Discussion
The present girl came to our observation at the age of 3 months for
severe epileptic seizures, started 3 weeks early with episodes of
loss-of-contact of brief duration, then evolved in severe drug-resistant
epilepsy mainly characterized by spasms in flexion; brain MRI disclosed
a complex brain malformation, with thin corpus callosum, polymicrogyria,
cortical dysplasia, heterotopias and asymmetric ventricles. These
findings, together with a chorioretinal atrophy and optic disc coloboma,
allowed to perform a clinical diagnosis of Aicardi syndrome (Aicardi,
2005). Submitted to a NGS-gene panel for epileptic encephalopathies and
brain malformation syndromes, the girl was found as harboring two
variants in trans (compound heterozygosis) of DCHS1 gene:
c.7408G>T: and c.3512G>A. In a in
silico evaluation, both showed pathogenic significance, even with
different grade of pathogenicity, as demonstrated by the analysis
through VarCards server, which reports the results from 23 distinct
prediction tools, including SIFT, Polyphen2 and CADD (Varcards Database,
2020). These variants are unlikely to affect splicing (only one splice
site is altered by the variant c.7408G>T) and are both
missense variants, therefore unlikely to result in a truncated protein.
However they could affect stability and localization of the protein, or
its interaction with other proteins of the Hippo-pathway; in this
aspect, western blots and/or immunofluorescence microscopy would be, in
future, helpful to clarify more the pathogenic role of such variants.
DCHS1 gene encodes a transmembrane cell adhesion molecule that
belongs to the protocadherin superfamily. DCHS1 is a ligand for FAT4,
which is another protocadherin; DCHS1 and FAT4 form an apically located
adhesive complex in the developing brain, but their functions
encompasses also an important role in cell-cell adhesion of fibroblasts,
which is thought to be necessary for wound healing (Cappello et al,
2013). It has been shown that mutations in genes encoding the
receptor-ligand cadherin pair DCHS1 and FAT4 lead to periventricular
neuronal heterotopia, both in humans and in experimental mice models,
with reduced expression of Dchs1 or Fat4 (Beste et al,
2016). In particular, within mouse embryonic neuroepithelium, it has
been observed an increased progenitor cell numbers and a reduction of
their differentiation into neurons, resulting in the heterotopic
accumulation of cells below the neuronal layers in the neocortex,
reminiscent of the human phenotype. A concurrent knockdown of Yap, a
transcriptional effector of the Hippo signaling pathway, was also
observed, thus confirming that Dchs1 and Fat4 proteins are upstream of
Yap as key regulators of mammalian neurogenesis (Cappello et al, 2013)
(Beste et al, 2016).
DCHS1 gene is the causative factor of a rare (less than 20
patients reported so far) and complex malformation syndrome, Van
Maldergem type 1, which is characterized, in its classical form, by
intellectual disability, typical craniofacial features, auditory
malformations, hearing loss, skeletal and limb malformations, brain
abnormalities with periventricular neuronal heterotopia and other
variable anomalies (van Maldergem et al, 1992) (Sotos et al, 2017)
(Ulubas et al, 2018) (Ivanovski et al, 2018). The function ofDCHS1 gene product in fibroblast adhesion can explain why some
allelic variants of this gene have been found in a familiar form of
mitral valve prolapse, with high penetrance (Durst et al, 2015).
An exact cause of Aicardi Syndrome has never been precisely elucidated
and its pursuit is still matter of debate (Wong et al, 2018). In the
last 15 years, many studies have suggested a potential role of
X-chromosome (Xp22) or other autosomal (1p, 3q, 6q, 12q) translocations.
