Model Of Lysosomal Storage Disease Research Articles

A PIKfyve modulator combined with an integrated stress response inhibitor to treat lysosomal storage diseases

Lysosomal degradation pathways coordinate the clearance of superfluous and damaged cellular components. Compromised lysosomal degradation is a hallmark of many degenerative diseases, including lysosomal storage diseases (LSDs), which are caused by loss-of-function mutations within both alleles of a lysosomal hydrolase, leading to lysosomal substrate accumulation. Gaucher's disease, characterized by <15% of normal glucocerebrosidase function, is the most common LSD and is a prominent risk factor for developing Parkinson's disease. Here, we show that either of two structurally distinct small molecules that modulate PIKfyve activity, identified in a high-throughput cellular lipid droplet clearance screen, can improve glucocerebrosidase function in Gaucher patient-derived fibroblasts through an MiT/TFE transcription factor that promotes lysosomal gene translation. An integrated stress response (ISR) antagonist used in combination with a PIKfyve modulator further improves cellular glucocerebrosidase activity, likely because ISR signaling appears to also be slightly activated by treatment by either small molecule at the higher doses employed. This strategy of combining a PIKfyve modulator with an ISR inhibitor improves mutant lysosomal hydrolase function in cellular models of additional LSD.

Characterization of early markers of disease in the mouse model of mucopolysaccharidosis IIIB

BackgroundMucopolysaccharidosis (MPS) IIIB, also known as Sanfilippo Syndrome B, is a devastating childhood disease. Unfortunately, there are currently no available treatments for MPS IIIB patients. Yet, animal models of lysosomal storage diseases have been valuable tools in identifying promising avenues of treatment. Enzyme replacement therapy, gene therapy, and bone marrow transplant have all shown efficacy in the MPS IIIB model systems. A ubiquitous finding across rodent models of lysosomal storage diseases is that the best treatment outcomes resulted from intervention prior to symptom onset. Therefore, the aim of the current study was to identify early markers of disease in the MPS IIIB mouse model as well as examine clinically-relevant behavioral domains not yet explored in this model.MethodsUsing the MPS IIIB mouse model, we explored early developmental trajectories of communication and gait, and later social behavior, fear-related startle and conditioning, and visual capabilities. In addition, we examined brain structure and function via magnetic resonance imaging and diffusion tensor imaging.ResultsWe observed reduced maternal isolation-induced ultrasonic vocalizations in MPS IIIB mice relative to controls, as well as disruption in a number of the spectrotemporal features. MPS IIIB also exhibited disrupted thermoregulation during the first two postnatal weeks without any differences in body weight. The developmental trajectories of gait were largely normal. In early adulthood, we observed intact visual acuity and sociability yet a more submissive phenotype, increased aggressive behavior, and decreased social sniffing relative to controls. MPS IIIB mice showed greater inhibition of startle in response to a pretone with a decrease in overall startle response and reduced cued fear memory. MPS IIIB also weighed significantly more than controls throughout adulthood and showed larger whole brain volumes and normalized regional volumes with intact tissue integrity as measured with magnetic resonance and diffusion tensor imaging, respectively.ConclusionsTogether, these results indicate disease markers are present as early as the first two weeks postnatal in this model. Further, this model recapitulates social, sensory and fear-related clinical features. Our study using a mouse model of MPS IIIB provides essential baseline information that will be useful in future evaluations of potential treatments.

Experimental Models of CNS Deficits in Lysosomal Storage Diseases

Lysosomal storage diseases are characterized by enzyme deficiency in the lysosomal appar-atus of the cell, triggering a pathological accumulation of undigested cellular material (proteins, lipids or carbohydrates) and tissue damage. Clinically and etiologically dive-rse, this group includes over 70 presently recognized hereditary conditions with no known effective therapy. Thus, the search for therapeutic strategies directed at these disorders represents an urgent unmet biomedical task, also necessitating the use of appropriate and valid experimental (animal) models. Here, we discuss the existing models of lysosomal storage diseases and the applicability of rodent and zebrafish as model organisms for probing these diseases.

Gene therapy for lysosomal storage diseases: Current clinical trial prospects.

