor weight gain and may be referred to feeding clinic and gastroenterology. The physician should confirm the “normal” newborn screen in these patients because the metabolic defect is so readily treated and symptoms reversed.

Newborn screening from dried blood spots can detect biotinidase deficiency within days after birth. Newborn screening identifies common mutations, which then leads to confirming the deficiency by biotinidase assay . Biotinidase activity varies from one individual to another even with the same mutation, indicating that there may be epigenetic factors affecting enzyme level. Severe biotinidase deficiency is rare where as partial biotinidase deficiency appears to be common and may be associated with mild clinical manifestations or none at all.

Who is at Risk for Developing this Disease?

Multiple carboxylase deficiency is an uncommon metabolic disorder affecting 1 in 80,000 to 1 in 120,000 individuals. These enzyme deficiencies are inherited as autosomal recessive trait after the birth of an affected child, the couple will have a 1 in 4 (25%) risk of having an affected child in each subsequent pregnancy. The carrier frequency (heterozygote) in the general population is estimated to be 1 in 150 to 1 in 350 individuals.

An affected individual (homozygote) can only transmit the deficient allele to offspring. The offspring will be obligate heterozygotes for the specific mutant allele and unaffected unless the individuals mate is a carrier of a mutant allele.

What is the Cause of the Disease?

Mutations in the genes for holocarboxylase synthetase and biotinidase produce deficient or defective enzymes inhibiting the activation of the four carboxylases and limiting the recovery of protein bound biotin for reactivation of the carboxylases.


The gene for olocarboxylase synthetase is located on chromosome 21q22.1 and has 11 exons. There are at least 35 mutations known. The gene for biotinidase deficiency (BPD) is located on chromosome 3p25 and has 4 axons. Over 100 mutations have been reported, but 5 common mutations account for about 60% of the abnormal alleles.

Almost all individuals with partial biotinidase deficiency have the 444H mutation in combination with a mutation for profound deficiency on the other allele. Individuals with homozygote D444H/D444H alleles tend to have mild or intermediate biotinidase deficiency in the 20-30 percent of mean normal enzyme activity. D444H in cis arrangement with a 171T mutation produces a severe reduction in enzyme activity. When this combination is associated with a second severe allele, profound biotinidase deficiency occurs.

Genotype may aid in predicting the phenotype and degree of enzyme deficiency in some individuals. The great variability in clinical expression limits these correlations even in members of the same family. The various enzyme deficiencies that result from the various genetic mutations result in abnormalities of fatty acid synthesis, gluconeogenesis and amino acid catabolism. Clinical presentations may vary greatly creating difficulty in achieving a diagnoses on clinical symptoms alone.

Systemic Implications and Complications

The presenting symptoms of untreated multiple carboxylase deficiency are metabolic ketoacidosis, poor feeding and vomiting, generalized muscle hypotonia and lethargy, alopecia, hair loss and hyperkeratotic skin changes, hearing loss, optic atrophy, encephalopathy, coma and death. The considerable clinical heterogeneity ranges from profound deficiency and multiple systemic failures to intermediate deficiencies with primary cutaneous hair and skin changes to global developmental delay and mental retardation.

A range of late onset forms occur. Before the application of newborn screening, most late onset forms went undetected. Mild-to-moderate deficiencies with enzyme levels in the 20-30% normal mean enzyme activity have now been reported in the third and fourth decades of life with skin changes, hearing loss and mild learning disabilities .Diagnostic tests include plasma carnitine, plasma acylcarnitine profile, plasma amino acids, and urine organic acids on a routine urine sample.

This metabolic abnormality should now be detected in all affected newborns through the expanded newborn screening using tandem mass spectrometry and lead to presymptomatic therapy. Once the enzyme deficiency is detected, the child and family are referred to a genetics center for confirmation of the diagnosis and initiation of treatment.

Treatment Options

The systemic abnormalities associated with holocarboxylase synthetase and biotinidase deficiencies can be avoided with biotin supplementation.

In the neonatal period, biotin 5-10mg per day is given orally. As the child grows, biotin dose may be increased to 15-20mg per day by mouth. Biotin may be supplied in tablet form and in liquid preparations. Biotin may be added to small amounts of cereal or other food. Dosage in adults may be as high as 100mg per day. The optimal dose of biotin is unknown, but current dosage is empirical and may be adjusted as the child grows. The doses currently recommended are not necessarily based on evidence, but there is no apparent biotin toxicity.

Measuring biotinidase activity periodically appears not to have an impact on clinical treatment. Biotinidase levels will vary over time and depend on laboratory methods. The biotinidase assay is temperature sensitive and care must be taken to ensure proper landing of the specimen to the laboratory.

Monitoring urinary organic acids has been suggested as a means of assessing the effectiveness of treatment , but the variability in excretion patterns in patients with various degrees of deficiency has precluded the use of urinary organic acids as a routine.

Optimal Therapeutic Approach for this Disease

The critical aspect in approach to treating this metabolic disorder is early recognition and prompt biotin therapy. In those cases in which the diagnosis may be delayed for weeks or months, immediate biotin supplementation is expected to reverse the clinical symptoms.

Refer the child and parents for genetic consultation to confirm the positive newborn screen by determining biotinidase activity as percent of mean normal level. Severe deficiency is 5% or less and likely to have clinical consequences if not treated immediately. Therapy is considered mandatory for levels below 30% mean normal level. Treatment is considered to be life-long. Levels between 15% and 30% mean normal level are considered intermediate or moderate deficiencies and are unlikely to be associated with metabolic decompensation during periods of stress such as infections or fluid imbalances with biotin therapy.

