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Miclea D., Alkhzouz C., Popp R.A., Zimmermann A., Lazar C., Lazea C., Bucerzan S., Farcas M., Grigorescu-Sido P.
▪ Testarea genetica efectuata la Laboratorul de Genetica al Spitalului Clinic de Urgenţă pentru
Copii, Centrul Regional de Genetica Medicala Cluj
▪ Testarea genetica – diagnostic pozitiv intr-o patologie genetica
▪ Genetica medicala in urmatorii 15 ani – medicina genomica (medicina de precizie)
▪ Diagnostic si evolutie: boli rare, boli comune
▪ Tratament: Farmacogenetica
Ghiduri de testare genetica, in Europa - utilizate de ≈ 3-5 ani▪ Patologia endocrina: DSD/hipostatura – panel de gene NGS
▪ Boli genetice de metabolism: panel de gene NGS
▪ DD/ID, TSA, epilepsie – array CGH/SNP, panel, exom NGS
▪ Aceasta se poate efectua pentru tot mai multe patologii şi ȋn ţara noastră
1) patologia endocrină (Miclea D)
▪ ADS/hiperplazia congenitala de corticosuprarenala şi anomalii ale pubertăţii
▪ hipostatura şi anumite displazii scheletale
▪ obezitatea
2) fibroza chistica şi anumite boli genetice de metabolism (Farcas M)▪ boala Gaucher, deficitul de alfa1antitripsină
3) patologia senzoriala (surditatea), psihiatrica şi neurologică (Farcas M, Miclea D)
▪ RD/DI, TSA, epilepsia - forme izolate sau sindromice
▪ aparatura▪ Laboratorului de Genetica Medicala al Spitalului Clinic de Urgenta pentru Copii, Centrul Regional Cluj
▪ Laboratorului de Genetica Medicala, UMF Cluj
▪ Parteneriate cu:▪ Centrul Imogen, Cluj-Napoca
▪ Centrul de Medicina Genomica, Timisoara
▪ Colaborare cu:▪ Centrul Regional de Genetica Medicala, Dolj
▪ Spitalul Robert Debre (Dr AC Tabet) si Spitalul Bicetre (Dr J Bouligand), Paris, Franta
▪ Tehnici utilizate▪ Citogenetica clasica
▪ FISH
▪ PCR si variante
▪ Stripassay
▪ MLPA
▪ SNP array/CGH array
▪ NGS
▪Anomalii de dezvoltare sexuala
NEONATAL
▪ OGE ANORMALE
▪ Hipertrofie clitoridiana izolata
▪ Hipospadias posterior izolat
▪ Criptorhidie/ectopie testiculara unilaterala+micropenis
▪ Criptorhidie/ectopie testiculara unilaterala+hipospadias
▪ Criptorhidie/ectopie bilaterala: gonade palpabile in pozitie inghinala sau nici o gonada palpabila
Anomalii de dezvoltare sexuala
PUBERTAR
▪ OGE ANORMALE
▪ Virilizare OGE la momentul reactivarii axului gonadotrop
▪ Deficit 5 alfa reductaza, Deficit 17 cetoreductaza testiculara, ovotestis
▪ RETARD PUBERTAR
▪ Disgenezie gonadala (Turner, Klinefelter)
▪ Cariotip 45,X/46,XY
▪ AMENOREE PRIMARA
▪ Rezistenta completa la androgeni
▪ Dozari hormonale▪ 17OH progesteron
▪ DHEAS
▪ delta4 androstendion
▪ Testosteron
▪ AMH
▪ Ecografie gonade+OGI
▪ Cariotip+SRY
▪ Testare deficit 21OH
▪ Panel gene ADS
▪ Cariotip+SRY
▪ Deficit 21 hidroxilaza – stripassay 11 mutatii mai frecvente
▪ Panel gene:
▪ TruSight One 4800 gene OMIM morbide
Patient age(yrs)
social
gender phenotype Caryotype
Hormonal
investigation Ultrasounds SNP array NGS
p1 10male
Micropenis. 5th
clinodactyly 46,XY Testo N DHT ↓
N gonads, no
mullerian
derivates no pathogen CNV N
p2 18female
clitoridian
hypertrophy 46,XX N gonads, OGI VN no pathogen CNV N
p3 1female
Right inguinal
hernia. Feminin
OGE. 2 aunts
maternal line with
primary
amenhoree 46,XY N
left gonads in
inguinal conduct.