Lysosomal storage diseases (LSDs) are a group of metabolic inborn errors caused by defective enzymes in the lysosome, resulting in the accumulation of undegraded substrates. LSDs are progressive diseases that exhibit variable rates of progression depending on the disease and the patient. The availability of effective treatment options, including substrate reduction therapy, pharmacological chaperone therapy, enzyme replacement therapy, and bone marrow transplantation, has increased survival time and improved the quality of life in many patients with LSDs. However, these therapies are not sufficiently effective, especially against central nerve system abnormalities and corresponding neurological and psychiatric symptoms because of the blood-brain barrier that prevents the entry of drugs into the brain or limiting features of specific treatments. Gene therapy is a promising tool for the treatment of neurological pathologies associated with LSDs. Here, we review the current state of gene therapy for several LSDs for which clinical trials have been conducted or are planned. Several clinical trials using gene therapy for LSDs are underway as phase 1/2 studies; no adverse events have not been reported in most of these studies. The administration of viral vectors has achieved good therapeutic outcomes in animal models of LSDs, and subsequent human clinical trials are expected to promote the practical application of gene therapy for LSDs.

Structure-performance relationships of four lysosomal markers used for the imaging of HT-29 cancer cells and a cellular model of lysosomal storage disease (Niemann-Pick C)

Four new BODIPY derivatives with a potential tendency to aggregation have been synthesized and characterized by means of NMR techniques, mass spectrometry and UV–Vis/fluorescence spectroscopies. The objective of this study is to determine which structural factors of the new molecules influence most notably the cellular uptake, intracellular location and fluorescence imaging abilities. The behaviour of the compounds in organic solvent and aqueous solution has been studied. In organic solvents (DMSO, ethanol and toluene), the photophysical properties of the new molecules are almost independent of the building blocks used to synthesize the pendant moieties (non-fluorogenic parts). In an aqueous environment (HEPES Buffer Solution, pH 7), at 10 μM, three of the compounds (1, 2 and 4) tend to form weakly emissive nanoparticles (DLS determination) whereas one of them (3) remains soluble and highly fluorescent. In the nanomolar range of concentration, all the compounds are aqueous soluble. The cellular internalisation of the compounds (10 nM) has been studied in human colon adenocarcinoma HT-29 cells by means of flow cytometry and confocal laser scanning microscopy. All the compounds were uptaken by HT-29 cells, but notably molecule 3 (made with lysine as a building block) was the one displaying a higher loading and a more clear lysosomal location (0.88 Pearson's correlation coefficient in colocalization assays using lysosomal fluorescent probe LysoTracker DND-99). Molecule 3 also performed better than the valine derivative 1 as a lysosomal marker in a cellular model (human adrenal carcinoma SW13/cl.2 cells) of lysosomal storage disease (Niemann-Pick type C).

Reproduction in Animal Models of Lysosomal Storage Diseases: A Scoping Review.

Background: Lysosomal storage diseases (LSDs) are caused by a mutation in a specific gene. Enzymatic dysfunction results in a progressive storage of substrates that gradually affects lysosomal, cellular and tissue physiology. Their pathophysiological consequences vary according to the nature of the stored substrate, making LSDs complex and multisystemic diseases. Some LSDs result in near normal life expectancies, and advances in treatments mean that more people reach the age to have children, so considering the effects of LSDs on fertility and the risks associated with having children is of growing importance. Objectives: As there is a lack of clinical studies describing the effect of LSDs on the physiology of reproductivity, we undertook a scoping review of studies using animal models of LSDs focusing on reproductive parameters. Methods: We searched six databases: MEDLINE, LILACS, Scopus, Web of Science, Embase and SciELO, and identified 49 articles that met our inclusion criteria. Results: The majority of the studies used male animal models, and a number reported severe morphological and physiological damage in gametes and gonads in models of sphingolipidoses. Models of other LSDs, such as mucopolysaccharidoses, presented important morphological damage. Conclusion: Many of the models found alterations in reproductive systems. Any signs of subfertility or morphological damage in animal models are important, particularly in rodents which are extremely fertile, and may have implications for individuals with LSDs. We suggest the use of more female animal models to better understand the physiopathology of the diseases, and the use of clinical case studies to further explore the risks of individuals with LSDs having children.

