Genetic Testing and Care for Newborns in NICU (neonatal

intensive care unit)Seema R. Lalani, MDAssociate Professor

Department of Molecular and Human GeneticsBaylor College of Medicine

Houston, TX 77030

Learning objectives

• Describe the genetic evaluation for a critically ill newborn• Discuss a case study

Burden of genetic disorders in NICUs• Congenital anomalies are a leading cause of infant mortality in the United States, accounting for about

20% of all infant deaths

• Chromosomal abnormalities are ascertained in one in 160 liveborn infants

• There are over 4500 Mendelian disorders that have a known genetic etiology at present, and a significant fraction of these present in the neonatal period

• Genetic assessment has become an essential aspect of neonatal medicine

• Healthcare professionals need to know when genetic evaluation is indispensable in the care of newborns

Why is it important to make a genetic diagnosis?

• Difficult to decide on plan of care without a certain diagnosis

• Obtaining a genetic diagnosis often assists in patient management

• Families dealing with serious medical issues in their children often endure years of diagnostic uncertainty

• Prevention of genetic disease in future children

Some reasons to consider genetic testing in the NICU• Multiple congenital anomalies • Neonatal hypotonia, feeding difficulties

• Neonatal seizures• Congenital heart disease • Cardiomyopathy

• Brain malformations• Skeletal dysplasia• Arthrogryposis

• Liver failure• Immunodeficiency• Lactic acidosis

http://www.chron.com/neighborhood/humble-news/article/Hospital-sees-triplet-births-three-times-in-half-1723835.php

Challenges of making a genetic diagnosis in critical care setting

Infants on life support- comprehensive clinical assessment can be difficult

Atypical presentation of a typical genetic disorder in the newborn period

Sequential testing may be time consuming in a critically ill child

Genetic evaluation of a newborn

Rudolph Pediatrics

Clinical tests that one can request • Karyotype• Aneuploidy FISH• CMA Comprehensive• Metabolic testing- Plasma amino acids, urine organic acids, acylcarnitine

profile, lactate, and ammonia • Global MAPS• MitoNGS• Methylation study• Single gene testing (CFTR, SMA, CHD7)• Whole exome sequencing (WES)• Critical WES

46,XX

CellNucleus

Chromosomes

Gene

Protein

~ 25,000 genes—packaged on 23 pairs of chromosomes

Chromosomes and genes

KaryotypeCytogenetic

abnormalities

DNA sequencing

Genomic disorders

Chromosomal microarray

analysis (CMA)

Chromosomal aberrations

Trisomy 21, 18, 13

Single gene disorders

Cystic fibrosis

Chromosomal abnormalities frequently cause multiple congenital anomalies

• Chromosomal abnormalities are seen in one in 160 liveborn infants

• Of these, the most common abnormality is Down syndrome (trisomy 21), seen in approximately one in 800 livebirths

Down syndrome• Congenital heart defects are seen in ~50% of children

with Down syndrome

• Most common genetic cause of intellectual disability

• Hypotonia

• Epicanthal folds, flat nasal bridge, upward-slanting palpebral fissures, small ears

• Midphalangeal hypoplasia of 5th digits, wide spacing between the first and second toes, single transverse palmar crease

• The risk of having a pregnancy affected by Down syndrome is highly correlated with maternal age, increasing from about 0.1% at 30, to 1% at 40

Karyotype should be done in all infants suspected to

have Down syndrome

Down syndrome-trisomy 21

Trisomy 18

• 1 in 6,000 live births • ~95% spontaneously abort• Growth retardation, small eyes,

small mouth and chin, overlapping fingers, rocker-bottom feet

• Multiple heart valves are affected

>50% of infants die in the first few weeks of life

Trisomy 18 (47,XY,+18)

Rapid FISH

Trisomy 13

Clefting abnormalities, scalp defects, extra digits, brain abnormality

>80% of infants die in the first month of life

Trisomy 13 (47,XX,+13)

Turner syndrome, 45,X

Doswell et al, 2006

Karyotype is important but many chromosomal deletions and

duplications are too small to be detected on chromosomal study

Chromosomal Microarray Analysis (CMA) checks for

smaller deletions/duplications in the genome

An entire gene is missing, along with other genes

Image taken from https://deescribewriting.wordpress.com/category/point-of-view-2/

Normal Chromosomal Microarray Analysis (CMA)

DiGeorge Syndrome

DiGeorge syndrome is a frequent cause of congenital heart defects in children

Chromosome 22q11 deletion

A baby with congenital heart defect (tetralogy of Fallot)Low calcium level in bloodOn examination, has small ears, cleft palate, and long digits

Hypotonic infant

Neonatal hypotonia and feeding difficultiesPrader-Willi syndrome (methylation study)

Wang et al. BMC Pediatr. 2016

Angelman/Prader-Willi deletion on chromosome 15

Consensus StatementThe diagnostic yield of CMA is significantly better (15-20%) than G-banded karyotype study in the evaluation of children with multiple

congenital anomalies

Miller et al. AJHG, 2010

Baby with a large tongue and hypoglycemia

• Two-month old infant with a large tongue (macroglossia)• History of neonatal hypoglycemia• Large for age• Abdominal wall defects

Abnormal methylation testing for Beckwith-Wiedemann syndrome

Whole Exome Sequencing

The exome (“sum total” of exons) makes up about 1.5% of the whole genome, but is predicted to harbor 85% of disease-causing variants

• We have about 20,000 plus genes• Human genome has about 3 billion letters

CMA checks for Missing pagesDuplicated pagesMissing paragraphs on a page

EXOME CHECKS FORMisspellingsMissing lettersExtra letters

Diagnostic yield

• Yield of CMA in the evaluation of MCA-15-20%

• Exome sequencing has helped in determining a cause in about 25-30% of children suspected to have a genetic diagnosis

When do we order exome sequencing?

