Newborn Screening 50th Year Celebration: the Utah experience

Marzia Pasquali PhD FACMG

Professor of Pathology and Pediatrics Medical Director, Biochemical Genetics and Newborn Screening

University of Utah and ARUP Laboratories

17 October 2013

Newborn screening

Public health activity aimed at the early identification of

conditions for which timely intervention can lead to the

elimination or reduction of mortality, morbidity, and

disabilities associated with these conditions.

History of Newborn Screening

1961 Dr. Robert Guthrie (New York) • First to use dried blood spot sample (often referred to

as a Guthrie card)

• Developed Bacterial Inhibition Assay (BIA) to detect

elevated levels of phenylalanine in dried blood spots

• Blood spots are placed on agar media containing B.

subtilis spores and a chemical inhibitor

• Spores can not grow unless phenylalanine is present

• Measure diameter of bacterial growth around blood

spot and compare to growth around blood spots

enriched with known amounts of phenylalanine (semi-

quantitative)

Newborn Screening: then

One analyte

(Phe)

One test

(Bacterial Inhibition Assay)

One disease

(Phenylketonuria)

One spot

Tandem Mass Spectrometry

• MS/MS can test

a number of

metabolites

simultaneously,

rather than one

analyte at a time

as in classic

screening

procedures.

Newborn Screening: now

Multiple analytes

Many diseases

487.47 1485440

459.40 1444288

437.33 756800 288.29

745216 403.21 702080

375.34 479104 347.35

450416 313.37 426912

298.43 346880

319.24 403520

347.04 311184

347.48 392144

347.86 105628

400.44 170080

376.23 123656

426.23 209408 404.10

161568

437.58 666432

438.40 253632

454.42 134736

460.22 481056

482.55 268048

461.36 111876

488.41 552960

577.46 166384

489.48 165424

549.59 108612 513.07

90616 523.35 50356

One test

(MS/MS)

One spot

MS/MS analysis Two main classes of metabolites can be detected using

tandem mass spectrometry (MS/MS):

– Amino acids. The level of one

or more amino acids increases in disorders of amino acid metabolism (metabolic block close to the actual amino acid).

– Aminoacidopathies and Urea Cycle defects: • Phenylketonuria, Maple

Syrup Urine Disease, Homocystinuria, Citrullinemia, Argininosuccinic aciduria, Tyrosinemia Type I

– Acylcarnitines. In disorders of the intermediary metabolism of amino acids or of fatty acid oxidation the abnormal metabolites are conjugated with carnitine to facilitate their excretion and to balance the Coenzyme A pool.

– Organic acidemias and fatty acid oxidation defects: • Glutaric acidemia Type I,

Propionic acidemia, Methylmalonic acidemia

• MCADD, VLCADD, CUD, CPT-I and CPT-II deficiencies.

Secretary Advisory Committee on Heritable

Disorders and Genetic Diseases in Newborns and

Children TANDEM MASS SPECTROMETRY

(MS/MS)

Traditional methods

(EIA, HPLC,etc.)

OTHERS

Acylcarnitines Amino acids

FAO (5) OA (9) AA (6) Hematology (3) Others (5)

MCAD IVA PKU Hb SS CH SCID

VLCAD GA-1 MSUD Hb S/bThal BIOT HEAR

LCHAD HMG HCY Hb S/C CAH CCHD

Critical congenital

heart disease

TFP MCD TYR I GALT Pompe

CUD MUT ASA CF

Cbl A,B CIT

3MCC

PROP

BKT

Newborn Screening in Utah

(2006)

• PKU

• Congenital Hypothyroidism

• Galactosemia

• Hemoglobinopathies

• Newborn Hearing

Screening

•MCAD Deficiency

•MSUD

•Homocystinuria

•Biotinidase deficiency

•Congenital adrenal

hyperplasia

•Others detectable by MS/MS

•Cystic Fibrosis (2009)

•SCID (2013)

•CCHD (2014)

Utah’s unique partnership

• A Pilot Project (2003-2005) was

established with the purpose of:

Evaluating the feasibility of expanding

newborn screening in Utah with a

private-public partnership involving the

Department of Health, ARUP

Biochemical Genetics laboratory, and

the Metabolic Center/Medical Genetics

at the University of Utah.

Pilot Project: Study Design (1)

• This study was approved by the Institutional

Review Board of the University of Utah (IRB_00011072, Supplemental Newborn Screening for the Early Diagnosis of Inborn Errors of Metabolism in Utah).

• Facility: University of Utah hospital (3,000-4,000 births per year)

• Subjects: Infants born at the U of U hospital (neonatal intensive care unit babies were excluded). Mothers had to sign a consent form.

Filter paper

Pilot Project: Study Design (2)

MS/MS

DoH

Nu

mb

er

of

ba

bie

s

0

1000

2000

3000

Total births

Pilot project

Oct 03-Dec 03 Jan 04-Sep 04 Oct 03-Sep'04

30%

68%

91%

Pilot Project: Participation

Oct 03-Dec 03 Jan 04-Sept 04 Oct 05-Sept 05

• 4,827 first screen and 690 second

screen samples were analyzed

– 1.7% first screen and 0.6% second screen

needed follow-up

– 0.1% needed confirmatory testing

– 1 infant with Glutaric acidemia type I was

identified and treated

Pilot Project: Results

Pilot Project: Conclusions

• Integration of private and public resources allows the prompt identification and treatment of children with metabolic disorders.

• Bidirectional data exchange between the State and the testing laboratory is necessary for good turnaround time.

• Education of health care professional and of the community is of paramount importance for a successful implementation of a new program.