Recapture Lysosomal Enzyme Deficiency via Targeted Gene Disruption in the Human Near-Haploid Cell Line HAP1.

Background: Advancement in genome engineering enables rapid and targeted disruption of any coding sequences to study gene functions or establish human disease models. We explored whether this approach can be used to study Gaucher disease, one of the most common types of lysosomal storage diseases (LSDs) in a near-haploid human cell line (HAP1). Results: CRISPR-Cas9 targeting to coding sequences of β-glucocerebrosidase (GBA), the causative gene of Gaucher disease, resulted in an insertional mutation and premature termination of GBA. We confirmed the GBA knockout at both the gene and enzyme levels by genotyping and GBA enzymatic assay. Characterization of the knockout line showed no significant changes in cell morphology and growth. Lysosomal staining revealed more granular lysosomes in the cytosol of the GBA-knockout line compared to its parental control. Flow cytometry analysis further confirmed that more lysosomes accumulated in the cytosol of the knockout line, recapturing the disease phenotype. Finally, we showed that this knockout cell line could be used to evaluate a replacement therapy by recombinant human GBA. Conclusions: Targeted gene disruption in human HAP1 cells enables rapid establishment of the Gaucher model to capture the key pathology and to test replacement therapy. We expect that this streamlined method can be used to generate human disease models of other LSDs, most of which are still lacking both appropriate human disease models and specific treatments to date.

Murine Models of Lysosomal Storage Diseases Exhibit Differences in Brain Protein Aggregation and Neuroinflammation.

Genetic, epidemiological and experimental evidence implicate lysosomal dysfunction in Parkinson’s disease (PD) and related synucleinopathies. Investigate several mouse models of lysosomal storage diseases (LSDs) and evaluate pathologies reminiscent of synucleinopathies. We obtained brain tissue from symptomatic mouse models of Gaucher, Fabry, Sandhoff, Niemann–Pick A (NPA), Hurler, Pompe and Niemann–Pick C (NPC) diseases and assessed for the presence of Lewy body-like pathology (proteinase K-resistant α-synuclein and tau aggregates) and neuroinflammation (microglial Iba1 and astrocytic GFAP) by immunofluorescence. All seven LSD models exhibited evidence of proteinopathy and/or inflammation in the central nervous system (CNS). However, these phenotypes were divergent. Gaucher and Fabry mouse models displayed proteinase K-resistant α-synuclein and tau aggregates but no neuroinflammation; whereas Sandhoff, NPA and NPC showed marked neuroinflammation and no overt proteinopathy. Pompe disease animals uniquely displayed widespread distribution of tau aggregates accompanied by moderate microglial activation. Hurler mice also demonstrated proteinopathy and microglial activation. The present study demonstrated additional links between LSDs and pathogenic phenotypes that are hallmarks of synucleinopathies. The data suggest that lysosomal dysregulation can contribute to brain region-specific protein aggregation and induce widespread neuroinflammation in the brain. However, only a few LSD models examined exhibited phenotypes consistent with synucleinopathies. While no model can recapitulate the complexity of PD, they can enable the study of specific pathways and mechanisms contributing to disease pathophysiology. The present study provides evidence that there are existing, previously unutilized mouse models that can be employed to study pathogenic mechanisms and gain insights into potential PD subtypes, helping to determine if they are amenable to pathway-specific therapeutic interventions.

Synaptic Function and Dysfunction in Lysosomal Storage Diseases.

Lysosomal storage diseases (LSDs) with neurological involvement are inherited genetic diseases of the metabolism characterized by lysosomal dysfunction and the accumulation of undegraded substrates altering glial and neuronal function. Often, patients with neurological manifestations present with damage to the gray and white matter and irreversible neuronal decline. The use of animal models of LSDs has greatly facilitated studying and identifying potential mechanisms of neuronal dysfunction, including alterations in availability and function of synaptic proteins, modifications of membrane structure, deficits in docking, exocytosis, recycling of synaptic vesicles, and inflammation-mediated remodeling of synapses. Although some extrapolations from findings in adult-onset conditions such as Alzheimer’s disease or Parkinson’s disease have been reported, the pathogenetic mechanisms underpinning cognitive deficits in LSDs are still largely unclear. Without being fully inclusive, the goal of this mini-review is to present a discussion on possible mechanisms leading to synaptic dysfunction in LSDs.

