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Acid sphingomyelinase deficiency (ASMD) is a lysosomal storage disorder caused by mutations in the gene-encoding acid sphingomyelinase (ASM). ASMD impacts peripheral organs in all patients, including the liver and spleen. The infantile and chronic neurovisceral forms of the disease also lead to neuroinflammation and neurodegeneration for which there is no effective treatment. Cellular accumulation of sphingomyelin (SM) is a pathological hallmark in all tissues. SM is the only sphingolipid comprised of a phosphocholine group linked to ceramide. Choline is an essential nutrient that must be obtained from the diet and its deficiency promotes fatty liver disease in a process dependent on ASM activity. We thus hypothesized that choline deprivation could reduce SM production and have beneficial effects in ASMD. Using acid sphingomyelinase knock-out (ASMko) mice, which mimic neurovisceral ASMD, we have assessed the safety of a choline-free diet and its effects on liver and brain pathological features such as altered sphingolipid and glycerophospholipid composition, inflammation and neurodegeneration. We found that the choline-free diet was safe in our experimental conditions and reduced activation of macrophages and microglia in the liver and brain, respectively. However, there was no significant impact on sphingolipid levels and neurodegeneration was not prevented, arguing against the potential of this nutritional strategy to assist in the management of neurovisceral ASMD patients.

Supplementary Materials and Methods, Figure Legends 1-3 from Nitric Oxide Boosts Chemoimmunotherapy via Inhibition of Acid Sphingomyelinase in a Mouse Model of Melanoma

Supplementary Figure 2 from Nitric Oxide Boosts Chemoimmunotherapy via Inhibition of Acid Sphingomyelinase in a Mouse Model of Melanoma

Cisplatin is one of the most effective anticancer drugs, but its severe toxic effects, including depletion of immune-competent cells, limit its efficacy. We combined the systemic treatment with cisplatin with intratumor delivery of dendritic cells (DC) previously treated ex vivo with a pulse of nitric oxide (NO) released by the NO donors (z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino]-diazen-1-ium-1,2-diolate or isosorbide dinitrate. We found that this chemoimmunotherapy, tested in the B16 mouse model of melanoma, was significantly more efficacious than cisplatin alone, leading to tumor regression and animal survival at low doses of cisplatin that alone had no effect. Tumor cure was not observed when combining cisplatin with DCs not exposed to NO donors, indicating the key role of the pretreatment with NO. We investigated the mechanisms responsible for the synergic effect of NO-treated DCs and cisplatin and found that NO-treated DCs were protected both in vitro and in vivo from cisplatin-induced cytotoxicity. Cisplatin triggered DC apoptosis through increased expression and activation of acid sphingomyelinase; pretreatment of DCs with NO donors prevented such activation and inhibited activation of the downstream proapoptotic events, including generation of ceramide, activation of caspases 3 and 9, and mitochondrial depolarization. The effects of NO were mediated through generation of its physiologic messenger, cyclic GMP. We conclude that NO and NO generating drugs represent promising tools to increase the efficacy of chemoimmunotherapies in vivo, promoting the survival and increasing the function of injected cells by targeting a key pathway in cisplatin-induced cytotoxicity. [Cancer Res 2007;67(16):7559–64]

Supplementary Figure 3 from Nitric Oxide Boosts Chemoimmunotherapy via Inhibition of Acid Sphingomyelinase in a Mouse Model of Melanoma

Supplementary Figure 1 from Nitric Oxide Boosts Chemoimmunotherapy via Inhibition of Acid Sphingomyelinase in a Mouse Model of Melanoma

Silicosis is a serious occupational disease with the highest incidence in China. However, its pathogenesis has not been fully elucidated. Studies have shown that the sphingomyelin signaling pathway may play an important role in different fibrotic diseases but its role in silicosis-mediated fibrosis is still unclear. In this study, the supernatant of human peripheral blood mononuclear cell line (THP-1)-derived macrophages exposed to silica (SiO2 ) was used to stimulate the transformation of human embryonic lung fibroblast cell line (HFL-1) into myofibroblasts, and the intervention effect of recombinant human acid ceramidase (rAC) was observed. The results showed that SiO2 stimulated the production of reactive oxygen species and malondialdehyde in the supernatant of THP-1-derived macrophages and increased the secretion of TGF-β1, TNF-α, and IL-8. In addition, we found that the expression levels of α-SMA, FN, Col I, and Col III in HFL-1 cells increased. Meanwhile, the activities of ASMase and ACase and the expression levels of Cer, Sph, and S1P were increased. Intervention by rAC can suppress these changes to different degrees. In conclusion, the present study shows that SiO2 dust poisoning may stimulate HFL-1 cell differentiation into myofibroblasts by inducing oxidative stress in THP-1-derived macrophages, thereby promoting the secretion of a variety of inflammatory factors and activating the sphingolipid signaling pathway in HFL-1 cells. Exogenous rAC can effectively interfere with the stimulation of HFL-1 cells by silica in vitro.

