Clinical features and genetic studies
The proband is an 8 year-old girl with epilepsy and severe
encephalopathy. She was born at term to healthy, unrelated parents after
uneventful pregnancy and delivery. A 17 year-old half-sister is healthy.
Birth weight was 2950g (27th pctl), height 47,5 cm
(13th pctl) and head circumference 32,5 cm
(11th pctl). After birth she was noted to be
hypotonic, irritable and difficult to feed. At age 1.5 months she
developed tonic seizures featuring right arm extension and head version.
The reported physical exam showed no dysmorphic features or
organomegalies, absent visual fixation and global hypotonia. An EEG
revealed multifocal epileptiform discharges and diffuse background
slowing. Initial tests performed between ages 2 and 4 months were normal
and included brain MRI (Figure 1A ), serum amino acids, urine
organic acids, CSF folate, purines and neurotransmitter metabolites and
CSF/plasma glucose ratio. Seizures persisted daily despite treatment
with pyridoxine, phenobarbital, sodium valproate or carbamazepine and by
age 4 months manifested as a combination of tonic spasms, myoclonic
jerks and dysautonomic signs. The patient did not attain any
developmental milestone during the first year of life. Further trials
with topiramate, lamotrigine, gabapentin, clobazam, levetiracetam,
lacosamide and eslicarbacepine acetate in various combinations did not
improve her condition. She was referred to our center at age 2y 11mo.
She displayed microcephaly with HC: 44.5 cm (-3SD), right occipital
plagiocephaly and poor contact despite brief visual fixation and smiling
in reaction to voice; other findings were horizonto-rotatory nystagmus,
spastic quadriparesis, hyperreflexia, extensor plantar responses and
occasional axial or segmental myoclonia. The patient suffered on average
2-3 seizures per day, mostly generalized tonic, heralded by a flexor
spasm and accompanied by oral automatisms and eyelid myoclonia, each
lasting 1-5 minutes and was on valproate and lacosamide. EEG revealed a
high-voltage background with multifocal spikes of occipito-temporal
predominance. Ictal recording during a tonic seizure showed initial
theta-delta generalized waves with irregular temporo-occipital spikes
and a posterior global attenuation. New investigations included array
CGH, serum lactate, sialotransferrin pattern, very long chain fatty
acids, biotinidase, biochemical screen for purine and pyrimidine defects
and whole exome sequencing (WES, see below). A repeat MRI at age 4 years
showed mild enlargement of subarachnoidal spaces, a relatively thick
corpus callosum and marked cerebellar atrophy with vermian predominance
(Figure 1A ). MRS showed the presence of creatine peak. She
received therapies for increasing spasticity, including oral baclofen,
botulinum toxin injections and multilevel lower limb tenotomies. She
currently remains wheel chair bound, with little awareness of her
surroundings (Figure 1B) . Seizures persist unchanged but their
frequency has been reduced to a few per week after perampanel was
substituted for the previous treatment. Melatonin has been added for
prominent insomnia.
Whole exome sequencing detected the NM_000738.2:c.1139C>T;
NP_000729.2:p.(Pro380Leu)de novo variant in the CHRM1 gene, encoding the muscarinic
acetylcholine receptor 1, which was later validated by Sanger sequencing
(Figure 1C ). This variant modifies a highly conserved residue
in the orthologous and paralogous receptors. The p.Pro380Leu variant is
considered as damaging or potentially disease causing by variousin silico predictors and is not present in general population
databases (see Supplementary Materials and Methods). Therefore, the
p.Pro380Leu variant is considered likely pathogenic. In the recently
solved 3D structure of CHRM1 (Maeda et al., 2019), residue Pro380
determines a kink in the C terminal part of helix 6 (Figure 1D )
No additional CHRM1 variants were found by WES reanalysis in a
cohort of over 102 patients with EIEE/DEE nor any matches were produced
upon sharing the novel variants in the Matchmaker Exchange platform
(Philippakis et al., 2015).