Activating Ras-MAPK pathway variants drive hippocampal clonal competition in human epilepsy

Mesial (<i>a.k.a.,</i> medial) temporal lobe epilepsy (MTLE) is the most common focal epilepsy<sup>1,2</sup> and, in drug-resistant cases, is treated by surgical removal of the anterior temporal lobe, which often shows neuronal loss and gliosis consistent with hippocampal sclerosis (HS)<sup>2</sup>. MTLE with HS has minimal contribution from germline genetic variation<sup>3</sup>, and is associated with prior precipitating insults such as prolonged childhood seizures and head trauma<sup>4-6</sup>. Somatic variants in Ras-MAPK pathway genes were recently reported in a few MTLE surgical specimens<sup>7,8</sup>, but their prevalence, clinical relevance, and underlying biological mechanisms remain unknown. Targeted duplex sequencing of hippocampal DNA from 462 surgical resections revealed significant enrichment of deleterious somatic variants in MTLE versus controls, with >40% of MTLE specimens harboring activating Ras-MAPK variants in <i>PTPN11</i>, <i>NF1</i>, <i>BRAF</i>, <i>KRAS</i>, and twelve genes not previously associated with focal epilepsy. Eight Ras-MAPK genes showed positive clonal selection in MTLE. Increased somatic variant burden predicted worse surgical outcome. Somatic Ras-MAPK variants at ultra-low (<0.5%) variant allele fractions were associated with older seizure onset and HS pathology, supporting a late prenatal or postnatal origin. Ras-MAPK variants in MTLE were enriched in cells derived from hippocampal progenitors-neurons, astrocytes, oligodendrocytes-in line with the known neuronal hyperexcitability and seizures induced by Ras-MAPK overactivation<sup>9,10</sup>; in contrast, Alzheimer disease hippocampi exhibited microglial enrichment of Ras-MAPK variants, consistent with prior reports<sup>11</sup>. Single-nucleus RNA sequencing showed increased expression of Ras-MAPK genes in neurons and upregulation of pathways mediating neurogenesis and neural development in MTLE. Functional validation of novel, recurrent <i>PTPN11</i> variants confirmed gain-of-function, while cellular modeling in induced pluripotent stem cells demonstrated proliferative/survival advantages for mutant cells in mosaic culture. Overall, our data suggest that somatic Ras-MAPK variants and acquired risk factors may converge on clonal competition in the hippocampus to modulate epilepsy risk.

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bioRxiv (Cold Spring Harbor Laboratory)
Published
2026-01-27
DOI
https://doi.org/10.64898/2026.01.26.701822
Primary Topic
Neuroscience and Neuropharmacology Research
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article
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article

Activating Ras-MAPK pathway variants drive hippocampal clonal competition in human epilepsy

Mingyun Bae, Michael B Miller, Rosita B Ramirez, August Yue Huang et al.
bioRxiv (Cold Spring Harbor Laboratory)
Neuroscience and Neuropharmacology Research
article

Activating Ras-MAPK pathway variants drive hippocampal clonal competition in human epilepsy

Mingyun Bae, Michael B Miller, Rosita B Ramirez, August Yue Huang, Dennis D Spencer, Zinan Zhou, James A King, Rani A Sarkis, Taufik A Valiante, Homeira Moradi Chameh, Amy E Roberts, Ingrid E Scheffer, Abbe Lai, Emily Lapinskas, Bertrand Mathon, Annapurna Poduri, Michael S Hildebrand, Kristopher T Kahle, Stephen C Blacklow, Ashton Tillett, Sofia M Zoullas, Brian Chhouk, Katherine S-M Brown, Jonathan Clark, R Mark Richardson, Samuel F Berkovic, Yilan Wang, Andrew P Morokoff, Edward Yang, Ingmar Blumcke, E Zeynep Erson-Omay, Alissa M D'Gama, Kristian Bulluss, Nihal DeLanerolle, Linus Marx, John D Rolston, Maria I Kontaridis, Terence J O'Brien, Patrick Kwan, Yasmine Chahine, Franck Bielle, Andrew Gao, Timo Krings, Luana Nunes Santos, Piero Perucca, Sanda Alexadrescu, Emily D Egan, Vincent Navarro, Eunjung Alice Lee, Christian G Bien, Roland Coras, Benjamin Finander, Samuel Gooley, Christopher A Walsh, Dipan Patel, Sattar Khoshkhoo, Marian Todaro, G Rees Cosgrove
article en