Sphingosine kinase1 (SphK1) is an acetyl-CoA dependent acetyltransferase which acts on cyclooxygenase2 (COX2) in neurons in a model of Alzheimer’s disease (AD). However, the mechanism underlying this activity was unexplored. Here we show that N-acetyl sphingosine (N-AS) is first generated by acetyl-CoA and sphingosine through SphK1. N-AS then acetylates serine 565 (S565) of COX2, and the N-AS-acetylated COX2 induces the production of specialized pro-resolving mediators (SPMs). In a mouse model of AD, microglia show a reduction in N-AS generation, leading to decreased acetyl-S565 COX2 and SPM production. Treatment with N-AS increases acetylated COX2 and N-AS-triggered SPMs in microglia of AD mice, leading to resolution of neuroinflammation, an increase in microglial phagocytosis, and improved memory. Taken together, these results identify a role of N-AS in the dysfunction of microglia in AD., Neuronal sphingosine kinase 1 (SphK1) acetylates COX2 which is needed for microglial phagocytosis activity, and release of pro-resolving mediators (SPMs) from neurons. Here the authors examine how SphK1-mediates COX2 acetylation, and how this leads to increased secretion of SPMs from neurons in the context of Alzheimer’s disease models.

Background Acid sphingomyelinase deficiency (ASMD), a rare lysosomal storage disease, results from mutations in SMPD1, the gene encoding acid sphingomyelinase (ASM). As a result, sphingomyelin accumulates in multiple organs including spleen, liver, lung, bone marrow, lymph nodes, and in the most severe form, in the CNS and peripheral nerves. Clinical manifestations range from rapidly progressive and fatal infantile neurovisceral disease, to less rapidly progressing chronic neurovisceral and visceral forms that are associated with significant morbidity and shorter life span due to respiratory or liver disease. Objectives To provide a contemporary guide of clinical assessments for disease monitoring and symptom management across the spectrum of ASMD phenotypes. Methods An international group of ASMD experts in various research and clinical fields used an evidence-informed consensus process to identify optimal assessments, interventions, and lifestyle modifications. Results Clinical assessment strategies for major organ system involvement, including liver, spleen, cardiovascular, pulmonary, and neurological/developmental are described, as well as symptomatic treatments, interventions, and/or life style modifications that may lessen disease impact. Conclusions There is currently no disease-specific treatment for ASMD, although enzyme replacement therapy with a recombinant human ASM (olipudase alfa) is in clinical development. Current monitoring addresses symptoms and multisystem involvement. Recommended interventions and lifestyle modifications are designed to address morbidity and disease complications and improve patient quality of life. While infantile neurovisceral ASMD is uniformly fatal in early childhood, patients with chronic visceral and chronic neurovisceral ASMD require appropriate management throughout childhood and adulthood by an interdisciplinary clinical team.

Mucopolysaccharidosis type IIIA (MPS IIIA) is a lysosomal storage disorder due to the deficient activity of sulfamidase (SGSH). Traditionally, measurement of this enzymatic activity has been performed using a fluorescently (4-MU) labeled glycoside substrate. While this substrate is inexpensive and readily available, the current method requires a 2-step procedure that is performed over 2 days. Here we report a new and simplified procedure using the 4-MU substrate. Major advantages of this assay method over the existing fluorescent method include a single step vs. 2-step procedure, an incubation time of 1 hour, and high sensitivity. The reaction is also run on UPLC equipment, which is available in most research labs and permits separation of the endogenous, autofluorescent material from the 4-MU signal. This assay method was developed using the MPS IIIA mouse model, and was validated using mouse plasma, liver and brain extracts, and dried blood spots. Human MPS IIIA skin fibroblasts and dried blood spots also were used to validate the method.

