Abstracts

Rationale: Recent studies have reported somatic mutations causing Rat Sarcoma (RAS) pathway activation in patients with mesial temporal lobe epilepsy (mTLE). Approximately half of these patients have a history of febrile convulsions, during the post-natal period when hippocampal neurogenesis is active. However, the relationship between somatic mosaicisms and hippocampal neurogenesis remains elusive. Our study aims to discover somatic variants in the neurogenic niche to uncover this relationship and elucidate the pathogenesis of mTLE.

Methods: Our population consists of 40 mTLE patients with a history of surgery, 34 of whom have matched blood samples while 6 do not. We obtained DNA using laser capture microdissection from neurogenic niche in dentate gyrus (DG) and cornus ammonis (CA). Due to the limited area of interest, the amount of DNA was inevitably small; hence, we utilized low-input DNA sequencing, which employs enzymatic fragmentation rather than acoustic fragmentation. We used FilterMutectCall and MosaicForecast following Mutect2 to call somatic variants and created an additional pipeline with SOBDetector to filter Formalin-Fixed Paraffin-Embedded (FFPE) artifacts, as the hippocampus samples had been stored as FFPE for several years.

Results: We performed deep whole exome sequencing and achieved approximately 500X depth of coverage in both regions. Overall, most of somatic variants were cytosine to thymine substitutions and were assigned to single base substitution (SBS) signature 1 and 5, consistent with previous study on focal cortical dysplasia published in Nat Genet 2023; 55.2(209-220). SBS signature 1 and 5, closely related to proliferation and stem cell activity, were more enriched in DG without statistical significance, supporting the neurogenic niche property of DG over CA. We discovered RAS-activating somatic variants in 12 patients out of 40 patients: gain of function mutations in oncogenes such as BRAF and FGFR2, and loss of function mutations in tumor suppressor genes such as NF1 and CBL. Patients with RAS-activating somatic variants limited to DG had a history of febrile convulsions, with pathology limited to the hippocampus (Figure 1). In contrast, the remaining patients showed inconsistency in the history of febrile convulsions or pathology outside the hippocampus, such as in the temporal cortex. Patients with RAS-activating somatic variants showed hypermutation in total mutation burden, a pattern that was more pronounced when comparing SBS signatures 1 and 5 as well as signature 10b, known for hypermutation (Figure 2).

Conclusions: Our findings support that variants limited to DG indicate post-natal somatic mosaicism, while other variants indicate post-zygotic somatic mosaicism. In conclusion, we propose that RAS-activating somatic mosaicism in neurogenic niche could cause the pathology of mTLE through post-natal hippocampal neurogenesis.

Funding: This research was supported by a grant of the MD-Phd/Medical Scientist Training Program through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea