The North Carolina Newborn
Screening Pilot for MPS I
NBSTRN Network Meeting
September 8, 2016
Background
• RTI International, in partnership with the North Carolina State
Laboratory of Public Health, Duke University and the University of
North Carolina at Chapel Hill, was awarded an Indefinite Delivery,
Indefinite Quantity (IDIQ) contract from the Eunice Kennedy Shriver
National Institute of Child Health and Human Development to
promote rapid implementation of newborn screening pilot studies
that will field-test conditions recently added to the Recommended
Uniform Screening Panel (RUSP) in 2015.
• The first Task Order for the IDIQ was for mucopolysaccharidosis type I
(MPS I)
34 Core
Conditions
Currently on
the
Recommended
Uniform
Screening
Panel
Added February 2016
MPS I
• Four states currently screening
• To assist in implementation
• NIH/NICHD – IDIQ Task Orders
• MPS I Pilot Studies
• Contracts awarded to North Carolina
(RTI, State Lab, UNC-CH, Duke) and
Georgia (State Lab, Emory)
https://www.newsteps.org/mps-i
Overview of Mucopolysaccharidoses
(MPS)
• Lysosomal enzyme deficiency
– Eleven known enzyme deficiencies comprise seven
different clinical types
• Rare (est. incidence 1:25,000)
• Tissue storage of glycosaminoglycans (GAG)
• Progressive disorders with multisystemic
involvement
• Wide range of clinical involvement in each MPS
disorder
Courtesy of Joseph Muenzer, MD, PhD
Mucopolysaccharidosis I (MPS I)
• Deficiency of lysosomal enzyme -Liduronidase
• Onset of symptoms before 6 months of
age in severe form (Hurler syndrome)
• The severe form has both somatic and
CNS involvement
• Early mortality in severe form (3 to 10
yrs of age)
• Rare (est. incidence 1:100,000)
• Autosomal recessive disorder
Age 4
Courtesy of Joseph Muenzer, MD, PhD
Biochemistry of MPS I
Iduronidase deficiency causes
a block in the sequential
breakdown steps of
glycosaminoglycans
MPS I
e.g. Dermatan Sulfate Degradation
Courtesy of Joseph Muenzer, MD, PhD
Enlarged lysosomes
[STORAGE]
Normal fibroblast
nucleus
MPS fibroblast
Courtesy of Joseph Muenzer, MD, PhD
Spectrum of Disease in MPS I
Attenuated
Severe
Hurler-Scheie
MPS I H/S
Hurler
MPS I H
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Severe intellectual disability
More progressive
Severe respiratory disease
Obstructive airway disease
Death before age 10 years
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Little or no intellectual defect
Respiratory disease
Obstructive airway disease
Cardiovascular disease
Joint stiffness/contractures
Skeletal abnormalities
Decreased visual acuity
Death in teens and 20’s
Scheie
MPS I S
• Normal intelligence
• Less progressive
physical problems
• Corneal clouding
• Joint stiffness
• Valvular heart disease
• Death in later decades
Courtesy of Joseph Muenzer, MD, PhD
Disease Progression: MPS I Severe
10 months
12 months
34 months
22 months
39 months
Courtesy of Joseph Muenzer, MD, PhD
MPS I Genotype – Phenotype Correlations
• Nonsense mutations – assume total lack of enzyme activity
• Homozygous or compound heterozygous for nonsense
mutations = severe disease
• 15 known nonsense mutations with W402X and Q70X, the two
most common
• Greater than 70% of MPS I patients have at least one nonsense
mutation
• Majority of patients with one missense mutation have
attenuated disease – R89Q most common
• Pseudodeficiency alleles have been reported
Courtesy of Joseph Muenzer, MD, PhD
MPS I Treatment
• Early hematopoietic stem cell
transplant (before 2 years of age) can
prevent cognitive decline, heart
disease, hepatosplenomegaly,
coarsening of features, hearing loss in
severe form
• Peripheral IV enzyme replacement
therapy effective for some somatic
complications (joint stiffening,
hepatosplenomegaly, cardiac disease)
but not neurological problems
http://thethomasfamilytrials.blogspot.com/2012/02/one
-year-post-transplant.html
IDUA Enzyme Activity Assay to Screen for MPS I
• Screening 80,000 newborns using dried blood spot specimens
• Testing done at the North Carolina State Laboratory of Public Health
• Initial enzyme activity assay using PerkinElmer reagents for the IDUA enzyme only.
