Phenotypic and genetic spectrum of ATP6V1A encephalopathy: a disorder of lysosomal homeostasis
Authors: Guerrini Renzo , Mei Davide , Kerti-Szigeti Katalin , Pepe Sara , Koenig Mary Kay , Von Allmen Gretchen , Cho Megan T , McDonald Kimberly , Baker Janice , Bhambhani Vikas , Powis Zöe , Rodan Lance , Nabbout Rima , Barcia Giulia , Rosenfeld Jill A , Bacino Carlos A , Mignot Cyril , Power Lillian H , Harris Catharine J , Marjanovic Dragan , Møller Rikke S , Hammer Trine B ,   The DDD Study, Keski Filppula Riikka , Vieira Päivi , Hildebrandt Clara , Sacharow Stephanie ,   Undiagnosed Diseases Network, Maragliano Luca , Benfenati Fabio , Lachlan Katherine , Benneche Andreas , Petit Florence , de Sainte Agathe Jean Madeleine , Hallinan Barbara , Si Yue , Wentzensen Ingrid M , Zou Fanggeng , Narayanan Vinodh , Matsumoto Naomichi , Boncristiano Alessandra , la Marca Giancarlo , Kato Mitsuhiro , Anderson Kristin , Barba Carmen , Sturiale Luisa , Garozzo Domenico , Bei Roberto ,   ATP6V1A collaborators, Masuelli Laura , Conti Valerio , Novarino Gaia , Fassio Anna 
Published: April 2022

Abstract
Vacuolar-type H+-ATPase (V-ATPase) is a multimeric complex present in a variety of cellular membranes that acts as an ATP-dependent proton pump and plays a key role in pH homeostasis and intracellular signalling pathways. In humans, 22 autosomal genes encode for a redundant set of subunits allowing the composition of diverse V-ATPase complexes with specific properties and expression. Sixteen subunits have been linked to human disease.
Here we describe 26 patients harbouring 20 distinct pathogenic de novo missense ATP6V1A variants, mainly clustering within the ATP synthase α/β family-nucleotide-binding domain. At a mean age of 7 years (extremes: 6 weeks, youngest deceased patient to 22 years, oldest patient) clinical pictures included early lethal encephalopathies with rapidly progressive massive brain atrophy, severe developmental epileptic encephalopathies and static intellectual disability with epilepsy. The first clinical manifestation was early hypotonia, in 70%; 81% developed epilepsy, manifested as developmental epileptic encephalopathies in 58% of the cohort and with infantile spasms in 62%; 63% of developmental epileptic encephalopathies failed to achieve any developmental, communicative or motor skills. Less severe outcomes were observed in 23% of patients who, at a mean age of 10 years and 6 months, exhibited moderate intellectual disability, with independent walking and variable epilepsy. None of the patients developed communicative language. Microcephaly (38%) and amelogenesis imperfecta/enamel dysplasia (42%) were additional clinical features. Brain MRI demonstrated hypomyelination and generalized atrophy in 68%. Atrophy was progressive in all eight individuals undergoing repeated MRIs.
Fibroblasts of two patients with developmental epileptic encephalopathies showed decreased LAMP1 expression, Lysotracker staining and increased organelle pH, consistent with lysosomal impairment and loss of V-ATPase function. Fibroblasts of two patients with milder disease, exhibited a different phenotype with increased Lysotracker staining, decreased organelle pH and no significant modification in LAMP1 expression. Quantification of substrates for lysosomal enzymes in cellular extracts from four patients revealed discrete accumulation. Transmission electron microscopy of fibroblasts of four patients with variable severity and of induced pluripotent stem cell-derived neurons from two patients with developmental epileptic encephalopathies showed electron-dense inclusions, lipid droplets, osmiophilic material and lamellated membrane structures resembling phospholipids. Quantitative assessment in induced pluripotent stem cell-derived neurons identified significantly smaller lysosomes.
ATP6V1A-related encephalopathy represents a new paradigm among lysosomal disorders. It results from a dysfunctional endo-lysosomal membrane protein causing altered pH homeostasis. Its pathophysiology implies intracellular accumulation of substrates whose composition remains unclear, and a combination of developmental brain abnormalities and neurodegenerative changes established during prenatal and early postanal development, whose severity is variably determined by specific pathogenic variants.