bridge 2010, bristol, may 13-14, 2010 identification of ssmc – case 1. 47,xy, +...

Bridge 2010, Bristol, May 13-14, 2010

Identification of sSMC – case 1.

47,XY, + mar(30%)/46,XY(70%) 47,XY,+der(8)

Clinical symptoms

- behaviour disturbances - mild mental disability (IQ:67) - long philtrum - V. finger clinodactyly - dysgenesis of the corpus callosum

With parental consent

To identify/clarify mosaicism uncertain cytogenetic changes microdeletions sSMC subtelomeric rearrangements


• Subtelomeric regions are rich in genes severe dysmorphic symptoms• 50% of cases proved to be inherited (balanced carrier parent)• 5-10% of cases with idiopathic severe mental disability (deletion, duplication, translocation)• Some of them are associated with specific phenotype (1p, 4p, 5p, 9q, 17p, 22q• Probes: Vysis ToTelVysion Probe Panel: mixture of 15 individual „ready-to-use” direct labelled probes specific for telomeric regions of the p and q arms of all chromosomes (excepted the p arms of acrocentric chromosomes)

To identify/clarify mosaicism uncertain cytogenetic changes microdeletions sSMC subtelomeric rearrangements

Detection of subtelomeric rearrangements.

FISH: ToTelVysion Multi-color DNA probe Mixture

Bridge 2010, Bristol, May 13-14, 2010With parental consent.

Clinical symptoms- Hypotonia- Eye: Retinitis pigmentosa, decoloration of papilla, nystagmus, div. strabism- Polydactyly- Facial dysmorphy: depressed premaxillary region, hypertelorism, ptosis, small, low set ears, downward slanting mouth - Severe mental disability- Delayed psychomotoric development

Detection of subtelomeric rearrangement – cases 1.-2.

8p subtelomeric deletion; 12p subtelomericregion trisomy in unbalanced translocation


Array Comparative genomic hybridisation (CGH)

Advantages • to identify the exact breakpoints, localization of

affected genes • to detect microdeletions, microduplications,

polymorphisms of pathological importance • to study genotype – phenotype correlations


Specific dystrophin gene array analysis

Sample 3: 114/1 affected boy at contiguous gene deletion syndrome (Score

dystrophin gene: -0,536; Score neighbouring genes: -0,742)

Complex glycerol kinase deficiency (CGKD): microdeletion of Xp21


Complex glycerol kinase deficiency (CGKD): Xp21 microdeletion

• dystrophin gene deletion by MLPA

Neighbouring genes: • GK gene: Xp21.1 – glycerol kinase deficiency • NR0B1 gene: Xp21.3-21.2 – adrenal hypoplasia • Il1RAP gene: Xp22.1-21.3 – mental disability• RPGR gene:

Accurate breakpoints Genotype-phenotype correlation


Genetic diagnostic approaches

Chromosome aberrationis compatible with the clinical features

Further steps delineated by the phenotype

Fine chromosome alterations: markers’ identification, sSMC, microdeletion syndromes, subtelomeric rearrangements

FISH, MFISH, CGHMolecular genetic tetsts(mutation analysis: PCR, sequencing, RFLP etc.)

Syndromes ofunknown origin

Syndromeidentification:„search” >5600

Chromosome analysisChromosome analysis

Numerical, structuralaberrations

Uncertain Normal karyotype

Known monogenic syndromes: Fra-X, chondrodysplasia, syndromic craniosynostoses etc.

Known chromosomal syndromes, syndromes of unknown origin, mental disability, dysmorphic symptoms

3.


