Neurodegenerative diseases characterized by aberrant accumulation of undigested cellular components represent unmet medical conditions for which the identification of actionable targets is urgently needed. Here we identify a pharmacologically actionable pathway that controls cellular clearance via Akt modulation of transcription factor EB (TFEB), a master regulator of lysosomal pathways. We show that Akt phosphorylates TFEB at Ser467 and represses TFEB nuclear translocation independently of mechanistic target of rapamycin complex 1 (mTORC1), a known TFEB inhibitor. The autophagy enhancer trehalose activates TFEB by diminishing Akt activity. Administration of trehalose to a mouse model of Batten disease, a prototypical neurodegenerative disease presenting with intralysosomal storage, enhances clearance of proteolipid aggregates, reduces neuropathology and prolongs survival of diseased mice. Pharmacological inhibition of Akt promotes cellular clearance in cells from patients with a variety of lysosomal diseases, thus suggesting broad applicability of this approach. These findings open new perspectives for the clinical translation of TFEB-mediated enhancement of cellular clearance in neurodegenerative storage diseases.

Infantile neuronal ceroid lipofuscinosis (INCL, or CLN1 disease) is an inherited neurodegenerative storage disorder caused by a deficiency of the lysosomal enzyme palmitoyl protein thioesterase 1 (PPT1). It was widely believed that the pathology associated with INCL was limited to the brain, but we have now found unexpectedly profound pathology in the human INCL spinal cord. Similar pathological changes also occur at every level of the spinal cord of PPT1-deficient (Ppt1^−/−) mice before the onset of neuropathology in the brain. Various forebrain-directed gene therapy approaches have only had limited success in Ppt1^−/− mice. Targeting the spinal cord via intrathecal administration of an adeno-associated virus (AAV) gene transfer vector significantly prevented pathology and produced significant improvements in life span and motor function in Ppt1^−/− mice. Surprisingly, forebrain-directed gene therapy resulted in essentially no PPT1 activity in the spinal cord, and vice versa. This leads to a reciprocal pattern of histological correction in the respective tissues when comparing intracranial with intrathecal injections. However, the characteristic pathological features of INCL were almost completely absent in both the brain and spinal cord when intracranial and intrathecal injections of the same AAV vector were combined. Targeting both the brain and spinal cord also produced dramatic and synergistic improvements in motor function with an unprecedented increase in life span. These data show that spinal cord pathology significantly contributes to the clinical progression of INCL and can be effectively targeted therapeutically. This has important implications for the delivery of therapies in INCL, and potentially in other similar disorders. Significance Infantile neuronal ceroid lipofuscinosis (INCL, or infantile Batten disease) is a fatal childhood disease that devastates the brain. Trying to treat the brain alone has so far proved ineffective, so we looked for effects of the disease in other parts of the body. Unexpectedly, we found that the spinal cord is severely affected before the brain and contributes to disease outcome. Directing gene therapy to just the spinal cord of INCL mice can improve their disease. However, treating both the spinal cord and brain provides the most effective therapeutic strategy ever seen in this disease. This combined therapy returned INCL mice to a near-normal life span and dramatically improved their quality of life.

The neuronal ceroid lipofuscinoses are fatal neurodegenerative disorders in which the visual system is affected early in disease progression. A typical accompanying feature is neuroinflammation, the pathogenic impact of which is presently obscure. Here we investigated the role of inflammatory cells in palmitoyl protein thioesterase 1-deficient ( Ppt1 -/-) mice, a model of infantile neuronal ceroid lipofuscinosis (CLN1 disease, infantile), predominantly focusing on the visual system. We detected an early infiltration of CD8+ T-lymphocytes and observed activation of microglia/macrophage-like cells. To analyse the pathogenic impact of lymphocytes, we crossbred Ppt1 -/- mice with mutants lacking lymphocytes ( Rag1 -/-), and scored axonal transport, axonal perturbation and neuronal survival. This lack of lymphocytes led to a significant amelioration of disease phenotypes, not only in the retino-tectal system, but also in other regions of the central nervous system. Finally, reconstitution experiments revealed a crucial role of CD8+ T-lymphocytes in pathogenesis. Our study provides novel pathomechanistic insights that may be crucial for developing treatment strategies. * Abbreviation : MHC : major histocompatibility complex

Juvenile neuronal ceroid lipofuscinosis (JNCL or CLN3 disease) is an autosomal recessive lysosomal storage disease resulting from mutations in the CLN3 gene that encodes a lysosomal membrane protein. The disease primarily affects the brain with widespread intralysosomal accumulation of autofluorescent material and fibrillary gliosis, as well as the loss of specific neuronal populations. As an experimental treatment for the CNS manifestations of JNCL, we have developed a serotype rh.10 adeno-associated virus vector expressing the human CLN3 cDNA (AAVrh.10hCLN3). We hypothesized that administration of AAVrh.10hCLN3 to the Cln3 ^Δex7/8 knock-in mouse model of JNCL would reverse the lysosomal storage defect, as well as have a therapeutic effect on gliosis and neuron loss. Newborn Cln3 ^Δex7/8 mice were administered 3×10 ¹⁰ genome copies of AAVrh.10hCLN3 to the brain, with control groups including untreated Cln3 ^Δex7/8 mice and wild-type littermate mice. After 18 months, CLN3 transgene expression was detected in various locations throughout the brain, particularly in the hippocampus and deep anterior cortical regions. Changes in the CNS neuronal lysosomal accumulation of storage material were assessed by immunodetection of subunit C of ATP synthase, luxol fast blue staining, and periodic acid-Schiff staining. For all parameters, Cln3 ^Δex7/8 mice exhibited abnormal lysosomal accumulation, but AAVrh.10hCLN3 administration resulted in significant reductions in storage material burden. There was also a significant decrease in gliosis in AAVrh.10hCLN3-treated Cln3 ^Δex7/8 mice, and a trend toward improved neuron counts, compared with their untreated counterparts. These data demonstrate that AAVrh.10 delivery of a wild-type cDNA to the CNS is not harmful and instead provides a partial correction of the neurological lysosomal storage defect of a disease caused by a lysosomal membrane protein, indicating that this may be an effective therapeutic strategy for JNCL and other diseases in this category.