Silicosis is a serious occupational disease with the highest incidence in China. However, its pathogenesis has not been fully elucidated. Studies have shown that the sphingomyelin signaling pathway may play an important role in different fibrotic diseases but its role in silicosis-mediated fibrosis is still unclear. In this study, the supernatant of human peripheral blood mononuclear cell line (THP-1)-derived macrophages exposed to silica (SiO

Ceramide is a bioactive signaling lipid involved in the pathogenesis of numerous diseases. It also plays an important role in ischemia reperfusion (IR) injury via activation of inflammatory/oxidative stress-stimulated signaling pathways, resulting in tissue damage. Acid ceramidase is a lipid hydrolase that modulates the levels of ceramide, and as such has a potential therapeutic role in many human diseases where ceramide has been implicated. Here we investigated the therapeutic potential of recombinant acid ceramidase in a murine model of hepatic IR injury. Serum ALT, AST, and LDH activities, as well as oxidative stress (MDA) and inflammatory (MCP-1) markers, were increased in mice subjected to IR compared to a sham group. In contrast, these elevations were significantly lower in an IR group pretreated with a single injection of acid ceramidase. Histological examination by two different assessment criteria also revealed that acid ceramidase pretreatment alleviated IR-induced hepatocyte damage, including reduced evidence of cell death and necrosis. In addition, elevated ceramide and sphingosine levels were observed in the IR group compared to sham, and were markedly reduced when pretreated with acid ceramidase. In contrast, the levels of the protective signaling lipid, sphingosine-1-phosphate (S1P), were reduced following IR and elevated in response to acid ceramidase pretreatment. These changes in sphingolipid levels could be correlated with changes in the activities of several sphingolipid-metabolizing enzymes. Overall, these results indicated that sphingolipid changes were an important pathologic component of hepatic IR injury, and that acid ceramidase administration ameliorated these lipid changes and other downstream pathologic changes.

Rationale: In cystic fibrosis the major cause of morbidity and mortality is lung disease characterized by inflammation and infection. The influence of sphingolipid metabolism is poorly understood with a lack of studies using human airway model systems. Objectives: To investigate sphingolipid metabolism in cystic fibrosis and the effects of treatment with recombinant human acid ceramidase on inflammation and infection. Methods: Sphingolipids were measured using mass spectrometry in fully-differentiated cultures of primary human airway epithelial cells and co-cultures with Pseudomonas aeruginosa. In situ activity assays, Western blotting and quantitative polymerase chain reaction were used to investigate function and expression of ceramidase and sphingomyelinase. Effects of treatment with recombinant human acid ceramidase on sphingolipid profile and inflammatory mediator production were assessed in cell cultures and murine models. Measurements and Main Results: Ceramide is increased in cystic fibrosis airway epithelium due to differential function of enzymes regulating sphingolipid metabolism. Sphingosine, a metabolite of ceramide with antimicrobial properties, is not upregulated in response to Pseudomonas aeruginosa by cystic fibrosis airway epithelia. Tumor necrosis factor receptor 1 is increased in the apical membrane of cystic fibrosis epithelia and activates NF-κB signaling, generating inflammation. Treatment with recombinant human acid ceramidase, to decrease ceramide, reduced both inflammatory mediator production and susceptibility to infection. Conclusions: Sphingolipid metabolism is altered in airway epithelial cells cultured from people with cystic fibrosis. Treatment with recombinant acid ceramidase ameliorates the two pivotal features of cystic fibrosis lung disease, inflammation and infection, and thus represents a therapeutic approach worthy of further exploration. This article is open access and distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives License 4.0 (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Acid sphingomyelinase deficiency (ASMD) leads to cellular accumulation of sphingomyelin (SM), neurodegeneration, and early death. Here, we describe the downregulation of the endocannabinoid (eCB) system in neurons of ASM knockout (ASM‐KO) mice and a ASMD patient. High SM reduced expression of the eCB receptor CB1 in neuronal processes and induced its accumulation in lysosomes. Activation of CB1 receptor signaling, through inhibition of the eCB‐degrading enzyme fatty acid amide hydrolase (FAAH), reduced SM levels in ASM‐KO neurons. Oral treatment of ASM‐KO mice with a FAAH inhibitor prevented SM buildup; alleviated inflammation, neurodegeneration, and behavioral alterations; and extended lifespan. This treatment showed benefits even after a single administration at advanced disease stages. We also found CB1 receptor downregulation in neurons of a mouse model and a patient of another sphingolipid storage disorder, Niemann–Pick disease type C (NPC). We showed the efficacy of FAAH inhibition to reduce SM and cholesterol levels in NPC patient‐derived cells and in the brain of a NPC mouse model. Our findings reveal a pathophysiological crosstalk between neuronal SM and the eCB system and offer a new treatment for ASMD and other sphingolipidoses., We are grateful to Steven Laffoon and the Wylder Nation Foundation (www.wyldernation.org) for the continuous support. We thank Dr. Melissa Wasserstein at the Children's Hospital at Montefiore for insightful comments and Dr. A. Rábano at the Fundación Cien and Dr. E. Martínez‐Sáez at the Hospital Vall d′Hebron for donating brain samples. We also thank the microscope facility at the CBMSO for their assistance with the confocal imaging and the institutional support of the Fundación Ramón Areces to the CBMSO. This work was funded by the Wylder Nation Foundation and by grants from the Spanish Ministry of Science and Education (SAF2017‐87698‐R) (AEI/FEDER, UE) to M.D.L and a MERIT award from the National Institutes of Health (2R37HD028607) to E.H.S.

