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Next generation sequencing technologies (NGS) forwhole exome/genome sequencing (WES/WGS):

impact on daily clinical practice, in dementia and beyond

9ο Πανελλήνιο Διεπιστημονικό Συνέδριο Νόσου AlzheimerΘεσσαλονίκη 14-17/5/2015

Ζαγανάς ΙωάννηςΝευρολόγος,Επίκουρος ΚαθηγητήςΝευρολογίας

Βογιατζή Εμμανουέλλα,Βιολόγος-Γενετίστρια

Λατσούδη Λένα,Βιολόγος-Γενετίστρια

Introduction: Clinical whole exome sequencing as a tool to end the“diagnostic Odyssey”

Ζαγανάς ΙωάννηςΝευρολόγοςΕπίκουρος Καθηγητής Νευρολογίας,Τμήμα Ιατρικής Πανεπιστημίου Κρήτης /Πανεπιστημιακό Γενικό Νοσοκομείο Ηρακλείου


Lu et al, N Engl J Med, 2014

Η δική μας εμπειρία

Πρόγραμμα«Γενετικές αναλύσεις νέας γενιάς σε ασθενείς

με νευρολογικές και άλλες παθήσεις»

18-year old woman with limb-girdlemuscular dystrophy

• Her sister similarly affected, but not herparents

• Central-type muscle weakness, severelydepressed tendon reflexes, calf hypertrophy,scapular winging, waddling gait

• Subject to multiple tests, including musclebiopsy, with no diagnostic results obtained

Sarcolemma and proteins involved in muscular dystrophies (Mercuri and Muntoni, 2013)

ISPD: isoprenoid synthasedomain containing


ISPD

Diagnosedby us(UOC)


ISPD

Diagnosedby us(UOC)

2-year old girl with recurrent lacticacidosis, epilepsy

• Hypotonia• Seizures• Mental retardation• Microcephaly

Diagnosedby us(UOC)

2 y.o. f.,lacticacidosis,epilepsy

2 y.o. f.,lacticacidosis,epilepsy

13 y.o. f.,myalgias,↑ CPK

6 y.o. f.,myopathy

Diagnosedby us(UOC)

62-year old man with progressivelyworsening muscle weakness (A)

• About 15 years ago, weakness and muscularatrophy in the periphery of his extremities wasnoted (progressively worsening since then)

• Deep tendon reflexes abolished in all 4extremities

• loss of sensory function in a stocking-glovedistribution (pain, temperature, vibration)

• FH: Brother died at a young age from an, furtherun-specified, neurological disorder that led toquadriplegia

62-year old man with progressivelyworsening muscle weakness (B)

• Electrophysiology: sensorimotor demyelinating-typepolyneuropathy (later on with a patchy distribution)

• LP: moderately increased protein• Testing for PMP22 duplication: negative• Patient subjected to several tests (at various

centers) with no diagnostic results• Treated with immunotherapy with no improvement

Rossor et al,Nat Rev Neurol,

2013

Rossor et al, Nat Rev Neurol, 2013

Diagnosedby us(UOC)

Jean-Martin Charcot(1825 – 1893)

Pierre Marie(1853-1940)

Howard Henry Tooth(1856–1925)

The application of whole exome sequencing in dementia and beyond:the clinician’s viewpoint

Ζαγανάς ΙωάννηςΝευρολόγοςΕπίκουρος Καθηγητής Νευρολογίας,Τμήμα Ιατρικής Πανεπιστημίου Κρήτης /Πανεπιστημιακό Γενικό Νοσοκομείο Ηρακλείου


Γενετικοί παράγοντες συνδεόμενοι με βεβαιότητα με τη νόσο Alzheimer

Χρωμό-σωμα

Αριθμόςμεταλλάξεων(οικογένειες)

Ηλικίαέναρξης

(έτη)

% περιπτώσεωνπρώϊμηςέναρξης

%συνολικών

περιπτώσεων

APP 21 33 (90) 45-65 <1 <0.1

PS1 14 185 (405) 30-60 50 1-2

PS2 1 13 (22) 40-80 <1 <0.1

Source: AD and FTD Mutation Database (Nov 2014)

Γονίδια σχετιζόμενα με FTLD ( AD and FTD Mutation Database, Nov 2014)

Μεταλλαγμένο Γονίδιο Φαινότυπος Locus Μεταλλάξεις(οικογένειες)

Expansion of a non-codinghexanucleotide repeat (C90RF72) FTLD/ALS 9p21 1 (336)

Progranulin(PGRN) FTLD 17q21-q22 69 (231)

Microtubule-associated proteintau (MAPT)

FTLD (FTDP-17) 17q21-q22 44 (134)

TDP43 protein (TADPR) ALS/FTLD 1p36 34 (131)

Fused in sarcoma (FUS) ALS/FTLD 16p11.2 23 (49)

Valosin-containing protein gene(VCP) IBMPFD 9p21-p12 18 (48)

Charged multivesicular bodyprotein 2B (CHMP 2B) FTLD 3p13-3p12 4 (5)

Genetic testing for familial dementia with additional neurological features (Loy et al, 2014)