Absence muller
residus no pathogen CNV AR
p4 5male
Penoscrotal
hipospadias.
Craniofacial
dysmorphism.
Blefarophimosis.
Short stature. Aortic
bicuspidia. DSV.
Development delay 46,XY N
gonads N, no
mullerian residus VOUS CNV SYCE1 N
p5 7male
Micropenis.
Testicular
hypoplasia 46,XY N VOUS CNV SYCE1
p6 18male
Micropenis.
Pubertary delay 46,XY N CNV dup 16p11.2
p7 1female
clitoridian
hypertrophy 46,XX 17OHP↑ uterus Del CYP21A2
penoclitoridian
gland, labial
hypertrophy
fusionated
Patient age(yrs)
social
gender phenotype Caryotype
Hormonal
investigation Ultrasounds SNP array NGS
p9 3male
micropenis.
Hipospadias 46,XY N no muller residus no pathogen CNV
p10 20female
amenoree. Hirsutism.
Gonadal dysgenesis 46,XX N no pathogen CNV
p11 1female
clitoris hypertrophy,
labial hypertrophy,
vaginal and uretral
meatus presents,
right inguinal hernia 46,XY T DHT ↑
in inguinal hernia-
ovalary tumors with
testicule
appearance no pathogen CNV AR
p12 2male
proximal
hypospadias.
Criptorchidism 46,XY no pathogen CNV
p13 3female
gonadal dysgenesis,
craniofacial
dysmorphisms, short
4th and 5th
metacarpals, 5th
clinodactyly, SGA,
short stature 46,XY AMH, testo↓ no muller residus no pathogen CNV
p14 3male severe hypospadias 46,XY N no pathogen CNV MAMLD1
p15 14male
Penian hypospadias.
Right cryptorchidism.
Ginecomastia.
Pubertal
development. 46,XY N
no muller residus,
left testicule in
inguinal conduct, a
smaller tumor with
the same
localisation in right
side no pathogen CNV
p16 3male
Bilateral
cryptorchidism.
Gonadal dysgenesis 46,XY LH,FSH ↑↑↑
left scleroatrophic
testicule, ecografic,
absent right
testicule no pathogen CNV
penoscrotal
hypospadias. Scrotal
17 patients – 3 CNV VOUS/pathogenic
10 p15.3 p15.1 p14 p13 p12.31 p12.1 p11.22 p11.21 q11.21 q11.22 q11.23 q21.1 q21.2 q21.3 q22.1 q22.2 q22.3 q23.1 q23.31 q23.33 q24.2 q25.1 q25.2 q25.3 q26.13 q26.2 q26.3
Base Position 130.205.223 130.797.393 131.389.563 131.981.733 132.573.903 133.166.073 133.758.243 134.350.413 134.942.583
Cytogenetic Band q26.2 q26.3
Sequence (+) No sequence data file found for this chromosome.
CLRN3
PTPRE
PTPRE
MKI67
MKI67
MGMT
EBF3
GLRX3 TCERG1L PPP2R2D
PPP2R2D
PPP2R2D
BNIP3
JAKMIP3
DPYSL4
STK32C
LRRC27
LRRC27
LRRC27
LRRC27
LRRC27
PWWP2B
PWWP2B
C10orf91
INPP5A
NKX6-2
C10orf93
GPR123
KNDC1
UTF1
VENTX
MIR202
ADAM8
ADAM8
ADAM8
TUBGCP2
ZNF511
CALY
PRAP1
PRAP1
C10orf125
C10orf125
ECHS1
SPRN
LOC619207
CYP2E1
SYCE1
SYCE1
SYCE1
FRG2B
LOC728410
DUX4
LOC653548
LOC653544
LOC653543
LOC653545
LOC728410
DUX4
LOC653543
16 p13.3 p13.2 p13.13 p13.12 p13.11 p12.3 p12.2 p12.1 p11.2 p11.1 q11.1 q11.2 q12.1 q12.2 q21 q22.1 q22.2 q23.1 q23.2 q23.3 q24.1 q24.2q24.3
Base Position 28.672.615 29.071.335 29.470.055 29.868.775 30.267.495 30.666.215 31.064.935 31.463.655 31.862.375
Cytogenetic Band p11.2
Sequence (+) No sequence data file found for this chromosome.
