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Page 314 Yasin et al. J Transl Genet Genom 2020;4:307-19 I https://doi.org/10.20517/jtgg.2020.30

observations below.

Developmental delay/ID and/or ASD is a primary characteristic feature in patients with PILBOS, SNIBCPS,
SIHIWES and ZFS [52,54-56] . Also, 27.5% of patients with CHARGE syndrome have been reported to have
[59]
[60]
ASD , and ID is also reported in patients with CHARGE syndrome .
Notably, in addition to neurodevelopmental and neurocognitive phenotypes such as developmental delay
and/or ID and/or ASD, patients from all four syndromes were reported to have macrocephaly. Speech
delays are noted for PILBOS, SNIBCPS and ZFS patients. Of other characteristic phenotypes found in ZFS
patients, hypotonia and some facial phenotypes such as hypertelorism and low-set ears in SNIBCPS, and
skeletal phenotypes in SIHIWES are noted. Cardiac defects are seen in SIHIWES, as they are in CHARGE
syndrome, and they have been reported in patients with ZFS. Finally, a characteristic facial gestalt is part of
the presentation of SIHIWES, SNIBPCS and ZFS but not of the other two syndromes.

Due to the discovery of thus far five of the nine CHDs being disease-causing, there has been a flourishing
of functional studies investigating pathophysiology in a number of model systems including animal and
[61]
several human cellular types. Others have comprehensively reviewed these studies for several CHDs ,
[16]
[16]
[29]
for all CHDs , for CHD2 , and for CHDs in stem cell function . As these reviews are limited in their
discussion of CHD8, we limit ourselves here to overviewing pertinent functional studies in CHD8 to fill the
lacunae.
Role of CHD8 in NDD
There were only seven publications with the key word “CHD8” when it was first discovered as a causative
[51]
gene for NDDs in 2007 . In just over the dozen years since then, a further 141 publications appear with
“CHD8” as a key word, greatly enriching our understanding of the pathogenic and functional contribution of
this gene. We note that several of these studies support an important role for CHD8 in cancer [62,63] , which is
beyond the scope of this paper. The burgeoning interest in CHD8 has no doubt been spurred by its reported
[64]
causation of up to 0.5% of all ASD . Extraordinarily, there are six different groups who have published
[71]
findings from mouse models alone [65-70] , as well as others who report functional studies with C. elegans
[72]
[73]
and Drosophila , and the first functional model study results were obtained with zebrafish . Furthermore,
a growing number of groups are exploring functional studies in human cellular models [74-77] . We will briefly
overview the main findings from these model studies below.
Human cellular model studies
[74]
To our knowledge, the first human cellular model study was presented by Sugathan et al. who used
induced pluripotent stem cell-derived neural progenitor cells (NPCs) to examine CHD8 regulation
in transcriptional networks. They knocked down CHD8 to single allele expression levels and then
examined whole transcriptome and genome-wide ChIP-seq data to derive genome-wide impacts of the
haploinsufficiency. They found that the expression of 1,756 genes was altered overall, with downregulated
genes enriched for pathways involved in brain development. In addition, a total of 5,658 genes were shown to
have possible CHD8-binding sites, and these were associated with transcriptional regulation and chromatin
modification. These data indicate widespread downstream targeting.

A later study examining CHD8 knockdown in commercial human NPCs derived from neuroblastoma
cells, discovered altered expression of both protein-coding and noncoding RNA genes, with a total of 1715
[77]
genes showing altered expression . This cited study also found that the differentially expressed genes
were enriched for neuronal development pathways and included known ASD candidate genes, similarly to
[74]
Sugathan et al. .
A separate group generated iPSC-derived NPCs from a human donor using CRISPR/Cas-9 heterozygous
[76]
gene disruption in CHD8 . They followed this by generating neurons and then brain organoids from