Background: Mucopolysaccharidoses (MPS) are a group of inherited metabolic diseases caused by impaired function or absence of lysosomal enzymes involved in degradation of glycosaminoglycans. Clinically, MPS are skeletal dysplasias, characterized by cartilage abnormalities and disturbances in the process of endochondral ossification. Histologic abnormalities of growth cartilage have been reported at advanced stages of the disease, but information regarding growth plate pathology progression either in humans or in animal models, as well as its pathophysiology, is limited. Methods: Histological analyses of distal femur growth plates of wild type (WT) and mucopolysaccharidosis type VI (MPS VI) rats at different stages of development were performed, including quantitative data. Experimental findings were then analyzed in a theoretical scenario. Results: Histological evaluation showed a progressive loss of histological architecture within the growth plate. Furthermore, in silico simulation suggest the abnormal cell distribution in the tissue may lead to alterations in biochemical gradients, which may be one of the factors contributing to the growth plate abnormalities observed, highlighting aspects that must be the focus of future experimental works. Conclusion: The results presented shed some light on the progression of growth plate alterations observed in MPS VI and evidence the potentiality of combined theoretical and experimental approaches to better understand pathological scenarios, which is a necessary step to improve the search for novel therapeutic approaches.

Two distinct metabolic derangements are included under the eponym “Niemann–Pick disease (NPD)”. The first is due to the deficient activity of the lysosomal enzyme acid sphingomyelinase (ASM). Patients with this disorder [also referred to as ASM-deficient (ASMD) NPD] are broadly classified as having types A, A/B, and B NPD depending on their degree of neurologic involvement. Type A NPD patients, also known as the infantile neurovisceral form, exhibit hepatosplenomegaly in infancy and profound central nervous system (CNS) involvement. They rarely survive beyond 2–3 years of age. Type B patients, also known as the chronic visceral form, have hepatosplenomegaly and pathologic alterations of their lungs, but there are usually no CNS signs. The age of onset and rate of disease progression varies greatly among type B patients, and many may survive into adulthood. Patients with phenotypes intermediate between types A and B NPD (type A/B), also known as the chronic neurovisceral form, also have been identified. These individuals represent the expected continuum caused by inheriting different mutations in the ASM gene (SMPD1). Patients in the second NPD category are designated as having types C and D NPD. These individuals may have mild hepatosplenomegaly, but the CNS is profoundly affected. Impaired intracellular trafficking of cholesterol causes types C and D NPD, and two distinct gene defects have been found. All type D patients originate from a common Nova Scotian ancestry.

Studies over the past several years have demonstrated the important role of sphingolipids in cystic fibrosis (CF), chronic obstructive pulmonary disease and acute lung injury. Ceramide is increased in airway epithelial cells and alveolar macrophages of CF mice and humans, while sphingosine is dramatically decreased. This increase in ceramide results in chronic inflammation, increased death of epithelial cells, release of DNA into the bronchial lumen and thereby an impairment of mucociliary clearance; while the lack of sphingosine in airway epithelial cells causes high infection susceptibility in CF mice and possibly patients. The increase in ceramide mediates an ectopic expression of β1-integrins in the luminal membrane of CF epithelial cells, which results, via an unknown mechanism, in a down-regulation of acid ceramidase. It is predominantly this down-regulation of acid ceramidase that results in the imbalance of ceramide and sphingosine in CF cells. Correction of ceramide and sphingosine levels can be achieved by inhalation of functional acid sphingomyelinase inhibitors, recombinant acid ceramidase or by normalization of β1-integrin expression and subsequent re-expression of endogenous acid ceramidase. These treatments correct pulmonary inflammation and prevent or treat, respectively, acute and chronic pulmonary infections in CF mice with Staphylococcus aureus and mucoid or non-mucoid Pseudomonas aeruginosa. Inhalation of sphingosine corrects sphingosine levels only and seems to mainly act against the infection. Many antidepressants are functional inhibitors of the acid sphingomyelinase and were designed for systemic treatment of major depression. These drugs could be repurposed to treat CF by inhalation.

