The genomic landscape across 474 surgically accessible epileptogenic human brain lesions

Abstract

Understanding the exact molecular mechanisms involved in the etiology of epileptogenic pathologies with or without tumor activity is essential for improving treatment of drug-resistant focal epilepsy. Here, we characterize the landscape of somatic genetic variants in resected brain specimens from 474 individuals with drug-resistant focal epilepsy using deep whole-exome sequencing (>350×) and whole-genome genotyping. Across the exome, we observe a greater number of somatic single-nucleotide variants (SNV) in low-grade epilepsy-associated tumors (LEAT; 7.92 ± 5.65 SNV) than in brain tissue from malformations of cortical development (MCD; 6.11 ± 4 SNV) or hippocampal sclerosis (HS; 5.1 ± 3.04 SNV). Tumor tissues also had the largest number of likely pathogenic variant carrying cells. LEAT had the highest proportion of samples with one or more somatic copy number variants (CNV; 24.7%), followed by MCD (5.4%) and HS (4.1%). Recurring somatic whole chromosome duplications affecting Chromosome 7 (16.8%), chromosome 5 (10.9%), and chromosome 20 (9.9%) were observed among LEAT. For germline variant-associated MCD genes such as TSC2, DEPDC5, and PTEN, germline SNV were frequently identified within large loss of heterozygosity regions, supporting the recently proposed ‘second hit’ disease mechanism in these genes. We detect somatic variants in twelve established lesional epilepsy genes and demonstrate exome-wide statistical support for three of these in the etiology of LEAT (e.g., BRAF) and MCD (e.g., SLC35A2 and MTOR). We also identify novel significant associations for PTPN11 with LEAT and NRAS Q61 mutated protein with a complex MCD characterized by polymicrogyria and nodular heterotopia. The variants identified in NRAS are known from cancer studies to lead to hyperactivation of NRAS, which can be targeted pharmacologically. We identify large recurrent 1q21-q44 duplication including AKT3 in association with focal cortical dysplasia type 2a with hyaline astrocytic inclusions, another rare and possibly under-recognized brain lesion. The clinical genetic analyses showed that the numbers of somatic SNV across the exome and the fraction of affected cells were positively correlated with the age at seizure onset and surgery in individuals with LEAT. In summary, our comprehensive genetic screen sheds light on the genome-scale landscape of genetic variants in epileptic brain lesions, informs the design of gene panels for clinical diagnostic screening, and guides future directions for clinical implementation of epilepsy surgery genetics.

Alterations inPTPN11and other RAS-/MAP-Kinase pathway genes define ganglioglioma with adverse clinical outcome and atypic histopathological features

“…One-hundred-thirty pre-selected samples of LEAT with uncharacteristic histopathology and clinical features were collected at the University Hospital in Erlangen, Germany; Klinikum Bethel-Mara, Bielefeld University, Germany; and Schoen-Klinik Vogtareuth, Germany. Seventy-two samples were snap frozen at -80°C and submitted to whole-exome sequencing and Single-Nucleotide-Polymorphism analysis (SNP) with Global Screening Array Microarrays v1.0 (Illumina, San Diego, CA, USA) as described elsewhere (Lopez-Rivera et al, 2022). Eighty-six samples were formalin-fixed and paraffin-embedded (FFPE) and submitted to DNA methylation analysis using 450K and 850K/EPIC arrays (Illumina, California, USA).…”

“…Following routine DNA extraction from the frozen tissue samples, whole-exome sequencing (WES) was performed at a coverage of >350x using Agilent SureSelect Human All Exon V7 enrichment and paired-end reads (151bp) Illumina sequencing. Paired-end FASTQ files were pre-processed following GATK best practices (DePristo et al, 2011) and genetic variants were identified as described previously (Lopez-Rivera et al, 2022). These samples were also genotyped using the Global Screening Array with Multi-disease drop-in (GSA-MD v1.0; Illumina, San Diego, CA, USA).…”

“…One case was compatible with the diagnosis of a multinodular and vacuolating tumor (MNVT; case #14). Quality assessment and significance levels of the genomic data were determined as described previously (Lopez-Rivera et al, 2022).…”

“…Protein Tyrosine Phosphatase Non-receptor Type 11 ( PTPN11 ) has been recently discovered as a new candidate gene in brain tissue obtained from drug-resistant structural epilepsies (Lopez-Rivera et al, 2022, Bedrosian et al, 2022). The PTPN11 gene encodes for an early non-receptor-type protein tyrosine phosphatase SHP-2 (src homology region 2-domain phosphatase-2) of the RAS-/MAP-Kinase pathway.…”

“…We recently identified an accumulation of PTPN11 alterations in low-grade epilepsy-associated developmental brain tumors (LEAT) (Lopez-Rivera et al, 2022), the second largest lesion category in drug-resistant focal epilepsies amenable to neurosurgical treatment (Blumcke et al, 2017, Lamberink et al, 2020). Ganglioglioma (GG) account for approx.…”