0-TESTO COMPLETO.pdf - Fondazione Santa Lucia

TEL.13 – Clinical and laboratory criteria for FSHD diagnosis in view of a national registry…
clinical phenotype. This observation points at the role played by each 4q D4Z4
allele in FSHD onset and progression and has clear implication for genetic
counseling. To confirm this result, we will extend our study to 18 additional
families we already identified.
2. Characterization of FSHD-like myopathic patients
not carrying the D4Z4 pathognomonic mutation
Previous studies suggested that FSHD is a genetically heterogeneous disorder.
In a 14-year period 1466 myopathic subjects were referred to our laboratories
for the molecular diagnosis of FSHD. All of them were investigated
using the standard technique. We failed to detect a p13E-11 allele shorter than
45 kb, in 332 of them. The majority of those cases is most likely represented
by various neurogenic and myogenic FSHD-like disorders could have been
clinically misdiagnosed. However, some of those myopathic patients may
carry a different molecular defect leading to FSHD phenotype. Indeed, the reevaluation
of 190 cases allowed us to select 73 FSHD-like patients. We aim to
continue this screening to verify whether their clinical phenotypes are consistent
with the consortium criteria for FSHD diagnosis. If necessary a complete
electrophysiological study and/or muscle biopsy with extensive study, as outlined
in a previous section will be performed by the clinical partner to exclude
other neuromuscular diseases. Selected patients presenting at least 3 of FSHD
clinical diagnostic criteria will be considered FSHD-like and will be further
investigated.
F. Detection of p13E-11 deletion – We must also consider that in 3% of
patients showing FSHD clinical features, there is a deletion of the region
proximal to the polymorphic D4Z4 repeat array that encompasses the p13E-
11probe [Lemmers et al. 2003]. In these cases the pathogenic D4Z4 allele is
undetectable by Southern hybridization of p13E-11 probe. To identify these
subjects among our cases, we will carry out an experimental procedure by
using 4qA and 4qB probes on HindIII digested DNA as described by Lemmers
et al. [2003]. Hybridization of HindIII digested DNA with 4qA/4qB probes
allows detection of deleted fragment in the 4q35 region, that were undetectable
in EcoRI digested DNA by p13E-11 hybridization, because lacking
the DNA sequence detected by probe p13E-11.
At present, through clinical re-evaluation and molecular analysis of 190
myopathic patients, we identified 5 subjects carrying a p13E-11 deletion.
Selected FSHD-like patients will be further investigated.
G. Methylation studies – To explain FSHD pathogenesis, we proposed that
D4Z4 deletion through modification of chromatin structure causes transcriptional
derepression that eventually leads to disease [Gabellini et al. 2002;
Tupler, Gabellini 2004].
In agreement with our hypothesis, Van Overvelds et al described D4Z4
hypomethylation in FSHD and in FSHD-like patients not carrying any D4Z4
deleted allele [van Overveld et al. 2003]. CpG DNA methylation is considered
the hallmark of gene repression and is known to be involved in X-chromosome
inactivation, imprinting and gene silencing [Robertson, Wolffe 2000]. It
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