The family needs to be aware that each child will have a 1 in 4 risk to have the familial mutations and have biotinidase deficiency. If there are other children and results of newborn screening can not be confirmed, the children should be tested.

Patient Management

The management of multiple carboxylase deficiency is evolving. Following the initial clinical evaluation and initiation of biotin supplementation, healthcare surveillance is standard well-child assessments with specific attention to achievement of developmental milestones.

Annual evaluation by a clinical geneticist is recommended to review the patient’s progress and search for any signs of biotinidase deficiency. Hearing and vision are assessed annually.

In the setting of acute decompensation, maintenance of fluid balance and correction of electrolyte abnormalities are paramount . Biotin may be given as the diagnostic tests are performed . Periodic assessment of biotinidase activity does not appear to play any roll in adjusting biotin supplementation. Biotinidase assays will very from month to month and year to year. This variation may have a physiological basis, but it also depends on laboratory standards and the manner in which the blood sample is collected and transported.

Unusual Clinical Scenarios to Consider in Patient Management

There is no evidence that vitamin supplements beyond biotin or additional metabolic cofactors are beneficial in this condition.

In any child who presents with unexplained seizures, encephalopathy, metabolic derangement, optic atrophy and developmental impairment, biotinidase deficiency should be considered. The patient’s medical record is considered incomplete unless the newborn extended screening is recorded.

Inborn errors of metabolism account for about 1% of heritable genetic conditions. In any patient at any age with evidence of systemic decompensation, screening for acute metabolic diseases would include complete blood count with platelets, electrolytes, plasma glucose, blood gases, ammonia level, plasma amino acids, carnitine and acylcarnitine profile, lactate and pyruvate with L/P ratio, very long chain fatty acids and, in the neonate and infant, confirmation of the newborn screening tests. Urine screening includes reducing substances, amino acid and organic acid excretion profiles.

What is the Evidence?

Wolf, B. “Clinical issues and frequent questions about biotinidase deficiency”. Mol Genet Metab. vol. 100. 2010. pp. 6-13. (This is the most current and comprehensive overview of biotinidase deficiency with a history goal perspective with a critical discussion of future clinical investigation.)

Wolf, B, Pagan, RA, Bird, TC, Dolan, CR, Stephens, K. “Biotinidase deficiency: late-onset biotin-responsive multiple carboxylase deficiency, late-onset multiple carboxylase deficiency”. Gene Reviews [Internet]. 1993-2010. (This review summarizes the disease characteristics, diagnosis and testing, management issues and genetic counseling issues associated with biotinidase deficiency.)

Wolf, B. “Biotinidase: Its role in biotinidase deficiency and biotin metabolism”. J Nutrit Biochem. vol. 16. 2005. pp. 441-5. (The biochemistry of biotinidase and biotin metabolism reviewed with emphasis on the effect oft he enzyme on biotin as a carrier protein.)

Pindolia, K, Jordon, M, Wolf, B. “Analysis of mutations causing biotinidase deficiency”. Hum Mutat. vol. 31. 2010. pp. 983-91. (This is a comprehensive a summary of the various mutations associated with biotinidase deficiency and the role of confirmational at testing when enzymatic results or equivocal.)

Seymons, K, De Moor, A, De Raeve, H, Lambert, J. “Dermatologic signs of biotin deficiency leading to the diagnosis of multiple carboxylase deficiency”. Pediatr Dermatol. vol. 21. 2004. pp. 231-5. (These authors review the skinand other systemic manifestations associated with multiple carboxylase deficiency.)

Gravel, RA, Narang, MA. “Molecular genetics of biotin metabolism: old vitamin, new science”. J Nutrit Biochem. vol. 16. 2005. pp. 428-31. (This is a thoughtful discussion of the role of bile tesselation of carboxylase in the relationship of biotin to histones and DNA regulatory functions.)

Arbuckle, HA, Morelli, J. “Holocarboxylase synthetase deficiency presenting as ichthyosis”. Pediatr Dermatol. vol. 2006. 23. pp. 142-4. (This is an informative case report of a newborn presenting with severe acidosis, feeding difficulties, reading abnormalities, vomiting, seizures, and progressive neurological depression with ichthyosiform changes of the eyebrows, eyelashes, and scalp.)

Tammachote, R, Janklat, S, Tongkobpetch, S, Suphapeetiporn, K, Shotelersuk, V. “Holocarboxylase synthetase deficiency: novel clinical and molecular findings”. Clin Genet. vol. 78. 2010. pp. 88-93. (This report reviews the clinical observations in four patients with multiple carboxylase deficiency in which comprehensive biochemical assays follow the patient through treatment with genotype-phenotype correlations.)

Monjaras, A, Cervantes-Roldan, R, Meneses-Morales, I, Gravel, RA, Reyes-Carmona, S, Solorzano-Vargas, S. “A. Impaired biotinidase activity disrupts holocarboxylase synthetase expression in late onset multiple carboxylase deficiency”. J Biochem. vol. 283. 2008. pp. 34150-8. (This is a comprehensive overview of impaired biotinidase activity in late onset multiple carboxylase deficiency presenting evidence to suggest that biotin supplementation does not only provide the product for defective biotinidase activity but also represses the holocarboxylase synthetase expression that is associated with biotin deficiency.)

Van Hove, JLK, Josefsberg, S, Freehauf, C, Thomas, JA, Thuy, LP, Barshop, BA. “Management of a patinet with holocarboxylase synthetase deficiency”. Mol Genet Metab. vol. 2008. 95. pp. 201-5. (This is a case report of the management of a patient with holocarboxylase synthetase deficiency with sequential plasma assessment of biotin concentrations and measurements in cerebral spinal fluid and blood establishing the pharmacokinetic variables that may aid in optimizing treatment.)