Screening Newborns for Inherited Metabolic Disease

Screening Newborns for
Inherited Metabolic Disease
Marsha F. Browning, MD, MPH; Deborah L. Marsden, MD
As treatment for hereditary
metabolic disorders continues
to evolve—especially early
intervention to prevent longterm
morbidity—neonatal
screening is emerging from
the theoretical realm to take its
place in the clinical setting.
Most inborn error-of-metabolism
(IEM) conditions are
single-gene disorders that
are inherited in an autosomal-recessive
fashion. They
result in a deficiency of a single enzyme
in the catabolic pathway of proteins, fatty
acids, or complex carbohydrates. Generally,
this deficiency leads to a biochemical
derangement, toxic accumulation, or depletion
of an end-product that can be acute or
chronic. Many of these disorders can now
be detected by newborn screening, enabling
presymptomatic diagnosis and possibly preventing
major morbidity. 1
Marsha F. Browning, MD, MPH, is assistant in Pediatrics,
Harvard Medical School, Massachusetts General
Hospital, Boston. Deborah L. Marsden, MD, is
assistant professor, Department of Pediatrics, Harvard
Medical School; and attending physician, Division of
Genetics (Metabolism Program), Children’s Hospital
Boston, Mass.
INHERITED METABOLIC DISEASES
Amino Acid Disorders
Amino acidopathies are caused by enzyme
defects early in the catabolic pathway of
the specific amino acid, resulting in its
toxic accumulation. The hallmark disorder
is phenylketonuria (PKU), 2 where a deficiency
of phenylalanine hydroxylase results
in the accumulation of phenylalanine, which
is toxic to the central nervous system and
causes severe, irreversible mental retardation
if not treated early. Treatment includes a
protein-restricted diet and supplementation
with a phenylalanine-free formula to ensure
adequate nutrition for normal development.
Adults in whom the phenylalanine-restricted
diet is relaxed or discontinued have significant
neuropsychiatric problems, including
anxiety and depression. 3,4 In “maternal
PKU,” the mother is affected with PKU
and the elevated phenylalanine levels are
teratogenic to the fetus. 5 Therefore, lifelong
dietary restriction of phenylalanine is now
recommended for all patients.
Organic Acid Disorders
Organic acidemias are due to enzyme deficiencies
in the catabolic pathways of one or more
amino acids, resulting in the accumulation
of intermediary organic acids. The result is
typically severe, recurrent metabolic acidosis
and ketonuria. Hypoglycemia and hyperammonemia
may be present in varying degrees
due to secondary inhibition of fatty acid
oxidation and the urea cycle, respectively.
Clinical presentation is often at 7 to 10 days
after birth, but some disorders can present at
age 2 to 3 days. Milder variants of organic
acidemias can present later in infancy or
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Browning and Marsden
childhood. Acute treatment requires aggressive
management of the acidosis, limitation
of protein intake, and an infusion of
> 10% dextrose solution to correct hypoglycemia
and reverse catabolism. 1 Long term,
these patients require protein restriction and
supplementation with a metabolic formula
that is depleted in the toxic precursor amino
acids. Propionic acidemia is a characteristic
organic acidemia caused by a deficiency of
propionyl-coenzyme A (CoA) carboxylase in
the catabolic pathway of isoleucine, valine,
methionine, threonine, cholesterol, and oddchain
fatty acids. 6
Fatty Acid Oxidation Disorders
Fatty acid oxidation defects (FAODs) are due
to enzyme deficiencies in the -oxidation
pathway of fatty acids in the mitochondria.
This results in accumulation of toxic fatty
acid intermediates and depletion of acetyl
CoA, which is the substrate for ketogenesis
and glucose—the final end-products of normal
fat metabolism. Characteristic features
include hypoketotic hypoglycemia, hyperammonemia,
and hepatic encephalopathy.