At the same time, male (46XY) patients have been reported as affected by
severe microcephaly (that is not a distinctive sign of the disease),
thus suggesting that the (possible) mutation in X-chromosome may not be
always lethal but (in some cases) related to a very severe phenotype in
males or that other genes may be involved in the pathogenesis of the
disease (Chappelow et al, 2008). Modern array-CGH studies, however, have
failed in reporting (small or large) CNVs in X-Chromosome, with only one
patient out of a total of 156 subjects found to have a de novo 157-kb
deleted region in Xq25, in which no known codifying exons of any known
gene were found (Wong et al, 2018).
A monogenic origin of the disease has been also investigated with
potential candidate genes. In particular FLNA gene, which encodes
for a protein (Filamin A) involved in brain cytoskeleton organization
and whose mutations are related to brain heterotopias; however, no
disease-causing mutations for this variants have been found in AS
patients (Anderson et al, 2009). Another gene whose variants were
potentially associated with AS was CDKL5 , which plays a role in
synaptic formations and is implicated in the etiology of early onset
drug-resistant epilepsies (Nemos et al, 2009), but a genetic analysis of
this gene in 10 patients affected by AS gave negative results. With the
development of NGS gene panels and the use of whole exome sequencing
(WES) or Whole genome sequencing (WGS) in large cohort of AS patients,
two novel variants in previously unidentified genes have been found
(Schrauwen et al, 2015): the genes involved, in two unrelated girls,
were TEAD1 (11p15.3) and OCEL1 (19p13.11): however a
further Sanger sequencing of these two genes in other 38 AS patients
failed to find pathogenic variants of these genes (Wong et al, 2017).
Hypomethylation of KCNAB3 (17p13.1) was also proposed among the
causes of the disease (Piras et al. 2017), but no consistent finding has
been found. More recently, a WES-based study on 11 patients affected by
AS has failed to find any significant variant in these patients (the
Authors analyzed in particular TEAD1 and OCEL1 ) (Lund et
al, 2016).
It is worthwhile to note that the genetic location of the mutated gene
of the present patient (DCHS1 ) is close to TEAD1. Given
the proximity of TEAD1 and DCHS1 , it cannot be excluded
that an imbalance in this region may be among the causes of AS, which
could be hypothetically caused by DCHS1 variants and not byTEAD1 mutations. If DCHS1 has a pivotal role, however, it
cannot be excluded that some (still unknown) gene regulators or
modifiers localized in X-chromosome may contribute to the phenotype.
By contrast, there are more than 50 genes in the region containingDCHS1 and TEAD1 , many of which are involved in
autoimmunity, therefore it is unlikely that mutations in all genes in
this region will share the same phenotype. A further in-silicoanalysis through geneMANIA tool (Warde-Farley et al, 2017) to assess
potential interactions between the two proteins suggested no physical
interaction, co-expression, co-localisation, shared pathway, protein
domains or genetic interactions. The only evidence found for any links
between the two proteins is that they are both involved in the Hippo
signaling pathway, causing (similar) structural brain changes.
Another hypothesis is that this patient could be the first report of a
new syndrome or a variant of Van Maldergem syndrome, presenting with the
same clinical characteristic triad of the AS syndrome, absence of facial
dysmorphism and only the dystonic posture of the hand as the clinical
symptoms of Van Maldergem syndrome (apart from some brain anomalies
present both in AS and Van Maldergem syndrome, as periventricular
heterotopia, brain asymmetry, corpus callosum hypoplasia) (table 1).
This hypothesis may be supported by the lower pathogenicity of
c.7408G>T variant, which resulted “pathogenic” only in 7
out of 23 prediction tools. Lastly, it cannot be excluded that this
variants in DCHS1 can be an incidental finding, and the phenotype
of the girl caused by other genetic anomalies.
Further researchers of variants in DCHS1 gene in patients
affected by Aicardi or Van Maldergem syndromes may contribute to
elucidate the role of this and other genes involved in the hippo-pathway
in the pathogenesis of the diseases, together with new studies on
low-level mosaicism and balanced rearrangements, as well as platforms
examining changes at the DNA and chromatin level affecting regulatory
regions, in particular for AS (Wong et al, 2018).