Molecular diagnosis of monogenic disorders

Diseases studied:• Fragile–X: FMR1 gene – Southern blot • Craniosynostoses: Apert, Crouzon, Pfeiffer

FGFR2 gene – PCR, sequencing, RFLP

• Achondroplasia: FGFR 3 gene - RFLP Hypochondroplasia: FGFR 2, 3 genes –

sequencing • Lissencephaly: LIS1 gene – FISH, sequencing


L TM TK1 TK2AIgI IgIIIIgII

Ser252Trp Pro253Arg: 99%

Apert syndrome: AD – FGFR2 mutation

Clinical symptomsCraniofacial dysmorphy: clover shaped cranium, high forehead,flat occiput, hypertelorism, deep nasal root, small, beaked nose, cleft palateSyndactyly, small muscular VSD



Patient

Healthy

172153118

54

300250

200150

100

50

E B M

c.C755G ( Ser252Trp) mutation

Genetic study

DNA isolationAmplification of exon 8 in FGFR2 gene using PCR techniqueSequencing, RFLP analysis (MboI)


Apert syndrome 2.

FGFR2c.C758G (p.Pro253Arg) Gain of function:

Increased ligand-binding capacityLoss of ligand-binding specificity

Genotype-phenotype correlation: Ser252Trp: cleft palate more common

better recovery after operationPro253Arg: more severe syndactyly



achondroplasia Apert sy DMP Rett sy FGFR2 FGFR3 dystrophin MECP2

PKU, CF, CAH, Werdnig-Hoffmann, Smith-Lemli-Opitz, glycosilation disturbances, haemophilia, Russel-Silver, Lysosomal storage diseases (Gaucher, Fabry, MPS I. Niemann-Pick, Pompe) , etc.

ORPHANET

Significance: • to avoid unncessary investigations• prognosis,• praenatal diagnosis • praesymptomatic diagnosis• prevent recurrence

Further genetic tests to detect gene mutationsperformed in collaboration inside Hungary


Genetic diagnostic approaches

Chromosome aberrationis compatible with the

clinical picture

Further steps delineated by the phenotype

Fine chromosome alterations: markers’ identification, sSMC, microdeletion syndromes, subtelomeric rearrengements

FISH, MFISH, CGHMolecular genetic tetsts(mutation analysis: PCR, sequencing etc.)

Syndromes ofunknown origin

Syndromeidentification:„search” >5600

Chromosome analysisChromosome analysis

Numerical, structuralaberrations

Uncertain Normal karyotype

Known monogenic syndromes: Fra-X, achondroplasia, Apert sy. etc.

Known chromosomal syndrome, syndromes of unknown origin, mental disability, dysmorphic symptoms

4.


• blepharophimosis• ptosis• external acustic meatus stenosis +/- deafness• hypotonia• mental disability

Diagnostic criteria:

Photos are shown with parental consent

Diagnosis: Ohdo syndrome


B=0 FA TRACE

Standard MRI vs. Neuroimaging:

T1 and T2 weighed MRI revealed no organic alteration in the brain, while fractional anisotropy and fiber trajectory imaging diclosed a demyelinated area close to the anterior horn of the ventricle causing a complete „communication” blockade

Magnetic resonance Imaging


CONCLUSION

• To identify the genetic background of congenital genetic disorders the whole spectrum of currently available genetic tests have to be applied according to a proper algorythm

• The genetic diagnosis (etiology) determines the syndrome-specific additional symptoms as well as pattern and severity of intellectual disability.

• The genetic diagnosis has to be considered when developing the early intervention, rehabilitation and preventive measurements.


Director of working group: Éva Oláh, MD, D.Sc.

Members:

Cytogenetics: Erzsébet Balogh, biol. PhD.

FISH, PCR: Anikó Ujfalusi, MD. PhD Beáta Bessenyei, mol. biol.

Syndrome diagnosis: Katalin Szakszon, MD. Gabriella P. Szabo, MD.

Technical assistants: Mrs. Gábor Orvos

Mrs. Ferenc BodnárMs. Éva Nagy

PhD students: Attila Mohánszky, mol. biol.

Gabriella P. Szabó, MD. Katalin Szakszon, MD. Ivett Körhegyi, MD.

: Student Scientific Circle: students

Clinical Genetic Center, University of Debrecen

bridge 2010, bristol, may 13-14, 2010 identification of ssmc – case 1. 47,xy, +...

Documents

p subtelomeric deletion

subtelomeric regions

p subtelomeric region

microdeletion of xp21

gk gene

mental disability rpgr

specific phenotype

unbalanced translocation