Cardiomyopathy may also occur due to
deposition of fatty acids in the myocardium,
especially in the long-chain defects. Symptoms
may present in the neonate, but are
TABLE 1. Disorders Covered by
Conventional Newborn Screening*
Biotinidase deficiency
Congenital adrenal hyperplasia
Cystic fibrosis
Congenital hyperthyroidism
Sickle cell disease
Galactosemia
Glucose
Table
-6-phosphate
not available
dehydrogenase
online
deficiency
Human immunodeficiency virus
Phenylketonuria
Toxoplasmosis
*Using Guthrie filter paper.
more common in infancy. 7-10 In addition,
maternal hemolysis, elevated liver enzymes,
and low platelets (HELLP syndrome) in
the third trimester may signal an FAOD
in the fetus; this seems to be
more prevalent with long-chain
defects such as long-chain
3-hydroxyacyl-CoA dehydrogenase
deficiency, although it
may occur with all FAODs. 11
The most common FAOD is
medium-chain acyl-CoA dehydrogenase
deficiency, with
a prevalence of one in every
12,000 in the United States
and a carrier rate estimated at
one in every 40. 7 The clinical
presentation is typically at age
9 to 15 months. 12 Approximately
30% of patients died
soon after presentation, but
since the advent of screening, mortality has
declined dramatically. 13 Treatment for FAODs
comprises avoidance of prolonged fasting and
early intervention during acute illness to
prevent hypoglycemia, including intravenous
dextrose bolus and 10% dextrose-saline infusion.
Carnitine and medium-chain triglyceride
supplementation may also be required.
NEWBORN SCREENING
Today, with
the introduction
of tandem mass
spectrometry,
more than
30 metabolic
disorders can
be detected
in newborns.
In 1959, Robert Guthrie, MD, developed a
bacterial-inhibition assay to measure serum
phenylalanine levels for children with known
PKU. Soon thereafter, a test was devised using
a filter-paper specimen to screen newborns. 2
From this test, a range of newborn screening
modalities have evolved (Table 1). Today, with
the introduction of tandem mass spectrometry
(MS/MS), 14,15 more than 30 metabolic disorders
can be detected in newborns. 16
Test Selection
Currently, each state determines its own newborn
screening policy on the basis of local demographics
and resources. Consequently, there is
great variation across the country in the number
of disorders screened. In 2000, the Newborn
Screening Task Force, convened by the Health
Resources and Services Administration (HRSA)
The Female Patient VOL. 31 DECEMBER 2006 19

Screening Newborns for Inherited Metabolic Disease
Blood Spots
Known
Internal
Standards
Inject
Into
Capillary
Inlet
Inert Gas
Collision Chamber
Mass Figure not + available - + online
- + -
Spectrometer
I
Parent Ions
First Quadrupole
Fragmentation
Second Quadrupole
FIGURE. Tandem mass spectrometry.
(Scanned)
+ Ions
of the US Maternal Child Health Bureau,
published guidelines in collaboration with the
American Academy of Pediatrics recommending
greater uniformity among screening programs. 17
In addition, the American College of Medical
Genetics (ACMG) has developed recommendations
for a standardized screening panel. 18
Expanded-panel Newborn Screening
The MS/MS technique uses two mass spectrometers
linked in tandem via a collision chamber
plus software capabilities (Figure), and takes less
than 2 minutes to process each sample. 14,15 It
identifies and quantifies amino acids and acylcarnitines—the
naturally occurring conjugates
of fatty acid and organic acid intermediates—by
recognizing characteristic mass spectra (molecular
weights) of these compounds in a single test.
The test can tentatively identify more than 25
additional metabolic diseases that were not previously
detectable using conventional newborn
screening techniques (Table 2). 19-21
Forty states currently offer testing using MS/
MS, but the actual number of tests offered
- Ions
Mass
Spectrometer
II
Daughter Ions
Unchanged
Ions
Third Quadrupole
Data Recorded
on Disk
(Scanned)
Dried blood spots are injected into the tandem mass spectrometer. They are separated into parent
ions in the first quadrupole and daughter ions in the third quadrupole. Data are recorded via
computer software at the beginning and end of the process to identify compounds by weight.
