Authors :
Presenting Author: Sattar Khoshkhoo, MD – Brigham and Women's Hospital
Yilan Wang, BS – Boston Children's Hospital; Yasmine Chahine, BS – Boston Children's Hospital; Ashton Tillett, BS – Brigham and Women's Hospital; August Yue Huang, PhD – Boston Children's Hospital; Stephanie Robert, MD, PhD – Yale School of Medicine; Brian Chhouk, BS – Boston Children's Hospital; Emre Kiziltug, BS – Yale School of Medicine; Carol Nelson-Williams, MS – Yale School of Medicine; Edward Stronge, BS – Boston Children's Hospital; H Phillips, MD – Stanford School of Medicine; Thiuni Adikari, BMSc (Hons) – University of Melbourne; Zimeng Ye, MS – University of Melbourne; Tom Witkowski, BSc (Hons) – University of Melbourne; Julie Lokan, MBBS – Austin Health; Dulcie Lai, PharmD, PhD – University of North Carolina, Chapel Hill; Edward Yang, MD, PhD – Boston Children's Hospital; Erin Heinzen, PharmD, PhD – University of North Carolina, Chapel Hill; Eyiyemisi Damisah, MD – Yale School of Medicine; Homeira Moradi, PhD – Krembil Brain Institute, University Health Network; Ronald Coras, MD – University Hospitals Erlangen; Bertrand Mathon, MD, PhD – Sorbonne Université, ICM, Salpêtrière Hospital; Vincent Navarro, MD – Sorbonne Université, ICM, Salpêtrière Hospital; Franck Bielle, MD, PhD – Sorbonne Université, ICM, Salpêtrière Hospital; Sanda Alexandrescu, MD – Boston Children's Hospital; Anita Huttner, MD – Yale School of Medicine; Ingrid Scheffer, MBBS, PhD – University of Melbourne; Samuel Berkovic, MD – University of Melbourne; Michael Hildebrand, PhD – University of Melbourne; Annapurna Poduri, MD, MPH – Boston Children's Hospital; Nihal DeLanerolle, DPhil, DSc – Yale School of Medicine; Dennis Spencer, MD – Yale School of Medicine; Taufik Valiante, MD, PhD – Krembil Brain Institute, University Health Network; Ingmar Blümcke, MD – University Hospitals Erlangen; Kristopher Kahle, MD, PhD – Massachusetts General Hospital; Eunjung Lee, PhD – Boston Children's Hospital; Christopher Walsh, MD, PhD – Boston Children's Hospital/HHMI
Rationale: Focal epilepsies which are associated with common lesions such as malformations of cortical development (MCD) and mesial temporal sclerosis (MTS), are frequently drug-resistant and require neurosurgical treatment in a about a third of patients, with variable outcomes and attendant morbidity. Post-zygotic mutations (i.e., somatic variants) are now a well-recognized cause of MCD, which are typically seen in children. However, the etiology of MTS which is the histopathologic hallmark of mesial temporal lobe epilepsy (MTLE) and typically seen in adults is still debated. Previously, using whole-exome sequencing we detected 10 pathogenic somatic variants activating Ras-MAPK signaling in 105 patients with MTLE and none in the controls. In this study, we investigated the extent to which low-abundance somatic Ras-MAPK variants in the hippocampus contribute to drug-resistant MTLE.
Methods: To significantly improve the sensitivity and specificity of detection for low-abundance somatic variants, we performed duplex sequencing on gene-panel enriched hippocampus-derived DNA (depth >1000X) from 218 surgically-treated MTLE patients and 56 neurotypical individuals. The gene-panel included a mix of Ras-MAPK genes, MCD genes (e.g., mTOR pathway), and common developmental and epileptic encephalopathy genes. A subset of the novel MTLE-associated somatic variants were evaluated experimentally using molecular assays.
Results: Gene-panel sequencing significantly improved the somatic variant detection sensitivity with 98 pathogenic somatic Ras-MAPK variants identified in 81 patients with MTLE (37% of 218) and no neurotypical controls which was statistically significant (p< 1e-5). This included variants in the FGFR1, PTPN11, SOS1, KRAS, NRAS,, BRAF, CBL, NF1, and LZTR1 genes. In comparison, only four variants were detected in MCD genes and one variant in developmental and epileptic encephalopathy genes, indicating a specific and likely causal association between pathogenic Ras-MAPK variants and MTLE. Strikingly, most Ras-MAPK variants (n=92, 93%) had allele frequencies < 1% which suggests a late gestational or early postnatal origin. All the patients with Ras-MAPK pathogenic variants had evidence of MTS on histopathology and only < 10% of the cases were associated with a secondary MCD or a low-grade epilepsy-associated tumor. Some of the novel, recurrent MTLE-associated SHP2 (protein encoded by
PTPN11) variants were examined using
in vitro assays that demonstrated activation of downstream Ras-MAPK signaling.
Conclusions: Somatic variants activating Ras-MAPK signaling likely cause MTLE in a significant subset of patients with sporadic, drug-resistant disease. Given the association of Ras-MAPK variants with MTS and improved surgical outcomes, somatic variants may be an important and clinically relevant biomarker that should be incorporated into the routine evaluation of patients with drug-resistant MTLE.
Funding: NINDS K08 Award K08-NS128272. Doris Duke Physician Scientist Fellowship