Acid sphingomyelinase deficiency (ASMD) is a rare lysosomal storage disorder with heterogeneous clinical manifestations, including hepatosplenomegaly and infiltrative pulmonary disease, and is associated with significant morbidity and mortality. Olipudase alfa (recombinant human acid sphingomyelinase) is an enzyme replacement therapy under development for the non-neurological manifestations of ASMD. We present a quantitative systems pharmacology (QSP) model supporting the clinical development of olipudase alfa. The model is multiscale and mechanistic, linking the enzymatic deficiency driving the disease to molecular-level, cellular-level, and organ-level effects. Model development was informed by natural history, and preclinical and clinical studies. By considering patient-specific pharmacokinetic (PK) profiles and indicators of disease severity, the model describes pharmacodynamic (PD) and clinical end points for individual patients. The ASMD QSP model provides a platform for quantitatively assessing systemic pharmacological effects in adult and pediatric patients, and explaining variability within and across these patient populations, thereby supporting the extrapolation of treatment response from adults to pediatrics.

Introduction: Farber disease is the lysosomal storage disorder due to the deficiency of acid ceramidase activity. Although it was described over six decades ago, it remains poorly described due to its unusual clinical presentation and rarity. No targeted therapies currently exist.Areas covered: The history of Farber disease, acid ceramidase and research leading to the development of therapy will be summarized, along with current knowledge regarding the clinical presentation and patient management.Expert opinion: Farber disease is often misdiagnosed as idiopathic juvenile arthritis. The clinical presentation is broad, with onset and survival ranging from early childhood to adults. Multiple organ systems, including the central nervous system, may be involved, but the most consistent finding in all patients is subcutaneous nodules that form mostly around joints. The nodules consist of lipid-filled macrophages, and inflammation is a major driver of the pathobiology. Pain, dysphonia and limitation of m...

A series of studies were carried out in Farber disease (OMIM #228000) cells and mice to evaluate the feasibility of enzyme replacement therapy (ERT) for this disorder. Media from Chinese hamster ovary (CHO) cells overexpressing human recombinant acid ceramidase (rhAC) was used to treat fibroblasts from a Farber disease patient, leading to significantly reduced ceramide. We also found that chondrocytes from Farber disease mice had a markedly abnormal chondrogenic phenotype, and this was corrected by rhAC as well. Acute dosing of rhAC in Farber mice confirmed the enzyme's bioactivity in vivo, and showed that it could be safely administered at doses up to 50 mg/kg. These studies also revealed little or no re-accumulation of ceramide in tissues for at least 7 days after enzyme administration. Once weekly administration of rhAC moderately improved survival of the mice, which could be enhanced by starting enzyme administration at an earlier age (3 days vs. 3 weeks). Repeat administration of the enzyme also led to normalization of spleen size, significantly reduced plasma levels of monocyte chemoattractant protein 1 (MCP-1), reduced infiltration of macrophages into liver and spleen, and significantly reduced ceramide and sphingosine in tissues. Overall, we conclude that ERT should be further developed for this debilitating and life-threatening disorder.

