2008 Barcelona - European Society of Human Genetics

Cytogenetics
otide-based array platform (244K chip, Agilent, Santa Clara, USA) on
a series of 10 cases that had prior been analyzed on the CytoChip
version 2 .0 (BlueGnome, Cambridge, UK) and partially validated by
FISH. Our findings indicate that cross validation using a second array
CGH platform is a suitable approach for the validation of BAC/PAC array
results that combines the advantages of other validation methods
such as FISH, qPCR or MLPA . The cross validation approach is characterized
by a robust protocol, a rapid work-up, no need for metaphase
spreads and moderate costs .
P02.026
characterization of balanced chromosome translocation
breakpoints associated to phenotype by microdissection and
acGH
O. Villa 1 , N. Kosyakova 2 , I. Cuscó 1 , M. Aragonés 3 , D. García-Cruz 4 , F. Solé 5 , A.
Plaja 6 , T. Liehr 2 , L. A. Pérez-Jurado 1,6 ;
1 Unitat de Genètica, Universitat Pompeu Fabra, U735 CIBERER, Barcelona,
Spain, 2 Institut für Humangenetik und Anthropologie, Jena, Germany, 3 Genes
and disease program. Center for genomic regulation (CRG), Barcelona, Spain,
4 Instituto de Genética Humana “Dr. Enrique Corona Rivera”. Departamento de
Biología Molecular y Genómica, CUCS, Universidad de Guadalajara, Guadalajara,
Mexico, 5 Laboratorio de Citogenética y Biología Molecular. Hospital del
Mar, Barcelona, Spain, 6 Programa de Medicina Molecular i Genètica, Hospital
Vall d’Hebron, Barcelona, Spain.
The positional cloning of genes affected by cytogenetic anomalies associated
with phenotype is one of the main methods for the identification
of disease-related genes . However, mapping translocation breakpoints
and rearranged chromosomal boundaries by molecular cytogenetics
is labour intensive . The isolation of the aberrant chromosomes
by either flow sorting or microdissection permits the specific analysis of
their genomic content by high throughput technologies such as aCGH
or sequencing . We have applied chromosome microdissection and
aCGH to map three apparently translocation breakpoints associated
with different phenotypes: 1) 46,XY,t(4;15)(q22;q26) in a chronic myelomonocytic
leukaemia; 2) 46,XX,t(11;13)(q21;q14) in a patient with
language and developmental delay; and 3) 46,XY,t(7;18)(p15 .1;q21 .1)
in a male with hypogonadotropic hypogonadism. We first isolated one
of the two derivative chromosomes (6-8 chromosomes), amplified the
DNA by DOP-PCR, and performed reverse FISH in order to verify the
appropriate cytogenetic location of the microdissected material . Then,
we hybridized the DNA onto a BAC array and the Agilent 244K CGH
oligoarray for precise mapping . All three translocation breakpoints
have been narrowed down to regions of ~8-15 Kb in each chromosome
with high reproducible signals . We are currently attempting to
further define the exact breakpoints by either tiling array or direct PCR
and sequencing . In summary, the manual microdissection of aberrant
chromosomes is a cheap and reliable method to obtain target DNA for
ulterior high throughput analysis with microarrays .
P02.027
High Resolution comparative Genomic Hybridization analysis in
patients with idiopathic mental Retardation
K. Nieto-Martinez, J. Duarte, C. A. Venegas, S. Kofman-Alfaro;
Facultad de Medicina, UNAM, Mexico City, Mexico.
Mental retardation (MR) and development delay (DD) occur in 2-3%
of the general population and are very heterogeneous entities . Clinical
characteristics observed in these patients are not always related
to a specific syndromes, so different technologies must be applied to
investigate the origin of MR . Conventional karyotyping has a resolution
of >5 Mb and detects chromosomal alterations in >5% of individuals
with unexplained MR, while Comparative Genomic Hybridization
(CGH) is a molecular cytogenetic technique that can characterize unbalanced
genetic material in a one-step global screening procedure .
CGH analysis also provides information about the origin of gains and
losses of chromosomal material and maps these imbalances to their
position on the chromosome . We have used high resolution CGH to
detect deletions and duplications in the range of 2-5 MB in order to
find genomic imbalances in 10 patients with idiopathic mental retardation.
