2009 Vienna - European Society of Human Genetics

Molecular basis of Mendelian disorders
of Medical Biology and Genetics, Adana, Turkey, 3 Cukurova University School
of Medicine, Department of Neurology, Adana, Turkey.
Scientific background: The spinal muscular atrophies (SMAs) causing
degeneration of the anterior horn cells of the spinal cord characterized
by progressive weakness of the lower motor neurons. Several types of
SMA have been described depend on age when accompanying clinical
features appear. The most common types are acute infantile (SMA
type I, or Werdnig-Hoffman disease), chronic infantile (SMA type II),
chronic juvenile (SMA type III or Kugelberg-Welander disease), and
adult onset (SMA type IV) forms. SMA is diagnosed with detection of
homozygous deletions of SMN1 (exon 7 - 8 or exon 7) gene in molecular
level.
Objectives: It is aimed to conduct molecular analysis of exon 7 and 8
of SMN gene in sixty five subjects of SMA (66 patients).
Materials and methods: PCR-RFLP method is used for detection of
homozygous exon 7 - 8 deletions. PCR-SSCP method was used either
to identify for intragenic mutations and especially compound heterozygotes
or to confirm some SMA patients homozygous deletions
detected by RFLP.
Conclusion: In this study, 94 % (62/66) of SMA patients including all
types were found homozygous for exon 7 and 8 deletions with RFLP
method. The rate of homozygous deletions determined was 94.7%
(18/19) in type I patients, 96% (22/23) in type II and 90% (18/20) in
type III. SSCP method was used only for 4 subjects who are clinically
diagnosed as SMA patients but not confirmed with RFLP analysis. The
results of SSCP analyses led to decision that patients may be of compound
heterozygous or intragenic mutations.
P12.151
New DNA microvariations described by smRt arrays.
P. Armero1 , B. Hernández-Charro1 , A. Hernández1 , R. Agudiez1 , J. Fdez-Toral2 ,
P. Madero1 ;
1 2 Centro de Análisis Genéticos, Zaragoza, Spain, Genetics Department. Hospital
Universitario Central de Asturias, Spain.
Introduction: Genome screening using array CGH has great potential
in the characterization of unexplained chromosomal aberrations. The
whole genome Sub-Megabase Resolution Tiling Array (SMRT array) is
capable of identifying microamplifications and microdeletions at a resolution
of 100 Kb. Other different techniques, such as MLPA or FISH,
are traditionally employed to detect these chromosomal alterations. In
this study we show the utility of the SMRT arrays to provide precise
information about the size and breakpoints of DNA copy number gains
and losses.
Methods: We present a patient with unexplained mental retardation
and a male normal karyotipe 46, XY. MLPA kit (from MRC-Holland)
technique was carried out. A SMRT array, (from Wan Lam Laboratory
at the BC Cancer Research Centre) analysis was performed to confirm
and describe the alteration.
Results: MLPA study showed a 14q deletion of about 1.5 Mb. The 14q
specific probe of MLPA kit was deleted. The SMRT array analysis of
the specific 14q32.33 region confirmed this microdeletion and allowed
us to exactly describe its size into 2.20 Mb.
Conclusions: The SMRT array study confirms a small deletion of 2.20
Mb unless than 1.5 Mb previously detected by MLPA. SMRT array
arises as an effective technique to detect DNA microvariations and
provides more information about their size and precise breakpoints.
P12.152
sNP array analyses can orient molecular diagnosis of autosomal
recessive heterogenous diseases in sporadic cases from
consanguinous families
M. C. Vincent1,2 , E. Schaefer3 , M. Cossée1,2 , C. Lagier-Tourenne1,4 , N. Dondaine1
, H. Dollfus3,2 , C. Tranchant5 , P. Charles6 , J. Amiel7 , C. Antignac7 , I.
Vuillaume8 , M. Koenig1,4 , J. L. Mandel1,4 ;
1 2 Laboratoire de Diagnostic Génétique, CHRU, Strasbourg, France, Laboratoire
de Génétique Médicale, EA3949, Faculté de Médecine, Strasbourg, France,
3 4 Service de Génétique Médicale, CHRU, Strasbourg, France, IGBMC (CNRS/
INSERM/ULP), Illkirch, France, 5Service de Neurologie, CHRU, Strasbourg,
France, 6Consultation de Génétique, Pitié Salpêtrière, AP-HP, Paris, France,
7Département de Génétique, Necker Enfants Malades,AP-HP, Paris, France,
8Centre de Biologie-Pathologie, CHRU, Lille, France.