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Dried blood spot sample is incubated with IDUA synthetic substrate and internal standard
In normal DBS samples, IDUA cleaves the substrate to produce a product
Product is detected using tandem mass spectrometry
Enzyme activity is calculated (µmol/L/h)
Normal newborns have high enzyme activity and screen positive infants have no or low enzyme activity
• Low activity samples are retested with a 3-Plex assay
• PerkinElmer reagents are used to test 3 LSD enzymes (IDUA, GAA and ABG)
• GAA and ABG enzymes are used as references to assess sample quality
• Molecular testing will be done on screen positive samples that have very low enzyme activity
• Sequencing of IDUA gene
• Sequencing done at Duke Molecular Diagnostics Laboratory
Assay Implementation
• Hired Lab personnel:
• IT programmer
• Chemist
• Med tech
• Instrumentation:
• Using existing mass spectrometer: 2 Waters Acquity TQD
• Purchases shaking incubators and other general lab equipment
Assay Validation
Clinical Validation
• Analysis of 2,148 specimens
• Quality Controls
• Positive and negative controls provided
by CDC (Low, Med High, Base Pool)
• Daily QC purchased from PerkinElmer
• Proficiency Testing panel from CDC
• Specimen exchange with Missouri
newborn screening program
(normal, carrier, pseudo-deficiency,
severe MPS I)
Normal Birth Weight (above 2500g)
N = 1866
Mean
Median
Max
Min
St. Dev
IDUA Activity
5.619
5.292
18.469
0.503
1.932
10
Population Median
5
Referral Cutoff
-1
C
D
C
M
P
S
-1
S
M
P
ir
m
ed
ie
nc
y
fic
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on
f
P
se
ud
od
e
C
ar
ri
e
r
0
N
or
m
al
IDUA Activity (mol/L/hr)
Activity of Screen Negative and
Screen Positive Samples
Courtesy of Jennifer Taylor, PhD
Screening Algorithm
2 other enzymes are in
normal range
With 3-Plex Reagents
Inconclusive
Two of the three enzymes have
low activity
Cutoff
Value
IDUA Activity
(µmol/L/h)
Initial
2.27
Referral
1.35
Work Flow for Positive MPS-I Newborn Screens
Abnormal Screen Result
NC State NBS Lab
Communicate by Phone Call or
Fax
Genetic Counselor UNC Division of
Pediatric Genetics and Metabolism
Phone Call
Infant’s Primary Care Provider
PCP contacts family directly or has
GC contact family and
appointment made to see infant
and parents at UNC-CH
Work Flow for Positive MPS-I
Newborn Screens
Family comes to UNC for confirmatory
testing
urine sample for glycosaminoglycan (GAG)
measurement , blood sample for enzyme
testing
Negative urine
GAG, normal
enzyme level =
false positive
screen
No follow-up needed
Negative urine
GAG,
decreased
enzyme, IDUA
pseudodeficiency
mutation
Likely pseudodeficiency
12 month follow-up at
UNC
MD Metabolic/Clinical Geneticist
and Genetic Counselor see
parents/patient. Blood & urine
samples sent to Duke. Results
communicated by Duke Lab to
UNC GC
Positive urine GAG and decreased or no
enzyme
Confirmed MPS I
NO
Two IDUA nonsense mutations
(associated with severe form)
q 3 month follow-up at UNC to evaluate for
somatic and CNS disease and determine
need for ERT and/or HSCT
YES
Referral to Duke for
hematopoietic
stem cell transplant
(HSCT)
MPS I Newborn Screening Pilot Education
• Presentation at state meeting of NC
Pediatric Society
• Website for providers and parents
developed by RTI through grant from
National MPS Society
• http://ncnewbornscreeningpilots.org/MP
S-I/index.php
• Webinar given for all primary care
providers in state and posted on
webiste
• Memo sent to all birthing hospitals,
physician offices and health
departments in state by director of
NCSLPH
MPS I Newborn Screening Pilot
• Screening began August 15, 2016
• Laboratory and clinical data collected will be shared with the LPDR
using common data elements developed by NBSTRN
Acknowledgements
Leadership Team for MPS I Pilot
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Don Bailey, PhD – RTI International
Jennifer Taylor, PhD – RTI International
Lisa Gehtland, MD – RTI International
Joseph Muenzer, MD, PhD – UNC-Chapel Hill
Alex Kemper, MD, MS, MPH – Duke University
Scott Zimmerman, DrPH, MPH, HCLD (ABB) – Director, NC State Lab of
Public Health
Hari Patel, MS – NC State Newborn Screening Lab
David Millington, PhD - Duke University (retired)
Kristin Clinard, MS – UNC-Chapel Hill
Cynthia Powell, MD – UNC-Chapel Hill
Acknowledgements
MPS I Newborn Screening External
Advisory Committee
• David Millington, PhD
• John Rusher, MD
• Barbara Wedehase, MSW
• Adam Zolotor, MD, MPH
• Mark Dant
Laboratory Directors - Duke
• Deeksha Bali, PhD
• Sarah Young, PhD
• Rebeccah Catherine Rehder, PhD
Newborn Screening Program
• Joe Orsini PhD (NY NBS Program)
• Tracy Klug, MO NBS Laboratory
• Patrick Hopkins, Chief of MO NBS
laboratory