• No clinical diagnosis • Extensive differential diagnosis• All other previous testing has been non-diagnostic• Rapid testing is needed for determining patient’s diagnosis and prognosis

Retterer et al., Genet Med. 2016

Test yield based on primary indication

50-HOUR WGS

2012

Twenty neonates studied, 8 diagnosed (40%)

2016

TCH Study• Retrospective study included infants <100 days old who

had WES completed from 2011 to January 2017 through Baylor Genetics Laboratories

• WES was completed in 278 infants with suspected genetic diseases

• The majority (~90%) were from intensive care units (NICU, CVICU, and PICU)

Results

• Overall, molecular diagnosis was achieved in 102/278 infants (~36%)

• Critical exome (TAT of 2 weeks) provided a diagnosis in 32/63 infants (50.8%)

• The diagnosis affected medical management in 53/102 (52%) infants

Medical management Overall Rate(n=278)

Affected medical management? 53/102(52.0%)

Re-direction of care 19 (35.8%)

Initiation of subspecialist care 27 (50.9%)

Change in treatment or diet 7 (13.2%)

Major procedures completed 5 (9.4%)

Clinical exome sequencing has a high impact on medical management

Five infants (9%) received hematopoetic stem cell (HSCT) or organ transplantation

• RAG1 -Severe combined immunodeficiency• UNC13D-Hemaophagocytic lymphohistiocytosis• FANCA-Fanconi anemia

• PTPN11-Noonan syndrome (severe HCM)• ACTC1- Left ventricular non-compaction

Medical impact-dietary and and medication changes

Gene Disease Treatment

ATP7AMenkes disease; Occipital horn syndrome; Spinal muscular atrophy, distal, X-linked 3

Receiving copper histidine injections under NIH clinical trial

PDHA1Leigh syndrome, X-linked; Pyruvate dehydrogenase E1-alpha deficiency Thiamine and ketogenic diet

ACAD9Mitochondrial complex I deficiency due to ACAD9 deficiency

Followed by Cardiology for LV trabeculations; Riboflavin

MUT Methylmalonic aciduriaPro-Phree/Propimex-1 formula and carnitine

ETFDH Glutaric acidemia IIC Treated with Carbaglu (carglumic acid) and Riboflavin

ENPP1 arterial calcification of infancy, generalized, 1 (GACI1) Treated with pamidronate

ALDH7A1 Epilepsy, pyridoxine-dependentTreated with large dose of Vitamin B6 (pyridoxine) with cessation of seizures

Mortality Exome Sequencing Overall Rate(n=278)

120-day death rate Diagnosed 30/102(29.4%)

Undiagnosed 28/170(16.5%)

Higher mortality in infants who are diagnosed by exome sequencing

For all the deceased infants (81/278), genetic disorders were molecularly confirmed in 48% infants by exome sequencing

Genetic Diagnoses Genes Affected infants

Kabuki syndrome KMT2D 4 (5%)

Noonan syndrome PTPN11 2

Noonan syndrome RAF1 2

Orofaciodigital syndrome I OFD1 2Combined oxidative

phosphorylation deficiency 12EARS2 2

Glycogen storage disease IV GBE1 2

Profound neonatal hypotonia PURA 2

Liver failure, transient infantile TRMU 2

Short-rib thoracic dysplasia 3 with or without polydactyly

DYNC2H12

Recurring diagnoses made by WES in infants in critical care units

When to consider Noonan syndrome diagnosis in NICU?

• Infants with Noonan syndrome can present with hypertrophic cardiomyopathy

• Pulmonic stenosis• Dysmorphic features

https://www.heart.org/en/health-topics/congenital-heart-defects/about-congenital-heart-defects/pulmonary-valve-stenosishttps://www.mayoclinic.org/diseases-conditions/hypertrophic-cardiomyopathy/symptoms-causes/syc-20350198

Approximately 39 out of 102 (38%) diagnosed individuals had atypical or unrecognized infantile

presentation of genetic disorders

When to consider the diagnosis of Kabuki syndrome in NICU (KMT2D)?Not an easy clinical diagnosis in this group

Left ventricular outflow tract defects, hypotonia, renal defects, feeding difficulties, and ear abnormalities-Think Kabuki!

Complex phenotype• Infant-4 days old-NICU

• Small for gestational age• Bilateral microphthalmos, microcornea, small palpebral

fissures, and iris colobomas• Pulmonary hypoplasia • Atrial septal defect• Left sided multicystic dysplastic kidney (pelvic), right kidney

was small • Anal atresia• Ambiguous genitalia• Microcephaly; brain MRI: Marked dilatation of the lateral

ventricles, with associated thinning of the brain parenchyma, and small cerebellum and brainstem

Diagnosis?

• Two changes (p.Y1655X and p.R2336P) in the BRCA2 gene were detected in this individual.

• Defects in BRCA2 are the cause of Fanconi anemia complementation group D1 (FANCD1)

Conclusions

• CMA and whole exome sequencing are powerful diagnostic tools in the evaluation of critically ill infants suspected to have genetic disorders

• Neonates often present with atypical presentation of recognizable disorders making clinical diagnosis challenging

• Rapid genetic testing (critical exome sequencing) provides earlier diagnosis in infants in ICUs, affecting clinical decision-making in a significant number of infants

Thank you!