The analysis by HR-CGH reveals 8 cases with normal profile
and two showed abnormal results, one with a gain in 11p15 .1 and
the second one a loss in 12q21 .3 . These cases illustrate the value of
molecular cytogenetic techniques as an important tool in the diagnosis
and assessment of MR as well as a phenotype-genotype correlation .
This work was supported CONACyT Grant No .2005-13947 and UNAM
Grant No . SDEI . PTID .05 .1
P02.028
Array-cGH diagnosis of cryptic chromosome aberrations
R. Stoeva1,2 , L. Grozdanova3 , J. P. Fryns2 , J. Vermeesch2 , I. Ivanov1 , I.
Patcheva1 , I. Stoev1 , B. Dimitrov2 , R. Thoelen2 , T. Krastev1 , M. Stefanova1,4 ;
1Department of Pediatrics and Medical Genetics, Medical University, Plovdiv,
Bulgaria, 2Center for Human Genetics, Katholieke Universiteit, Leuven, Belgium,
3Department of Medical Genetics, University Hospital, Plovdiv, Bulgaria,
4Center for Human Genetics, Free Flemish University Hospital, Brussels, Belgium.
Chromosome abnormalities are the most frequent cause of congenital
malformations/mental retardation syndromes . Recently it was demonstrated
that molecular karyotyping improves the detection rate of submicroscopic
aberrations up to 5-17% . Here we present the molecular
and clinical data of three unrelated individuals with different chromosome
aberrations, depicted by array-CGH analysis .
The smallest described so far deletion 1q44, only 200 kb in size, appeared
de novo in a 4-year-old girl with clinical features inconsistent
with the currently known phenotype of 1qter deletion syndrome . She
presented mild mental retardation, speech delay, epilepsy, persistent
foramen ovale and facial dysmorphism .
A 123 kb duplication of 8p11 .1 chromosome region was detected in a
severely mentally retarded 13-year-old boy with microcephaly, downslanted
palpebral fissures, midface hypoplasia, bird-like nose, long
philtrum and retrognathia . Very few patients with duplications involving
the same chromosome region were reported till now . Surprisingly, both
the mother and healthy brother had cytogenetically balanced translocation
t(9;22)(q22 .1;q13 .1) .
Duplication of CHKAD clone (Shaikh et al ., 2000) within the critical
Di George region was found in an 11-month-old-boy . He was a product
of twin pregnancy and had developmental delay, large fontanel,
hypertelorism, depressed nasal bridge, anteverted nostrils, long philtrum,
microretrognathia and inverted nipples . Few mentally retarded
individuals with 22q11 .2 duplication, but larger in size, approximately
3-6 Mb, were described . The phenotype was variable, characterized
mainly with neurological disturbances .
The reported herein chromosome aberrations, smaller than 1 Mb, contribute
to the interpretation of such anomalies in the clinical practice,
an evolving issue after the routine application of array-CGH .
P02.029
De novo balanced chromosomal rearrangements in patients with
mental retardation and/or multiple congenital abnormalities reanalysed
by array cGH
B. Sikkema-Raddatz, N. Hanemaaijer, Y. Swart, T. Dijkhuizen, C. v. Ravenswaaij-Arts,
K. Kok;
Department of Genetics, Groningen, The Netherlands.
De novo balanced chromosome rearrangements, as detected by cytogenetic
banding techniques, occur in approximately one in 2500 newborns
. The risk of mental retardation (MR) and/ or multiple congenital
abnormalities (MCA) is twice as high as in random populations . It is
plausible that in approximately half of these cases the observed phenotype
is caused by deletion, disruption or otherwise inactivation of a
gene(s) at the breakpoint region(s) . As the resolution of standard cytogenetic
banding techniques is approximately 5 Mb, the unbalanced
nature of some of these translocations may have escaped detection .
We analyzed 19 patients with MR/ MCA and an apparently de novo
balanced chromosome rearrangement at minimal 550 banding level,
by high resolution arrayCGH using an 244K oligo array (Agilent) . Eighteen
patients carried a translocation, one of these patients had a complex
translocation involving three chromosomes . A further patient carried
an inversion . With the 244K oligo array we detected an unbalance
at the breakpoint regions for four patients; at 5 out of 39 breakpoints .
In addition cryptic microdeletions and duplications not located at the
translocation/inversion breakpoints were seen for several patients .
The de novo nature of these aberrations is still under investigation .
The clinical relevance of our findings will be discussed.