Novel splice-site and recurrent p.Arg729* CNKSR2 variants in ESES/CSWS: insights into sex-dependent expression

Abstract

Purpose: Pathogenic variants in CNKSR2 (Xp22.12) cause an X-linked neurodevelopmental disorder with intellectual disability, language impairment, and a distinctive epilepsy phenotype, including encephalopathy with status epilepticus during slow-wave sleep (ESES/CSWS). Hemizygous males are typically severely affected, whereas symptomatic females remain rare and incompletely characterized. We describe two unrelated patients with de novo CNKSR2 variants to expand the mutational and sex-dependent phenotypic spectrum of this disorder. Methods: Both patients underwent clinical, electroencephalographic, and neuroimaging evaluation. CNKSR2 variants were identified by whole exome sequencing with parental segregation, and the splice-site variant was assessed in silico (SpliceAI, MaxEntScan, Human Splicing Finder). Results: Patient 1, a 17-year-old female, harbored a novel canonical splice-site variant (c.64+1G>A) in the Nterminal region and presented with a relatively mild phenotype. In silico analysis supported abolition of the canonical donor splice site. Patient 2, an 8-year-old male, carried a de novo nonsense variant (c.2185C>T, p. Arg729*) and exhibited drug-resistant ESES, autism, and severe language impairment. p.Arg729* had previously been reported in one independent male. Our case represents its second independent occurrence, a CGA>TGA transition at a CpG dinucleotide consistent with a mutational hotspot. Conclusion: Together, these cases expand the mutational spectrum and provide further evidence for sexdependent phenotypic variability in CNKSR2-related epilepsy. Our observations support the hypothesis that Xchromosome inactivation may contribute to phenotypic variability in females, although XCI was not assessed here, and support inclusion of CNKSR2 in epilepsy gene panels regardless of sex.