SBK1
EIF3CL
EIF3C
EIF3C
CLN3
CLN3
APOB48R
IL27
NUPR1
NUPR1
CCDC101
SULT1A2
SULT1A2
SULT1A1
SULT1A1
SULT1A1
SULT1A1
SULT1A1
EIF3C
EIF3C
EIF3CL
ATXN2L
ATXN2L
ATXN2L
ATXN2L
ATXN2L
TUFM
SH2B1
SH2B1
SH2B1
SH2B1
SH2B1
ATP2A1
ATP2A1
RABEP2
CD19
SPNS1
LOC653390
RUNDC2C
LOC606724
BOLA2
BOLA2B
GIYD1
GIYD2
GIYD2
GIYD1
SULT1A3
SULT1A4
SULT1A3
SULT1A4
LOC388242
LOC388242
IMAA
MMAA
SPN
SPN
QPRT
C16orf54
ZG16
KIF22
MAZ
MAZ
PRRT2
C16orf53
MVP
MVP
CDIPT
LOC440356
LOC440356
SEZ6L2
SEZ6L2
SEZ6L2
ASPHD1
TMEM219
TMEM219
TAOK2
TAOK2
HIRIP3
INO80E
DOC2A
C16orf92
C16orf92
ALDOA
ALDOA
ALDOA
ALDOA
PPP4C
TBX6
CORO1A
LOC606724
BOLA2
BOLA2B
GIYD1
GIYD1
GIYD2
GIYD2
SULT1A4
LOC613037
LOC595101
IMAA
CD2BP2
CD2BP2
TBC1D10B
MYLPF
SEPT1
ZNF48
ZNF771
ZNF771
DCTPP1
SEPHS2
ITGAL
ITGAL
ZNF768
ZNF747
ZNF764
ZNF688
ZNF688
ZNF785
ZNF689
PRR14
FBRS
SRCAP
SNORA30
PHKG2
C16orf93
RNF40
ZNF629
BCL7C
MIR762
CTF1
CTF1
NCRNA00095
FBXL19
ORAI3
SETD1A
HSD3B7
HSD3B7
HSD3B7
STX1B
STX4
ZNF668
ZNF646
POL3S
VKORC1
VKORC1
BCKDK
BCKDK
MYST1
FUS
FUS
FUS
FUS
PYCARD
PYCARD
TRIM72
ITGAM
ITGAM
ITGAX
ITGAD
COX6A2
ZNF843
ARMC5
ARMC5
TGFB1I1
TGFB1I1
TGFB1I1
SLC5A2
C16orf58
ERAF
CSDAP1
C16orf67
ZNF720
ZNF267 HERC2P4
6 p24.3 p22.3 p22.1 p21.2 p21.1 p12.3 p12.1 q12 q13 q14.1 q14.3 q15 q16.1 q16.3 q21 q22.1 q22.31 q23.2 q23.3 q24.1 q24.3 q25.1q25.2 q25.3 q26 q27
Base Position 31.470.581 31.829.331 32.188.081 32.546.831 32.905.581 33.264.331 33.623.081 33.981.831 34.340.581 34.699.331
Cytogenetic Band p21.33 p21.32 p21.31
Sequence (+) No sequence data file found for this chromosome.
