Mucopolysaccharidosis VI (MPS VI) is caused by deficient arylsulfatase B (ARSB) activity resulting in lysosomal storage of glycosaminoglycans (GAGs). MPS VI is characterized by dysostosis multiplex, organomegaly, corneal clouding, and heart valve thickening. Gene transfer to a factory organ like liver may provide a lifetime source of secreted ARSB. We show that intravascular administration of adeno-associated viral vectors (AAV) 2/8-TBG-felineARSB in MPS VI cats resulted in ARSB expression up to 1 year, the last time point of the study. In newborn cats, normal circulating ARSB activity was achieved following delivery of high vector doses (6 × 10(13) genome copies (gc)/kg) whereas delivery of AAV2/8 vector doses as low as 2 × 10(12) gc/kg resulted in higher than normal serum ARSB levels in juvenile MPS VI cats. In MPS VI cats showing high serum ARSB levels, independent of the age at treatment, we observed: (i) clearance of GAG storage, (ii) improvement of long bone length, (iii) reduction of heart valve thickness, and (iv) improvement in spontaneous mobility. Thus, AAV2/ 8-mediated liver gene transfer represents a promising therapeutic strategy for MPS VI patients.

Mucopolysaccharidoses (MPSs) are lysosomal storage disorders characterized by progressive accumulation of glycosaminoglycans (GAGs) in various tissues. Enzyme replacement therapy (ERT) for several MPSs is available to date. However, the efficacy of ERT is limited, in particular in compartments such as bone, cartilage, the brain, and the eyes. We selected a rodent model of an MPS, with no central nervous system storage, to study the impact, on systemic features of the disease, of various stable levels of exogenous enzymes produced by adeno-associated viral vector (AAV)-mediated liver gene transfer. Low levels (6% of normal) of circulating enzyme were enough to reduce storage and inflammation in the visceral organs and to ameliorate skull abnormalities; intermediate levels (11% of normal) were required to reduce urinary GAG excretion; and high levels (>or=50% of normal) rescued abnormalities of the long bones and motor activity. These data will be instrumental to design appropriate clinical protocols based on either enzyme or gene replacement therapy for MPS and to predict their impact on the pathological features of MPS.

Autophagy and energy metabolism are known to follow a circadian pattern. However, it is unclear whether autophagy and the circadian clock are coordinated by common control mechanisms. Here, we show that the oscillation of autophagy genes is dependent on the nutrient‐sensitive activation of TFEB and TFE3, key regulators of autophagy, lysosomal biogenesis, and cell homeostasis. TFEB and TFE3 display a circadian activation over the 24‐h cycle and are responsible for the rhythmic induction of genes involved in autophagy during the light phase. Genetic ablation of TFEB and TFE3 in mice results in deregulated autophagy over the diurnal cycle and altered gene expression causing abnormal circadian wheel‐running behavior. In addition, TFEB and TFE3 directly regulate the expression of Rev‐erbα (Nr1d1), a transcriptional repressor component of the core clock machinery also involved in the regulation of whole‐body metabolism and autophagy. Comparative analysis of the cistromes of TFEB/TFE3 and REV‐ERBα showed an extensive overlap of their binding sites, particularly in genes involved in autophagy and metabolic functions. These data reveal a direct link between nutrient and clock‐dependent regulation of gene expression shedding a new light on the crosstalk between autophagy, metabolism, and circadian cycles.

Ammonia is a potent neurotoxin that is detoxified mainly by the urea cycle in the liver. Hyperammonemia is a common complication of a wide variety of both inherited and acquired liver diseases. If not treated early and thoroughly, it results in encephalopathy and death. Here, we found that hepatic autophagy is critically involved in systemic ammonia homeostasis by providing key urea-cycle intermediates and ATP. Hepatic autophagy is triggered in vivo by hyperammonemia through an α-ketoglutarate-dependent inhibition of the mammalian target of rapamycin complex 1, and deficiency of autophagy impairs ammonia detoxification. In contrast, autophagy enhancement by means of hepatic gene transfer of the master regulator of autophagy transcription factor EB or treatments with the autophagy enhancers rapamycin and Tat-Beclin-1 increased ureagenesis and protected against hyperammonemia in a variety of acute and chronic hyperammonemia animal models, including acute liver failure and ornithine transcarbamylase deficiency, the most frequent urea-cycle disorder. In conclusion, hepatic autophagy is an important mechanism for ammonia detoxification because of its support of urea synthesis, and its enhancement has potential for therapy of both primary and secondary causes of hyperammonemia.