18-year old woman with progressivemyoclonic epilepsy and dementia

– At the age of 14 y.o. visual phenomena– Since 2-3 years myoclonus and possibly absence

seizures– Gradual decline in cognitive functions– FH: negative for neurologic disease– MRI: Normal– Initial response to levetiracetam, later evolution to

generalized seizures despite addition of valproate

PME Type Gene Locus Protein

Unverricht- Lundborgdisease

EMP1 (CSTB) 21q22 Cystatin (proteaseinhibitor)

Lafora disease EMP2AEMP2B/NHLRC1

6q246p22

LaforinMalin

Neuronal ceroidlipofuscinosis (CLN1-8, CLN10)

PPT1 (CLN-1)……

1p32 Palmitoyl-proteinThioesterase- 1

MERRF syndrome A8344G Mitochondrial DNA Lys tRNA

Sialidosis Type Ι NEU1 6p21.33 Neuraminidase(glycoproteinmetabolism)

DRPLA ATN1 12p13.31 Atrophin 1 (nuclearprotein)

Progressive myoclonic epilepsy and dementia

Τύπος PME Γονίδιο Locus Πρωτείνη


EMP 1 (CSTB) 21q22 Cystatin (proteaseinhibitor)


6q246p22

LaforinMalin


PPT1 (CLN-1)……



Sialidosis Type Ι NEU 1- 6p21.33 Neuraminidase(glycoproteinmetabolism)

DRPLA ATN 1 12p13.31 Atrophin 1 (nuclearprotein)

ΕΠ: Μ. Τζαρδή





6q246p22

LaforinMalin


PPT1 (CLN-1)……









6q246p22

LaforinMalin


PPT1 (CLN-1)……





65 year old man with muscular weakness,fibrillations and behavioral disorder

– Since 2010, according to his wife, language problems(“poor speech”), inability with rational associations,change in behavior, recklessness, poor grooming, flataffect, inability to perform his work, loss of interests

– On 2011 weakness of the left upper extremity, graduallyworsening and extending centripetally and then to theother extremities and bulbar muscles, fibrillations,atrophies (on examination increased tendon reflexes)

– FH: Negative for neurological disorders


Turner et al, 2013

Γονίδια σχετιζόμενα με FTLD ( AD and FTD Mutation Database, Nov 2014)

Μεταλλαγμένο Γονίδιο Φαινότυπος Locus Μεταλλάξεις(οικογένειες)

Expansion of a non-codinghexanucleotide repeat (C90RF72) FTLD/ALS 9p21 1 (336)

Progranulin(PGRN) FTLD 17q21-q22 69 (231)

Microtubule-associated proteintau (MAPT)

FTLD (FTDP-17) 17q21-q22 44 (134)

TDP43 protein (TADPR) ALS/FTLD 1p36 34 (131)

Fused in sarcoma (FUS) ALS/FTLD 16p11.2 23 (49)

Valosin-containing protein gene(VCP) IBMPFD 9p21-p12 18 (48)

Charged multivesicular bodyprotein 2B (CHMP 2B) FTLD 3p13-3p12 4 (5)

• Whole exome sequencing

Pure FTD

MAPT

Progranulin

The middlegroundC9ORF72

FUS

VCP

TDP- 43

Pure ALS

SOD 1



Pure FTD

MAPT

Progranulin


FUS

VCP

TDP- 43

Pure ALS

SOD 1



Pure FTD

MAPT

Progranulin


FUS

VCP

TDP- 43

Pure ALS

SOD 1



Pure FTD

MAPT

Progranulin


FUS

VCP

TDP- 43

Pure ALS

SOD 1


- Clinical application of Whole Exome / Genome Sequencing for rare oryet undiagnosed diseases has dramatically increased our diagnosticsuccess rates

- A final diagnosis, even without changing their therapeuticmanagement, gives significant information and comfort to the patientand his family by terminating a long stressful diagnostic process(often involving several different physicians)

- Financially beneficial, as a time-consuming process (diagnosticOdyssey), involving several tests (MRIs, biopsies, blood tests etc) isavoided for a relative minimal cost

Conclusions

Pierre Marie

Jean-MartinCharcot

Henri Parinaud

GeorgesGilles de la Tourette

JosephBabinski

"To learn how to treat a disease, one must learn howto recognize it. The diagnosis is the best trump in thescheme of treatment.”

“Για να μάθει κανείς πως να θεραπεύσει μία νόσο,πρέπει πρώτα να μάθεις πως να την αναγνωρίζεις. Ηδιάγνωση είναι το καλύτερο ατού στο σχήμα τηςθεραπείας.»

Jean-Martin Charcot (1825 – 1893)

Ευχαριστίες (Πρόγραμμα: «Γενετικές αναλύσεις νέας γενιάςσε ασθενείς με νευρολογικές και άλλες παθήσεις»)

• Εμμανουέλλα Βογιατζή• Λένα Λατσούδη• Νευρολογική Κλινική ΠΑΓΝΗ (Γ. Αμοιρίδης, Α. Πλαϊτάκης)• Κλινικοί που μας εμπιστεύτηκαν τα δείγματα των ασθενών

τους (Α. Ευαγγελίου, Β. Μαστοροδήμος, Δ. Κοτζαμάνη, Μ.Τζαγκουρνισάκης, Μ. Μαυρίδης, Π. Βοργιά, Δ. Ζαφειρίου, Α.Μόντη, Γ. Μπριασούλης, κτλ.)

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