Abstract

Mesial (<i>a.k.a.,</i> medial) temporal lobe epilepsy (MTLE) is the most common focal epilepsy<sup>1,2</sup> and, in drug-resistant cases, is treated by surgical removal of the anterior temporal lobe, which often shows neuronal loss and gliosis consistent with hippocampal sclerosis (HS)<sup>2</sup>. MTLE with HS has minimal contribution from germline genetic variation<sup>3</sup>, and is associated with prior precipitating insults such as prolonged childhood seizures and head trauma<sup>4-6</sup>. Somatic variants in Ras-MAPK pathway genes were recently reported in a few MTLE surgical specimens<sup>7,8</sup>, but their prevalence, clinical relevance, and underlying biological mechanisms remain unknown. Targeted duplex sequencing of hippocampal DNA from 462 surgical resections revealed significant enrichment of deleterious somatic variants in MTLE versus controls, with >40% of MTLE specimens harboring activating Ras-MAPK variants in <i>PTPN11</i>, <i>NF1</i>, <i>BRAF</i>, <i>KRAS</i>, and twelve genes not previously associated with focal epilepsy. Eight Ras-MAPK genes showed positive clonal selection in MTLE. Increased somatic variant burden predicted worse surgical outcome. Somatic Ras-MAPK variants at ultra-low (<0.5%) variant allele fractions were associated with older seizure onset and HS pathology, supporting a late prenatal or postnatal origin. Ras-MAPK variants in MTLE were enriched in cells derived from hippocampal progenitors-neurons, astrocytes, oligodendrocytes-in line with the known neuronal hyperexcitability and seizures induced by Ras-MAPK overactivation<sup>9,10</sup>; in contrast, Alzheimer disease hippocampi exhibited microglial enrichment of Ras-MAPK variants, consistent with prior reports<sup>11</sup>. Single-nucleus RNA sequencing showed increased expression of Ras-MAPK genes in neurons and upregulation of pathways mediating neurogenesis and neural development in MTLE. Functional validation of novel, recurrent <i>PTPN11</i> variants confirmed gain-of-function, while cellular modeling in induced pluripotent stem cells demonstrated proliferative/survival advantages for mutant cells in mosaic culture. Overall, our data suggest that somatic Ras-MAPK variants and acquired risk factors may converge on clonal competition in the hippocampus to modulate epilepsy risk.

bioRxiv (Cold Spring Harbor Laboratory)
Broad Institute (US), Boston Children's Hospital (US), Beth Israel Deaconess Medical Center (US), Cleveland Clinic (US), The Royal Melbourne Hospital (AU), Royal Children's Hospital (AU), University Health Network (CA), Howard Hughes Medical Institute (US), Harvard University (US), Inserm (FR), The University of Melbourne (AU), University of Toronto (CA), Bielefeld University (DE), The Alfred Hospital (AU), Yale University (US), Sorbonne Université (FR), Massachusetts General Hospital (US), Universitätsklinikum Erlangen (DE), Masonic Medical Research Laboratory (US), Assistance Publique – Hôpitaux de Paris (FR), Evangelisches Krankenhaus Bielefeld (DE), Lahey Medical Center (US), Boston VA Research Institute (US), Dana-Farber Cancer Institute (US), Pitié-Salpêtrière Hospital (FR), Institut de Psychiatrie et Neurosciences de Paris (FR), Laboratory Krone (DE), Murdoch Children's Research Institute (AU), Epilepsy Foundation (AU), Ontario Brain Institute (CA), Alfred Health (AU), Austin Health (AU), Dana-Farber/Boston Children's Cancer and Blood Disorders Center (US), Mass General Brigham (US), Monash University (AU)
U.S. Department of Defense, Alzheimer's Association, American Heart Association, Brigham and Women's Hospital, Massachusetts General Hospital, University of Miami, University of Pittsburgh, Deutsche Forschungsgemeinschaft, National Institutes of Health, Intellectual and Developmental Disabilities Research Center, Harvard Stem Cell Institute, National Health and Medical Research Council
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