Acid Ceramidase Deficiency (Farber disease, FD) is an ultra-rare Lysosomal Storage Disorder that is poorly understood and often misdiagnosed as Juvenile Idiopathic Arthritis (JIA). Hallmarks of FD are accumulation of ceramides, widespread macrophage infiltration, splenomegaly, and lymphocytosis. The cytokines involved in this abnormal hematopoietic state are unknown. There are dozens of ceramide species and derivatives, but the specific ones that accumulate in FD have not been investigated. We used a multiplex assay to analyze cytokines and mass spectrometry to analyze ceramides in plasma from patients and mice with FD, controls, Farber patients treated by hematopoietic stem cell transplantation (HSCT), JIA patients, and patients with Gaucher disease. KC, MIP-1 alpha, and MCP-1 were sequentially upregulated in plasma from FD mice. MCP-1, IL-10, IL-6, IL-12, and VEGF levels were elevated in plasma from Farber patients but not in control or JIA patients. C16-Ceramide (C16-Cer) and dhC16-Cer were upregulated in plasma from FD mice. a-OH-C18-Cer, dhC12-Cer, dhC24:1-Cer, and C22:1-Cer-1P accumulated in plasma from patients with FD. Most cytokines and only a-OH-C18-Cer returned to baseline levels in HSCT-treated Farber patients. Sphingosines were not altered. Chitotriosidase activity was also relatively low. A unique cytokine and ceramide profile was seen in the plasma of Farber patients that was not observed in plasma from HSCT-treated Farber patients, JIA patients, or Gaucher patients. The cytokine profile can potentially be used to prevent misdiagnosis of Farber as JIA and to monitor the response to treatment. Further understanding of why these signaling molecules and lipids are elevated can lead to better understanding of the etiology and pathophysiology of FD and inform development of future treatments. (C) 2016 Elsevier B.V. All rights reserved.

Background/Aims: Cystic fibrosis (CF) is dominated by chronic inflammation and infection of the lung resulting in lung destruction and early death of patients. The lungs of CF patients are characterized by a massive accumulation of neutrophils. It requires definition why these massive numbers of neutrophils fail to eliminate typical CF pathogens like Staphylococcus aureus and Pseudomonas aeruginosa (P. aeruginosa) in CF lungs. Methods: We determined ceramide, sphingosine and reactive oxygen species (ROS) in neutrophils from wildtype and CF mice and determined the effect of sphingosine and ROS alone or in combination on killing of different P. aeruginosa strains. Results: We demonstrate that wildtype neutrophils are able to kill non-mucoid and mucoid clinical P. aeruginosa strains, while neutrophils from CF mice are insufficient to kill these P. aeruginosa strains, although both types of neutrophils infected with P. aeruginosa produce comparable levels of superoxide. All three analyzed P. aeruginosa strains are resistant to reactive oxygen species. The inability of CF neutrophils to kill P. aeruginosa is caused by a marked decrease of surface sphingosine levels in CF neutrophils. Wildtype neutrophils contain much higher concentrations of surface sphingosine than CF neutrophils. Further, wildtype neutrophils, but not CF neutrophils, release sphingosine, most likely as microparticles, upon infection. Sphingosine kills P. aeruginosa in vitro at low micromolar concentrations. Reconstitution of sphingosine in CF neutrophils restores their ability to kill these pathogens, demonstrating the significance of sphingosine for bacterial killing. Conclusion: The data provide evidence for a new paradigm explaining how neutrophils kill ROS-resistant P. aeruginosa, i.e. by sphingosine that kills P. aeruginosa at low concentrations. This mechanism is defective in CF neutrophils.

The eponym Niemann-Pick disease (NPD) refers to a group of patients who present with varying degrees of lipid storage and foam cell infiltration in tissues, as well as overlapping clinical features including hepatosplenomegaly, pulmonary insufficiency and/or central nervous system (CNS) involvement. Due to the pioneering work of Roscoe Brady and co-workers, we now know that there are two distinct metabolic abnormalities that account for NPD. The first is due to the deficient activity of the enzyme acid sphingomyelinase (ASM; “types A & B” NPD), and the second is due to defective function in cholesterol transport (“type C” NPD). Herein only types A and B NPD will be discussed. Type A NPD patients exhibit hepatosplenomegaly in infancy and profound CNS involvement. They rarely survive beyond 2–3 years of age. Type B patients also have hepatosplenomegaly and pathologic alterations of their lungs, but there are usually no CNS signs. The age of onset and rate of disease progression varies greatly among type B patients, and they frequently live into adulthood. Intermediate patients also have been reported with mild to moderate neurological findings. All patients with types A and B NPD have mutations in the gene encoding ASM (SMPD1), and thus the disease is more accurately referred to as ASM deficiency (ASMD). Herein we will review the clinical, pathological, biochemical, and genetic findings in types A and B NPD, and emphasize the seminal contributions of Dr. Brady to this disease. We will also discuss the current status of therapy for this disorder.