Courtesy of Marsha F. Browning, MD, MPH.
by each state still varies
widely. 16 Concerns
about expanded-panel
screening include the relatively
high initial capital
costs and the level
of technical expertise
required for testing and
interpretation. However,
the incremental
cost for additional tests
is low. 19-21 The availability
of trained metabolic
specialists and resources
for confirmation and
follow-up of abnormal
screening results must
be addressed as well.
Screening Versus
Diagnosis
The cut-off values for
abnormal metabolic
screening results are set
to detect most affected
infants without missing
cases or yielding an
excessive burden of false-positive results.
An abnormal elevation in a particular analyte
is not diagnostic, but rather requires
further testing. Each screening laboratory
determines its own appropriate cut-off values.
In Massachusetts, where expanded
newborn screening has been in place for
over 7 years, the rate of false-positive
results has been approximately 0.05% 22 —
ie, within the acceptable range. Causes of
false-positive findings include physiologic
enzyme immaturity, which can cause transient
analyte elevations.
Several compounds have the same molecular
weight. Examples include leucine, isoleucine,
and hydroxyproline, so that further testing
is required to differentiate between a
metabolic disease—in this case, maple syrup
urine disease (elevated branched-chain amino
acids)—and hydroxyprolinemia, which is a
benign disorder. 22 Thus, infants with positive
screening results should be referred to a metabolic
center for further diagnostic evaluation
and treatment.
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Browning and Marsden
Testing Algorithms
If an analyte value is outside the normal range,
the result is determined to be either “likely
affected” (requiring immediate referral to a
metabolic center), or “borderline abnormal”
(necessitating additional evaluation by the
primary care physician and a repeat filterpaper
specimen). Some programs recommend
specialist referral for any out-of-range specimen.
Certain disorders require immediate specialist
evaluation after an initial abnormal
result because of potentially devastating early
symptoms—eg, propionic acidemia, methylmalonic
acidemia, isovaleric acidemia, maple
syrup urine disease, citrullinemia, very–longchain
acyl-CoA dehydrogenase deficiency, trifunctional
protein deficiency, and long-chain
hydroxyacyl-CoA dehydrogenase deficiency. 23
For out-of-range results evaluated in the primary
care setting, clinical symptoms that may
indicate serious disease include irritability,
lethargy, vomiting, poor feeding, tachypnea,
hepatomegaly, ketonuria, and hypoglycemia.
Any clinical concern warrants immediate consultation
with, or referral, to a specialist.
Controversies
Although expanded-panel newborn screening
has greatly enhanced the ability to detect many
metabolic disorders presymptomatically, it is
not without disadvantages. Questions have
arisen about what to do with “benign” variants
and/or diseases that have no treatment
at this time. Additionally, there are wide
state-to-state discrepancies in availability of
the MS/MS technology and in opinions as
to which disorders to include in testing. The
expanded screening panel recommended for
all states by the ACMG has been posted on
the HRSA Web site to an overwhelmingly
positive public response. 24
CONCLUSION
Tandem mass spectrometry has vastly broadened
the horizons in newborn screening. The
technique can also be used for DNA analysis,
and may ultimately increase the sensitivity of
an initial abnormal result through second-tier
screening. This process is not yet fast enough
for IEM disorders, where early notification
may be crucial. Disorders that may be screened
by MS/MS in the future include lysosomal