Modeling Lysosomal Storage Diseases in the Zebrafish.

Lysosomal storage diseases (LSDs) are a family of 70 metabolic disorders characterized by mutations in lysosomal proteins that lead to storage material accumulation, multiple-organ pathologies that often involve neurodegeneration, and early mortality in a significant number of patients. Along with the necessity for more effective therapies, there exists an unmet need for further understanding of disease etiology, which could uncover novel pathways and drug targets. Over the past few decades, the growth in knowledge of disease-associated pathways has been facilitated by studies in model organisms, as advancements in mutagenesis techniques markedly improved the efficiency of model generation in mammalian and non-mammalian systems. In this review we highlight non-mammalian models of LSDs, focusing specifically on the zebrafish, a vertebrate model organism that shares remarkable genetic and metabolic similarities with mammals while also conferring unique advantages such as optical transparency and amenability toward high-throughput applications. We examine published zebrafish LSD models and their reported phenotypes, address organism-specific advantages and limitations, and discuss recent technological innovations that could provide potential solutions.

Pre-clinical Mouse Models of Neurodegenerative Lysosomal Storage Diseases.

There are over 50 lysosomal hydrolase deficiencies, many of which cause neurodegeneration, cognitive decline and death. In recent years, a number of broad innovative therapies have been proposed and investigated for lysosomal storage diseases (LSDs), such as enzyme replacement, substrate reduction, pharmacologic chaperones, stem cell transplantation, and various forms of gene therapy. Murine models that accurately reflect the phenotypes observed in human LSDs are critical for the development, assessment and implementation of novel translational therapies. The goal of this review is to summarize the neurodegenerative murine LSD models available that recapitulate human disease, and the pre-clinical studies previously conducted. We also describe some limitations and difficulties in working with mouse models of neurodegenerative LSDs.

Lipid Antigen Presentation by CD1b and CD1d in Lysosomal Storage Disease Patients.

The lysosome has a key role in the presentation of lipid antigens by CD1 molecules. While defects in lipid antigen presentation and in invariant Natural Killer T (iNKT) cell response were detected in several mouse models of lysosomal storage diseases (LSD), the impact of lysosomal engorgement in human lipid antigen presentation is poorly characterized. Here, we analyzed the capacity of monocyte-derived dendritic cells (Mo-DCs) from Fabry, Gaucher, Niemann Pick type C and Mucopolysaccharidosis type VI disease patients to present exogenous antigens to lipid-specific T cells. The CD1b- and CD1d-restricted presentation of lipid antigens by Mo-DCs revealed an ability of LSD patients to induce CD1-restricted T cell responses within the control range. Similarly, freshly isolated monocytes from Fabry and Gaucher disease patients had a normal ability to present α-Galactosylceramide (α-GalCer) antigen by CD1d. Gaucher disease patients' monocytes had an increased capacity to present α-Gal-(1-2)-αGalCer, an antigen that needs internalization and processing to become antigenic. In summary, our results show that Fabry, Gaucher, Niemann Pick type C, and Mucopolysaccharidosis type VI disease patients do not present a decreased capacity to present CD1d-restricted lipid antigens. These observations are in contrast to what was observed in mouse models of LSD. The percentage of total iNKT cells in the peripheral blood of these patients is also similar to control individuals. In addition, we show that the presentation of exogenous lipids that directly bind CD1b, the human CD1 isoform with an intracellular trafficking to the lysosome, is normal in these patients.

Low-dose Gene Therapy Reduces the Frequency of Enzyme Replacement Therapy in a Mouse Model of Lysosomal Storage Disease.