CCHCR1
CCHCR1
CCHCR1
TCF19
TCF19
POU5F1
POU5F1
PSORS1C3
HCG27
HLA-C
HLA-B
MICA
HCP5
HCG26
MICB
MCCD1
BAT1
BAT1
SNORD117
SNORD84
ATP6V1G2
ATP6V1G2
NFKBIL1
NFKBIL1
NFKBIL1
NFKBIL1
LTA
LTB
AIF1
BAT2
APOM
C6orf47
BAT4
CSNK2B
LY6G5C
LY6G6F
G6e
G6E
LY6G6D
LY6G6C
C6orf25
C6orf25
C6orf25
C6orf25
C6orf26
VARS
LSM2
HSPA1L
HSPA1A
HSPA1B
C6orf48
C6orf48
NEU1
SLC44A4
EHMT2
EHMT2
ZBTB12
C2
C2
CFB
RDBP
MIR1236
SKIV2L
DOM3Z
STK19
STK19
C4A
C4B
CYP21A2
CYP21A2
TNXB
TNXB
ATF6B
ATF6B
FKBPL
PRRT1
PPT2
PPT2
EGFL8
AGPAT1
AGPAT1
RNF5
RNF5
AGER
NOTCH4
C6orf10
BTNL2
HLA-DRA
HLA-DRB5
HLA DRB4
HLA-DRB1
HLA-DQA1
HLA-DQB1
HLA-DQA2
HLA-DQB2
HLA-DOB
TAP2
TAP2
PSMB8
PSMB8
TAP1
PSMB9
PSMB9
PPP1R2P1
HLA-DMB
HLA-DMA
BRD2
BRD2
HLA-DOA
HLA-DPA1
HLA-DPB1
HLA-DPB2
COL11A2
COL11A2
COL11A2
COL11A2
RXRB
SLC39A7
SLC39A7
HSD17B8
MIR219-1
RING1
VPS52
RPS18
B3GALT4
WDR46
TAPBP
ZBTB22
LYPLA2
KIFC1
PHF1
PHF1
PHF1
CUTA
CUTA
CUTA
CUTA
CUTA
SYNGAP1
ZBTB9
BAK1
GGNBP1
C6orf227
ITPR3
C6orf125
IP6K3
IP6K3
LEMD2
LEMD2
MLN
MLN
MIR1275
GRM4
HMGA1
HMGA1
HMGA1
HMGA1
HMGA1
HMGA1
C6orf1
C6orf1
C6orf1
NUDT3
RPS10
PACSIN1
SPDEF
C6orf106
C6orf106
Patient chr start stop size CNV Gene
Interpretatio
n
4 10 135257091 135378802 121711 DupSYCE1 VOUS
5 10 135252347 135378802 126455 DupSYCE1 VOUS
6 16 29595483 30192561 597078 Dup16p11.2 Pathogenic
7 6 32005904 32006896 992
Homo
delCYP21A2 Pathogenic
▪ 9 tested patients
▪ 6 patients
▪ MAMLD1 (Xq28): c.1066C>T exonic stopgain mutation
▪ Xlr
▪ Clinical picture – micropenis, hypospadias, bifid scrotum
▪ UMD predictor: pathogenic
▪ EXAC: 2 alleles for 121410 chromosomes
▪ AR gene - 2 patients
I. Forma centrala
Hipogonadism hipogonadotrop:
- tranzitor
- permanent
PANEL GENE
II. Forma periferica
Hipogonadism hipergonadotrop:
- afectare gonadala primara – intotdeauna patologic
- cel mai frecvent disgenezie gonadala secundara anomaliilor cromozomilor sexuali
CARIOTIP
▪ Suspiciune clinica daca• micropenis si/sau criptorhidism la nou-nascutii de sex masculin
• retardul pubertar la o varsta osoasa mai mare de 13 ani
▪ izolat (este afectat doar axul gonadotrop)
▪ 50% din cazuri cu anosmie/hipoosmie = sindrom Kallmann
▪ asociat altor afectari endocrine
▪ non-sindromic
▪ sindromic anomalii ale liniei mediane -despicaturi labio-palatine, agenezii dentare- anomalii ale urechii si surditate, agenezii renale, malformatii cardiace, anomalii scheletice ale extremitatilor, sinkinezii bimanuale
•HHC izolat – NGS – 9 pacienti• Panel gene: GNRHR, GNRH1, KISS1R, KISS1, TACR3, TAC3, KAL1, FGFR1, FGF8, PROKR2,
PROK2, WDR11, CHD7, SEMA3A, NSMF, HS6ST1, FSHB, LHB, SOX3, FGF17, IL17RD, DUSP6,SPRY4, FLRT3, PROP1, NR0B1, PCSK1, LHX4, HESX1, OTX2, RNF216, OTUD4, SOX2, POU1F1,SOX10, KALP, CUL4A, CUL4B, GNRH2, NRP1, NRP2, SIX6, PDYN, OPRK1, TAC1, TACR1,TACR2, NPVF, NPFFR1, PLXNA1, SEMA7A, LHX3, NPY, LHX2, POU2F1, POU3F2, SLIT2,ROBO3, LEPR, LEP, SLIT3, CGA, INHBA, PRLR, PCSK2, PLXNC1, DCC, ZIC1, LIFR, FARP2
•HHC+obezitate – MS-MLPA Prader Willi
OBEZITATE
+▪ Hipotonie+tulburari de alimentatie la nou-nascut
▪ Hipostatura, anomalii scheletale (brahimetatarsie)
▪ Endocrinopatii: hipogonadism
▪ Retard psihomotor
▪ Tulburare vizuala, auditiva, retard limbaj
▪ Epilepsie, tulburare de comportament
▪ Sindrom dismorfic, sindrom malformativ
=>OBEZITATE GENETICA
▪ Prader Willi: MS-MLPA - deletii si UPD
▪ Obezitate+Retard mental: PCRq - CNV 16p11.2; 15q11.1
▪ Obezitate+retard mental+sindrom dismorfic+sindrom malformativ: MLPA microdeletii
▪ SNP array/CGH array – ideal!
