Learner outcomes and its application to speech, hearing, At the end of this session, the learner will and craniofacial disorders (2020) • Understand the significance of identifying a genetic diagnosis • Learn how a medical geneticist differentiates syndromic vs isolated Nathaniel H. Robin, MD • Know the benefits and limitations of current Department of Genetics and future genetic testing University of Alabama at Birmingham

Overview What’s new?

• Approach to the patient with craniofacial anomaly • Determine syndromic v. isolated • Examples – Cleft lip and/or palate – –

Dysmorphology vs. Genomic Medicine

1 Genomic Medicine ‘Traditional’ genetics “ … the routine use of genotypic analysis, usually in the form of DNA testing, to enhance the quality of Dysmorphology (the study of abnormal form) medical care.” - A. Beaudet, 1998 ASHG Presidential Address (AJHG 64:1-13 1999) • Evaluation of child (adult, fetus) with unusual Examples facial characteristics +/- other abnormal • Cancer genetics findings in an effort to reach a genetic • Neurogenetics () diagnosis • Pharmacogenetics • Hearing

Indications for genetics evaluation Why is it important to make a diagnosis?

• Multiple anomalies • Prognosis – Including intellectual disability and growth failure • Management • Single (recognized) anomaly with known • Recurrence risk counseling genetic contribution • Access support groups – Cleft lip, hearing loss, craniosynostosis • Treatment • Unusual facial appearance • ‘Why’ • “Positive” family history

How to identify a genetic syndrome • Major anomaly: has severe medical or cosmetic consequence – congenital heart defect, deafness, intellectual • Look for other problems* in patient and family disability, cleft lip/palate members • Minor anomaly: clinically insignificant • Both similar and seemingly unrelated departure from normal development – wide set eyes, single palmar creases, café au lait spot *both major and minor anomalies – vs. normal variant

2 Is this child normal?

“The best clues are the rarest… (T)hese are not the most obvious anomalies nor even the ones that have the greatest significance for the patient’s health.” …. John Aase, M.D.

Cleft lip and/or cleft palate

• Oro-facial clefting is most common in the USA: 1/600 – Sharp rise in last 10 years (why?) • Requires extensive multidisciplinary team evaluation & management – Genetics is an important team member • >300 genetic with CLP • Isolated CLP is also genetic (multifactorial)

3 Cleft lip with/without cleft palate is NOT the same as cleft palate alone

• CLP: failure of fusion of premaxillary process with lateral maxillary procese(s) at ~42 days gestation

• CPA: Failure of fusion of palantine ridges to fuse at ~63 days gestation

CL/P vs CPA: epidemiology Mixed clefting • Occurrence of cleft lip/palate and cleft Cleft lip/palate Cleft palate alone palate alone in same family • Incidence/prevalence • No racial variation • Causes varies by race • Male/ female: 1/2 – Chance • Male/ female: 2/1 • Assoc’d malf’s: 22-61% – Van der Woude syndrome CL/P LP • Ass’d malf’s: 8-28% • Syndromic: 25-40% – Popliteal Pterygia • Syndromic: 10-25% – EEC CL/P CPA, CLA • Recurrence risk counseling for isolated CL/P vs.CPA – Rapp-Hodgkin LP is also different; based on empiric data – Hay-Wells

Van der Woude syndrome Importance of the premaxilla

• Cleft lip, cleft palate, lip pits • Genetics: AD, variable • Gene - Interferon regulatory factor 6 (1q32) – VDWS allelic to PPS – microdeletions: VDWS plus MR – IRF6 involved in NS CLP

4 Pierre Robin sequence (Hereditary arthroophthalmopathy) • Primary defect: micrognathia • Other findings: “U-shaped*” • Major clinical manifestations: cleft palate, glossoptosis – Myopia, retinal changes • 50% syndromic – Early/progressive arthritis , mild SED • Stickler syndrome – Sensorineural hearing loss • 22q11.2DS (VCFS) – Cleft palate/Pierre Robin sequence – Genes: COL2A1, COL11A1, COL11A2 (AD); COL9A1, COL9A2, COL9A3 (AR)

22q11.2 deletion syndrome (22q11.2DS) Treacher Collins syndrome

• Common (1/2-4000) & very variable (many names) • Autosomal dominant – – Velocardiofacial, DiGeorge, CTAF, Cayler, Kousoff… 60% sporadic – Also isolated conotruncal CHD, neonatal hypocalcemia • Very variable rd • Main clinic manifestations • Absence lateral 1/3 lower eyelashes – Characteristic facial appearance • Gene: TREACLE, on 5q32 – Congenital cardiovascular disease • Nager syndrome: TCOS with thumb – Speech, cognitive delays (VPI) anomalies – Psychological and behavioral problems