storage conditions, X-linked adrenoleukodystrophy,
type 1 diabetes, severe combined
immunodeficiency, hereditary hemochroma-
TABLE 2. Disorders Screened by
Tandem Mass Spectrometry
Aminoacidopathies
Argininosuccinate lyase deficiency
Argininosuccinate acidemia
Citrullinemia
Homocystinuria
Maple syrup urine disease
Phenylketonuria
Hyperphenylalaninemia
Hyperammonemia hyperornithinemia
homocitrullinemia
Tyrosinemia (types 1 and 2)
Organic Acidemias
Glutaric acidemia, type 1
3-Hydroxy-3-methylglutaryl-CoA lyase deficiency
Isovaleric acidemia
Table not available online
3-Methylcrotonyl-CoA carboxylase deficiency
Mitochondrial acetoacetyl-CoA thiolase deficiency
(-ketothiolase deficiency)
Propionic acidemia
Methylmalonic acidemia
Fatty Acid Oxidation Defects
Carnitine transporter defect
Carnitine palmitoyltransferase (types 1 and 2)
Very–long-chain acyl-CoA dehydrogenase deficiency
Long-chain acyl-CoA dehydrogenase deficiency
Medium-chain acyl-CoA dehydrogenase deficiency
Short-chain acyl-CoA dehydrogenase deficiency
Trifunctional protein deficiency
Glutaric acidemia, type 2
The Female Patient VOL. 31 DECEMBER 2006 23

Screening Newborns for Inherited Metabolic Disease
tosis, and lymphoblastic leukemia. 25-30 As the
technology continues to advance, the need for
close cooperation among public health personnel,
primary care providers, and metabolic
referral centers will only increase.
REFERENCES
1. Fearing MK, Marsden D. Expanded newborn
screening. Pediatr Ann. 2003;32(8):509-515.
2. Efron ML, Young D, Moser HW, MacCready
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Coding for Screening Newborns
for Inherited Metabolic Disease
Frank Vidal, MMC
Virtually all of the diagnostic tests discussed here, including
mass spectrometry (82541), are performed by laboratories
outside the physician’s office, and therefore are
not coded or billed by the rendering physician. However,
ordering these tests is part of the comprehensive
evaluation and management (E/M) codes (ie, well-child
visits) during which these screenings are performed. Procedural
elements are also covered by the E/M codes pertinent
to the newborn.
• 99381— Initial comprehensive preventive-medicine
E/M for new patient (infant, aged < 1 year),
including age- and gender-appropriate
history, physical examination, counseling,
anticipatory guidance, risk-reduction interventions,
and ordering of immunization(s),
laboratory/diagnostic procedures
• 99391— Periodic comprehensive preventive-medicine
reevaluation and management for established
patient (infant, aged < 1 year)
DIAGNOSTIC ELEMENTS
• 270.1— Phenylketonuria
–Hyperphenylalaninemia
Diabetes with Ketoacidosis (Acidosis and Ketonuria)
—Diabetic
• Acidosis without mention of coma
• Ketosis without mention of coma
• 250.10—Type 2 or unspecified type, not stated
as uncontrolled
• 250.11—Type 1, not stated as uncontrolled
• 250.12—Type 2 or unspecified type, uncontrolled
• 250.13—Type 1, uncontrolled
• 775.6—Neonatal hypoglycemia
• 251.1—Other specified hypoglycemia
–Hyperinsulinemia
–Not otherwise specified
–Ectopic
–Functional
• 270.6—Disorders of urea cycle metabolism
–Hyperammonemia
• 277.85—Disorders of fatty acid oxidation
–Long-chain 3-hydroxyacyl-coenzyme A
(CoA) dehydrogenase deficiency
–Long-chain/very–long-chain acyl-CoA
dehydrogenase deficiency
–Medium-chain acyl-CoA dehydrogenase
deficiency
–Short-chain acyl-CoA dehydrogenase
deficiency
Frank Vidal, MMC, is chairman, United States
chapter, International Academy of Medical Coding.
24 The Female Patient VOL. 31 DECEMBER 2006

Browning and Marsden
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Acknowledgement
The authors wish to acknowledge the assistance
of the New England Newborn Screening
Program, Jamaica Plain, Mass.
The Female Patient VOL. 31 DECEMBER 2006 27