UTAH Newborn Screening Program

The efficiency and effectiveness of a newborn screening program is dependent upon the smooth integration of sample collection, laboratory testing, follow-up, diagnosis, timely

treatment, and tracking of outcomes.

Follow-up

Clinical care Education

Evaluation

Medical Home

MS/MS screening in Utah: 2006-2013

• Total screens: 416,557

– Total metabolic disorders: 188 1:2,216

– Total maternal cases: 24

• PKU/Hyperphe/Biopt

• MCAD deficiency

• 3-Methylcrotonylglycinuria

• VLCAD deficiency

• Glutaric acidemia type I

• Primary carnitine deficiency (CUD)

• LCHAD deficiency

Thank You

• ARUP

– Newborn Screening and Biochemical Genetics Laboratories

– Dr. N. Kusukawa

– Dr. C. Kjeldsberg

• Utah Department of Health

– Kim Hart, Dr. H. Randall, Dr. R. Atkinson, N. Brown

– Fay Keune, Dr. F. Tait

• University of Utah Metabolic Clinic

– Dr. N. Longo, Dr. L. Botto, Dr. A. Warnock, S. Ernst, Krista

Viau

SCID Newborn Screening in Utah

Patricia Slev, PhD, D(ABCC)

Medical Director, Serologic Hepatitis and Retrovirus Laboratory

Co-Director, Immunogenetics Laboratory

Assistant Professor of Pathology,

University of Utah, School of Medicine

Timeline

• 2010 Uniform Panel Recommendation

• 2011 Utah SCID Subcommittee

Parent Advocate, March of Dimes, Pediatric Immunology, Pediatric

Bone Marrow Transplant Division

• 2012 Utah Genetics Advisory Committee and Newborn Screening

Subcommittee recommend screening for SCID

• 2013 Governor approves kit fee increase

• 2013 Pilot

• 2013 Implementation (July)

Why Screen ?

• Fatal without treatment

• Asymptomatic at birth

• Effective Treatment

hematopoietic stem cell transplant (HSCT)

enzyme replacement

gene therapy

• Early intervention is key

<3 months = 95% survival

>3 months = 65% survival

March of Dimes

Severe Combined Immunodeficiency Genotypes

T-Cell Lymphopenia

• Marker/Target – T-cell Receptor Excision Circle (TREC)

Circular, episomal, DNA fragment formed as a byproduct of successful

T-cell receptor rearrangement, during maturation in the thymus

TREC do not replicate with mitosis and therefore are diluted with

cell division

Peripheral concentrations correlate with T-cell production in the

thymus

Marker for T- cell lymphopenia not SCID

• Assay

Real-time PCR

Reported cut-offs and concentrations vary by laboratory due to

differences in reagents, calibrators and instrumentation

TREC Assay for SCID

Retrospective Utah Pilot Study

• 4,999 dried blood samples (DBS)

4,665 non- NICU

344 NICU

• TREC Singleplex Assay

TREC concentrations

b -actin concentrations

TREC Concentrations

Red - NBS samples

Green - CDC SCID negative screen/normal

Black - CDC SCID positive screen/SCID

Reference Gene Concentrations

(b -actin)

Red - NBS samples

Green - CDC SCID negative screen/normal

Black - CDC SCID positive screen/SCID

Pilot Study Results

0

100

200

300

400

500

600

Sa

mp

le n

um

bers

TREC concentration (copies/µl whole blood)

TREC concentration distribution in healthy and NICU babies

Healthy babies frequency

NICU babies frequency

TREC Concentrations vs. Birth Weight

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

0 1000 2000 3000 4000 5000 6000

Tre

c c

on

cen

trati

on

(co

pie

s/µ

L)

Birth weight (gr)

0

500

1000

1500

2000

2500

3000

3500

4000

4500

0 500 1000 1500 2000

TR

EC

co

ncen

trati

on

(c

op

ies/μL)

Birth weight <2000 gr

Birth weight (gr)

Detecting SCID

• Established Cutoff

• Detected CDC SCID positive DBS

• Detected SCID DBS

Reticular Dysgenesis

Adenosine Deaminase Deficiency

Algorithm

≤ cutoff

TREC & b -actin

(duplicate)

Non-NICU

low TREC/low b -actin – request 2nd DBS

low TREC/normal b -actin - request 2nd DBS

absent TREC/normal b -actin - flow cytometry

NICU

low TREC/low b -actin – request 2nd DBS

low TREC/normal b -actin –request 2nd DBS

TREC

> cutoff

screen negative

> cutoff

screen negative

The First Quarter

• Screened 13,200 babies

• NICU population accounts for the

majority of repeat DBS testing

• Confirmatory Testing – 3 babies

1 mild lymphopenia

2 profound lymphopenia

Acknowledgements

• Utah Department of Health

Dr. Harper Randall, Kim Hart , Dr. Robin Atkinson, Norm Brown

• Dr. Mei Baker

• ARUP/University of Utah

– Dr. Marzia Pasquali, Dr. Noriko Kusukawa, Dr. Harry Hill, Dr. Carl Wittwer

– New Technology Group, Newborn Screening Lab, Molecular Genetics Lab

– Wei Xiong, Jorja Warren, Dr. Orly Ardon, Dr. Jeff Stevenson

• Primary Children’s Medical Center, Immunology Division

Dr. Karin Chen

• SCID Subcommittee

Dr.Rich Harward (UDOH), Julie Drake (March of Dimes), Michael Pulsipher (Bone Marrow Transplant) Nan Streeter (UDOH), Jill Heaps (Parent Advocate)