Genetic testing would provide an Nonsyndromic clefting genes added level of insight • ‘Familial’ nature of NS CLP has been recognized for a long time (Trew, 1757) – Genetic contribution 12-20% (1920’-40’s), now as much as 50% • ‘Multifactorial trait’ threshold model – Mix of genetic & environmental factors • 2020: no useful genetic test* – No predicative help – Only empiric recurrence risk counseling

5 Isolated Clefting NS CLP: more than meets the eye

NS CLP is a systemic disorder of development • Other findings – Developmental asymmetry – Non right-handedness – Abnormal craniofacial anatomy – Hypoplasia orbicularis orbis – Dental anomalies – Velopharyngeal insufficiency – Suicide – Cancer

Deafness/hearing loss Environmental •ototoxic drugs •acoustic trauma •infections Syndromic • Clinically significant hearing loss (>25dB) affects •Alport several million children (~1/500 newborns) ~50% •Pendred • Early identification is key ~30% •Waardenburg Deafness •Usher • Universal newborn screening Genetic ~50% • Genetic advances occurring rapidly Non-syndromic – 1993: 7 known genes (syndromic and nonsyndromic) ~70% ~22% AD (DFNA1-40) ~77% AR (DFNB1-30) – 2020: 90+ nonsyndromic hearing loss genes identified ~1% X-linked (DFN1-8) – Diagnostic AND therapeutic implications ~1% Mitochondrial

Multifactorial Environmental Multifactorial •ototoxic drugs •ototoxic drugs •acoustic trauma •acoustic trauma •infections Syndromic •infections Syndromic •Alport •Alport ~50% •Pendred ~50% •Pendred ~30% •Waardenburg ~30% •Waardenburg Deafness •Usher Deafness •Usher Genetic Purely Genetic ~50% ~50% Non-syndromic Non-syndromic Purely Genetic ~70% ~22% AD (DFNA1-40) ~70% ~22% AD (DFNA1-40) ~77% AR (DFNB1-30) ~77% AR (DFNB1-30) ~1% X-linked (DFN1-8) ~1% X-linked (DFN1-8) ~1% Mitochondrial ~1% Mitochondrial

6 Evaluation of the child with hearing Syndromic hearing loss loss • There are several hundred genetic syndromes with hearing loss as one finding • Type: Conductive, sensorineural, mixed – 30% prelingual hearing loss • Severity: mild-profound – all types (conductive, SN, mixed) • Age of onset: prelingual, progressive, adult- onset • Syndromic or isolated

Syndromes with hearing loss

• Abruzzo-Erickson • Fountain • acroosteolysis • frontometaphyseal dys •Nager • Achondroplasia • glactosialidosis •oculo-auticulo-veterbral spectrum • Adelaide/FGFR3 • gengival fibromatosis •oral-facial-digital II Some syndromes are obvious…. • Apert • Hersh •osteogenesis imperfecta I-IV • auralcephalosyndactyly • hyperphosphatasemia •osteopathis striata • auro-digital-anal • •osteopetrosis • Boston-type craniosynostosis -microtia-clefting •otodental • branchio-oto-renal • hypodontia •otopalatodigital I & II • campomelic dysplasia • Johanson-Blizzard •Pallister-Killian • Camurati-Engelmann • Kartagener • cat eye • Keipert •postaxial acrofacial dysostosis • cervico-oculo-acoustic CHARGE • Keutel •premature aging and multiple nevi •sclerosteosis • cleidocranial dysplasia • KID •Stickler • Cockayne • Klippel-Feil •symphalangism- • craniodiaphyseal dys • Kniest •Townes-Brocks • craniometaphyseal dys • LADD •Van Buchem disease • Crouzon • LEOPARDmandibulofacial dysotosis •velocardiofacial • EEC •Waardenburg • mannosidosis • facio-auriculo-radial dys •Wildervanck • focal dermal hypoplasia • Marshall-Stickler syndrome • MPS I, II, IV, VI

Other syndromes are more subtle… Cleidocranial dysplasia

Crouzon Syndrome

Oculoauriculovertebral spectrum

7 Waardenburg Syndrome

• Type I – White forelock – Dystopia canthorum – Iris heterochromia – PAX3 • Type II – Lacking telecanthus – MITF • Type III – Like type I with upper limb deficiency and NTD • Type IV (Waardenburg-Shah) Velocardiofacial Syndrome Stickler Syndrome – Plus Hirschprung disease (Del22q11.2) – EDNRB