HIPOSTATURA
Factori genetici > 80% Factori de mediu
• Factori etnici, talia parentala
Cresterea - Proces multifactorial
Factori de mediu!
Ultimii 150 ani – trend secular
HIPOSTATURA - CLASIFICARE• primara: afectiuni intrinseci ale osului
• Displazii scheletale (3%)
• Sindroame genetice (21%)
• RCIU fara recuperare (12%)
• secundara: afectiuni ce modifica fiziologia cartilajului de crestere
• Endocrine (11%)
• Afectiuni cronice (3%)
• Malnutritie, cauze metabolice, psihosociale
• Idiopatica (50%)
• Hipostatura Constitutionala (idiopatica), familiala
Lam WFF, et al., Chin Med J 2002:115, 607-6117
▪ La fete – cromozomi sexuali si SHOX - MLPA
▪ La baieti – 45,X/46,XY? – cariotip
Afectare endocrina – Hipostatura proportionata
▪ PROP1
▪ Ideal Panel gene
Afectare scheletala – Hipostatura disproportionata
▪ SHOX - MLPA
▪ FGFR3 - PCR
▪ Ideal panel de gene
▪ Daca exista suspiciune clinica
▪ test genetic tintit
▪ Daca nu exista suspiciune clinica
▪ Evaluare genomica: MLPA, SNP array, NGS – panel de gene, exom
SINDROM NOONAN
▪ PTPN11
from, Baraitser and Winter, Color Atlas of Congenital Malformation Syndromes, 1996
SINDROM RUSSELL-SILVER
▪ 10% - UPD 7mat
▪ Anomalii ale metilarii 11p15.5-35%
(from, Baraitser and Winter, Color Atlas of Congenital Malformation Syndromes, 1996)
SINDROM RUBINSTEIN-TAYBI
• <10% del 16p13.3
• 10-20% del/dup CREBBP
(from, Baraitser and Winter, Color Atlas of Congenital Malformation Syndromes, 1996)
SINDROM WILLIAMS
• 99%: del 7q11.23
(from, Baraitser and Winter, Color Atlas of Congenital Malformation Syndromes, 1996)
▪ FIBROZA CHISTICA
▪ BOLI GENETICE DE METABOLISM
Boala Gaucher detectia a 8 mutatii comune GBA prin metoda StripAssay
Deficit de alfa 1 antitripsina A1AT – identificarea alelelor M,S si Z
▪ SURDITATEA nonsindromica – 328 cazuri analizate
▪ GJB2 - 35delG, W24X
▪ GJB6 - D13S1830 si D13S1854
▪ MLPA
Testare c35delG
Homozigot heterozigot negativă
Testare pW24X
Homozigot Heterozigot negativă
(compus)
Testare GJB6
Homozigot Heterozigot negativă
(compus)
SECVENȚARE
Lazar C, Int J Pediatr Otorhinolaryngol, 2010
-X fragil
-cariotip (1200 cazuri analizate)
-MLPA microdeletii (430 cazuri analizate)
-SNP array/CGH array (210 cazuri analizate)
DIAGNOSTIC MAI BUN
▪ un randament diagnostic bun şi astfel un bun raport cost-beneficiu derivă din indicaţia adecvată a acestor teste –
centre de expertiza cu echipa multidisciplinara
▪ Cluj – Deficit 21 hidroxilaza, Boli lizozomale
▪ utilizarea ghidurilor pentru testare genetică!
▪ testarea genetica presupune din ce in ce mai mult folosirea unor tehnologii de analiza a genomului, utilizate
deja la noi in tara – pentru o interpretare buna – trebuie un numar limitat de patologii/centru
TRATAMENT MAI BUN
▪ intelegerea unui mecanism molecular va conduce la alegerea unei terapii de precizie (ex.tulburarile din
spectrul autist, oncologie)
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