Authors :
Michael Hildebrand, PhD – University of Melbourne; Zimeng Ye, MS – Department of Medicine, Austin Health – University of Melbourne; Mark Bennett, PhD – Population Health and Immunity Division – The Walter and Eliza Hall Institute of Medical Research; Andrew Neal, MBBS, PhD – Department of Neurology, Alfred Hospital – Monash University; Joshua Laing, MBBS – Department of Neurology, Alfred Hospital – Monash University; Martin Hunn, MBChB – Department of Neurosurgery, Alfred Hospital – Monash University; Thanomporn Wittayacharoenpong, MS BioMedEng – Department of Neurology, Alfred Hospital – Monash University; Marian Todaro, PhD – Department of Neurology, Alfred Hospital – Monash University; Sheila Patel, PhD – Department of Medicine, Austin Health – University of Melbourne; Melanie Bahlo, PhD – Population Health and Immunity Division – The Walter and Eliza Hall Institute of Medical Research; Patrick Kwan, MD, PhD – Department of Neurology, Alfred Hospital – Monash University; Terrence O'Brien, MD – Department of Neurology, Alfred Hospital – Monash University; Ingrid Scheffer, MBBS, PhD – Department of Medicine, Austin Health – University of Melbourne; Samuel Berkovic, MD – Department of Medicine, Austin Health – University of Melbourne; Piero Perucca, MD, PhD – Department of Medicine, Austin Health – University of Melbourne
Rationale: Mosaic pathogenic variants restricted to brain are increasingly recognized as a cause of focal epilepsies. We aimed to identify a mosaic pathogenic variant and its anatomical gradient in brain DNA derived from trace tissue on explanted stereo-electroencephalography (SEEG) electrodes.
Methods: We studied a patient with non-lesional multifocal epilepsy undergoing pre-surgical evaluation with SEEG. Following explantation, electrodes were divided into 3 pools based on their brain location (right posterior quadrant, left posterior quadrant, hippocampus/temporal neocortex).
Tissue from each pool was processed and DNA whole genome amplified prior to high-depth exome sequencing. Droplet digital PCR was performed to validate mosaicism. Brain-specific GFAP protein assay enabled cell-of-origin analysis.
Results: We demonstrated a mosaic gradient for a novel pathogenic KCNT1 loss-of-function variant, c.530G >A, p.W177X, predicted to lead to nonsense-mediated decay. Strikingly, the mosaic gradient correlated strongly with the SEEG findings as the highest mutant allele fraction was in the right posterior quadrant, reflecting the most epileptogenic region on EEG studies. Elevated GFAP level indicated enrichment of brain-derived cells in SEEG cell suspension.
Conclusions: This study demonstrates proof-of-concept that mosaic gradients of pathogenic variants can be established using trace tissue from explanted SEEG electrodes.
Funding: This study was funded by the National Health and Medical Research Council Australia, The Medical Research Future Fund Australia, and Sir Weary Dunlop Medical Research Foundation Australia. This work was also made possible through the Victorian State Government Operational Infrastructure Support and Australian Government National Health and Medical Research Council (NHMRC) independent research Institute Infrastructure Support Scheme (IRIISS). MFB was supported by a Taking Flight Award from CURE Epilepsy. MB was supported by an NHMRC Investigator Grant (1195236). PK was supported by a MRFF Practitioner Fellowship (MRF1136427). IES was supported by a NHMRC Investigator Grant (1172897). SFB was supported by an Investigator Grant (1196637). TJO was supported by a NHMRC Investigator Grant (APP1176426). MFB, IES, SFB, PP and MSH were supported by an MRFF Genomics Health Futures Mission Grant (2007707). SFB and MSH were supported by a NHMRC Project Grant (1129054). PP was supported by an NHMRC Early Career Fellowship (APP1163708), the Epilepsy Foundation, The University of Melbourne, Monash University, Brain Australia, and the Weary Dunlop Medical Research Foundation.