Molecular diagnosis of rare autosomal recessive diseases with extensive
genetic heterogeneity represents a real challenge because
clinical data do not in most cases suggest a particular defective gene.
Consanguinity is frequent in such families. Genome wide SNP array
analysis allows, by searching for homozygous regions in such patients,
the selection of one or few candidate genes in which to search for mutations.
We report 8 such cases including 6 sporadic ones, where the
disease causing gene and mutation were found using this approach
(see table).
case Form Disease
Number of candidatehomozygous
segments
mutated gene
1 Sporadic Myopathy 1 TRIM32
2 Familial Spastic paraplegia 1 SPG11
3 Familial Ataxia 1 AOA1
4 Sporadic Achromatopsia 2 CNGB3
5 Sporadic Bardet Biedl 1 BBS1
6 Sporadic Ataxia 6 FXN
7 Sporadic Bardet Biedl 2 BBS6
8 Sporadic Bardet Biedl 2 BBS5
Homozygosity mapping using 50K micro-arrays (Affymetrix) was performed only on
the patients and allowed us to identify causative mutation in a significant proportion of
sporadic cases affected with different neuromuscular or neurosensory diseases (see
table.). This rapid and not too expensive approach is particularly useful for diseases
with extensive genetic heterogeneity like Bardet Biedl syndrome (14 genes published
to date) , limb girdle muscular dystrophy or sporadic ataxias, by selecting only one or
two genes for sequencing and identify the private mutation. In some cases, SNP array
analysis can reveal consanguinity that was unknown to or denied by the family
P12.153
Twenty novel mutations in SPG11/spatacsin identified using
both direct sequencing and mLPA
G. Stevanin1,2 , C. Depienne1,2 , E. Denis2 , E. Fedirko2 , E. Mundwiller1 , S. Forlani1
, C. Cazeneuve2 , E. Le Guern2 , A. Durr1,2 , A. Brice1,2 ;
1 2 CRicm UMRS975/NEB, Paris, France, Département de Génétique et Cytogénétique,
Paris, France.
Objective: To extend the SPG11 mutation spectrum and establish the
frequency of genomic rearrangements in this gene.
Background: Truncating point mutations in SPG11/spatacsin are the
major cause of autosomal recessive spastic paraplegia with thin corpus
callosum. Recently genomic rearrangements were also involved.
Methods: 45 unrelated patients with spastic paraplegia with thin corpus
callosum +/- mental retardation or cognitive delay were screened
using direct sequencing and MLPA.
Results: 25 different SPG11 point mutations, 18 of which were novel,
were identified in 16 patients (36%). All mutations but one introduced
premature termination codon in the protein sequence and were compatible
with a degradation of the corresponding mRNA by the nonsense-mediated
mRNA decay. The remaining mutation was a missense
variant which alters a highly conserved amino-acid of the protein
and was found associated with a truncating mutation. In addition,
MLPA analysis detected heterozygous SPG11 micro-rearrangements
in two patients who already had a single heterozygous point mutation.
Analysis of the affected relatives and parents when possible showed
that the mutations segregated with the disease and that heterozygous
compound mutations were inherited each from a healthy parent. Only
two patients out of 16 had homozygous mutations; the remaining 14
patients had heterozygous compound mutations. Finally, we identified
new missense polymorphisms that did not segregated with the disease.
Conclusions: These findings expand the SPG11 mutation spectrum
and highlight the importance of screening the whole coding region with
both direct sequencing and a quantitative method.
Rare missense polymorphisms are frequent in SPG11, complicating
interpretation of diagnosis.
P12.154
sPG4 mutations can mimic primary progressive multiple
sclerosis on clinical, biological and mRi aspects
P. Charles1 , C. Depienne1 , B. Fontaine2 , C. Lubetzki2 , O. Lyon-Caen2 , A. Durr1 ,
A. Brice1 ;
1 2 Département de Génétique et Cytogénétique, Paris, France, Fédération des
Maladies du Système Nerveux, Paris, France.
The most common form of autosomal dominant hereditary spastic
paraplegia (AD-HSP) is caused by mutations in the SPG4/SPAST