Branchio-Oto-Renal syndrome Pendred syndrome

• Hearing loss (89%) • Genetics: AD, variable • Normal facial appearance • Preauricular pits (77%) expressivity; gene: EYA1 • Sensorineural hearing impairment, abnormality of (8q13.3) • Branchial fistula/cysts (63%) bony labyrinth (req temporal bone CT), and • • Renal dysplasia (66%) Abnormal pinnae (41%) • Malformed middle/inner ear euthyroid goiter • Lacrimal duct stenosis – Vestibular function is abnormal in majority – Goiter not present at birth, develops in early puberty (40%) or adulthood (60%). • Genetics: AR, SLC26A4 gene (7q22-q31) Ear pits Renal failure, – Detected in ~ 75% of patients on dialysis – in SLC26A4 also cause isolated EVA (enlarged vestibular aqueduct) Stillbirth SNHL, ear pits

Multifactorial Autosomal dominant inheritance •ototoxic drugs •acoustic trauma •infections Syndromic •Alport ~50% •Pendred A a a a ~30% •Waardenburg Deafness •Usher Purely Genetic A a ~50% a a Non-syndromic ~70% ~22% AD (DFNA1-40) ~77% AR (DFNB1-30) ~1% X-linked (DFN1-8) a a a a A a A a ~1% Mitochondrial A a

8 Autosomal recessive inheritance Empiric recurrence risk for profound childhood deafness of unknown cause

A a • 1 affected child 1/6 • 1 child, consanguinity 1/4 A a A a AA Aa • 1 child, 2+ normal sibs 1/10 A normal, normal normal, carrier • 2 affected 1/4 aA aa • 1 parent, 1 child 1/2 a a A a A A A a a normal, carrier Affected, Homozygous • 1 parent 1/20 • parent + parent’s sibs 1/100 • sibs of parent, parent normal <1/100 Punnet square

GJb2 (Connexin 26) Genetic testing for SNHL (Jan 2012)

• First clinically relevant SNHL gene Prior to Jan 2012 – 30% singlet cases, 55% autosomal recessive • Many genes known, but no single gene worth – RR: positive test-1/4, negative 1/7, no test 1/6 testing (except GJb2) unless suggestive findings – Provides valuable prognostic information too present (eg, auditory neuropathy and OTOF) • Ethnic variation • – Common in Caucasians (~3% carriers; 35delG), Ashkenazi Sequential testing low yield, very $, slow Jews (>6%; 167delT), every population studied except Africans Jan 2012 • OTOSCOPE

What is OtoSCOPE®? Still not perfect

• Platform using the newest DNA sequencing • NextGen testing resulted in identification of the underlying genetic cause for hearing loss in 440/1119 patients (39 %). methods (“massively parallel”) – 49 genes, all types of variants [missense (49 %), large & small CNVs (18 % • Instead of analyzing one gene at a time, will each), nonsense (8 %), splice-site (6 %), promoter (<1 %)]

simultaneously sequence 66 96 105 122 136* • The diagnostic rate highest for patients with a positive family genes – all known to cause non-syndromic history of hearing loss or when the loss was congenital and symmetric. sensorineural hearing impairment • Diagnostic sensitivity and specificity of >99%

Sloan-Heggen et al., Hum Genet (2016) 135:441–450

9 What will better testing mean?

• Improved recurrence risk counseling • Gene specific therapies • Molecularly targeted therapies, ?cures

• Non syndromic mimics

Nonsyndromic mimics

• Conditions that present as isolated hearing loss, but the hearing loss is actually the presenting finding for a more complex disorder – Usher – Defness-Infertility

Usher syndrome Impact of early diagnosis of Usher syndrome • Genetically heterogenous disorder characterized by congenital SNHL & progressive vision loss (retinitis Wright et al., 2020* pigmentosa). • No differences in emotions, suggesting earlier – Vision loss late childhood - adolescence. diagnosis does not lead to increased anxiety or • Historically, Usher syndrome has been diagnosed via psychosocial issues for parents. onset of vision loss • Earlier diagnosis allows parents to process and • NextGen testing -> earlier diagnosis, before onset of prepare the child for independence throughout life. vision loss (psychosocial impact? )

*Wright et al., (2020). Next-generation Sequencing Experience: Impact of Early Diagnosis of Usher Syndrome. Chpt 2 in A Comprehensive Guide to Genetic Counseling, Benjamin A. Kepert (Editor). Nova Science.

10 Other ethical issues What is next beyond “NextGen”?

Attitudes towards genetics • Genome/exome sequencing • Among Deaf (Middleton et al, ’98): very negative – vs NextGen, chromosomal microarray • Among deaf (Brunger et al, ’00): much more positive

• Among Deaf (Mascia et al., 2020): – Increased interest in having genetic testing (45%), genetic counseling (23%), prenatal testing (45% v 16%) – Negative attitudes towards genetics persist, but less

Extra concerns with genome Conclusions sequencing • Cost 1. Genetic testing is here, and only growing in • Timing scope and significance • VOUS • Incidental findings 2. Genetic testing should only be done in • Secondary findings conjunction with genetic counseling

3. We are here to help: 205.934.4983

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