Niemann-Pick disease type A (NPD-A) is a lysosomal storage disorder characterized by neurodegeneration and early death. It is caused by loss-of-function mutations in the gene encoding for acid sphingomyelinase (ASM), which hydrolyzes sphingomyelin into ceramide. Here, we evaluated the safety of cerebellomedullary (CM) cistern injection of adeno-associated viral vector serotype 9 encoding human ASM (AAV9-hASM) in nonhuman primates (NHP). We also evaluated its therapeutic benefit in a mouse model of the disease (ASM-KO mice). We found that CM injection in NHP resulted in widespread transgene expression within brain and spinal cord cells without signs of toxicity. CM injection in the ASM-KO mouse model resulted in hASM expression in cerebrospinal fluid and in different brain areas without triggering an inflammatory response. In contrast, direct cerebellar injection of AAV9-hASM triggered immune response. We also identified a minimally effective therapeutic dose for CM injection of AAV9-hASM in mice. Two months after administration, the treatment prevented motor and memory impairment, sphingomyelin (SM) accumulation, lysosomal enlargement, and neuronal death in ASM-KO mice. ASM activity was also detected in plasma from AAV9-hASM CM-injected ASM-KO mice, along with reduced SM amount and decreased inflammation in the liver. Our results support CM injection for future AAV9-based clinical trials in NPD-A as well as other lysosomal storage brain disorders. © 2019 The Authors., This work was supported by Wylder Nation Foundation and by grants from Spanish Ministry of Economy and Competitivity (SAF-2014-57539-R and SAF2017-87698-R) to M.D.L. and from NIH-NINDS (R01NS073940) to K.S.B. A.P-C was a recipient of the FPU pre-doctoral fellowship from the Spanish Ministry of Economy and Competitivity. L.S. was a recipient of the Edward H. Schuchman Research Fellowship from the National Niemann-Pick Disease Foundation. E.H.S. and X.H. were supported by a MERIT award from the NIH (R37 HD28607) to E.H.S.

Gliosis in Niemann-Pick type C (NP-C) disease is characterized by marked changes in microglia and astrocytes. However, the gliosis onset and progression in NP-C has not been systematically studied, nor has the mechanism underlying this finding. Here, we found early gliosis in the subventricular zone (SVZ) of NP-C mice. Neural progenitor damage by Npc1 mutation suppressed vascular endothelial growth factor (VEGF) expression and further induced microglia activation followed by astrogliosis. Interestingly, excessive astrogliosis in the SVZ induced neural progenitor retention and/or migration into thalamus via astrocyte-derived VEGF, resulting in acceleration of thalamic and cortical gliosis through thalamo-cortical pathways. Transplantation of VEGF-overexpressing neural stem cells into the SVZ improved whole-brain pathology of NP-C mice. Overall, our data provide a new pathological perspective on NP-C neural pathology, revealing abnormalities in the subventricular-thalamo-cortical circuit of NP-C mouse brain and highlighting the importance of the SVZ microenvironment as a therapeutic target for NP-C disease.

Acid ceramidase (AC) hydrolyzes ceramides into sphingoid bases and fatty acids. The enzyme is overexpressed in several types of cancer and Alzheimer’s disease, and its genetic defect causes different incurable disorders. The availability of a method for the specific visualization of catalytically active AC in intracellular compartments is crucial for diagnosis and follow-up of therapeutic strategies in diseases linked to altered AC activity. This work was undertaken to develop activity-based probes for the detection of AC. Several analogues of the AC inhibitor SABRAC were synthesized and found to act as very potent (two-digit nM range) irreversible AC inhibitors by reaction with the active site Cys143. Detection of active AC in cell-free systems was achieved either by using fluorescent SABRAC analogues or by click chemistry with an azide-substituted analogue. The compound affording the best features allowed the unprecedented labeling of active AC in living cells., We thank Prof. H. Overkleeft and Dr. C. Ouairy (Leiden Institute of Chemistry) for a kind gift of BODIPY-carmofur, as well as A. Garcia (IQAC–CSIC), P. Rayo (IQAC–CSIC), I. Pérez-Pomeda (IQAC–CSIC), E. Dalmau (IQAC–CSIC), and Dr. M. Bosch (CCiT-UB) for their excellent technical assistance. This work has been partly funded by the Spanish Ministry of Economy, Industry and Competitiveness (Grants CTQ2014-54743-R, CTQ-2013-44334-P, and CTQ2017-85378-R).