Enzyme replacement therapy (ERT) is the standard of care for several lysosomal storage diseases (LSDs). ERT, however, requires multiple and costly administrations and has limited efficacy. We recently showed that a single high dose administration of adeno-associated viral vector serotype 8 (AAV2/8) is at least as effective as weekly ERT in a mouse model of mucopolysaccharidosis type VI (MPS VI). However, systemic administration of high doses of AAV might result in both cell-mediated immune responses and insertional mutagenesis. Here we evaluated whether the combination of low doses of AAV2/8 with a less frequent (monthly) than canonical (weekly) ERT schedule may be as effective as the single treatments at high doses or frequent regimen. A greater reduction of both urinary glycosaminoglycans, considered a sensitive biomarker of therapeutic efficacy, and storage in the myocardium and heart valves was observed in mice receiving the combined than the single therapies. Importantly, these levels of correction were similar to those we obtained in a previous study following either high doses of AAV2/8 or weekly ERT. Our data show that low-dose gene therapy can be used as a means to rarify ERT administration, thus reducing both the risks and costs associated with either therapies.

Diminished MTORC1-Dependent JNK Activation Underlies the Neurodevelopmental Defects Associated with Lysosomal Dysfunction

Here, we evaluate the mechanisms underlying the neurodevelopmental deficits in Drosophila and mouse models of lysosomal storage diseases (LSDs). We find that lysosomes promote the growth of neuromuscular junctions (NMJs) via Rag GTPases and mechanistic target of rapamycin complex 1 (MTORC1). However, rather than employing S6K/4E-BP1, MTORC1 stimulates NMJ growth via JNK, a determinant of axonal growth in Drosophila and mammals. This role of lysosomal function in regulating JNK phosphorylation is conserved in mammals. Despite requiring the amino-acid-responsive kinase MTORC1, NMJ development is insensitive to dietary protein. We attribute this paradox to anaplastic lymphoma kinase (ALK), which restricts neuronal amino acid uptake, and the administration of an ALK inhibitor couples NMJ development to dietary protein. Our findings provide an explanation for the neurodevelopmental deficits in LSDs and suggest an actionable target for treatment.

280. Combination of Low-Dose Gene Therapy and Monthly Enzyme Replacement Therapy Improves the Phenotype of a Mouse Model of Lysosomal Storage Disease

Enzyme replacement therapy (ERT) is the current standard of care for Mucopolysaccharidosis type VI (MPS VI) that is caused by deficiency of arylsulfatase B (ARSB), which results in widespread accumulation and excretion of toxic glycosaminoglycans (GAGs). However, ERT is associated with inconvenient multiple and costly administrations and fails to ameliorate cardiac, visual and bone abnormalities. To overcome ERT limitations, we developed a successful gene therapy approach based on a single administration of AAV2/8 that targets liver of MPS VI animal models. Importantly, we showed that a single systemic administration of AAV2/8 at high doses (2×10e12 gc/kg) is at least as effective as the current ERT therapeutic regimen based on weekly infusions of recombinant human ARSB (rhARSB). If this translates to humans, gene therapy could replace ERT for MPS VI. However, the administration of high doses of AAV2/8 requires a challenging and costly production process, and results in cell-mediated immune responses that eliminate transduced hepatocytes. Similarly, weekly ERT infusions are costly and require high patient compliance. We therefore evaluated in a mouse model of MPS VI whether the combination of low doses of AAV2/8 at 6×10e11 or 2×10e11 gc/kg with a rarified ERT schedule (1mg/kg once a month) may be as effective as the single treatments at high doses or frequent regimen. Significant increase of ARSB activity was found in liver of all treated mice. Detectable but low activity was variably observed in spleen and kidney and was associated with significant reduction of tissue GAGs, regardless of treatment and ARSB activity levels, similarly to what observed in mice treated with high doses of AAV2/8 or weekly ERT. This supports previous data indicating that low enzymatic levels improve MPS VI visceral phenotype. Evaluation of GAG storage in myocardium and heart valves is in progress. Urinary GAG, which are a sensitive biomarker of systemic clearance of lysosomal storage and, thus of therapeutic efficacy, are slightly reduced in mice treated with either monthly ERT or 2×10e11 gc/kg of AAV2/8. The reduction is more consistent at 6×10e11 gc/kg. Importantly, urinary GAG decreased more in mice receiving the combined therapy than in those receiving single treatments. In particular, urinary GAG reduction in mice treated with both 6×10e11 gc/kg of AAV2/8 and monthly ERT was comparable to that obtained following administration of either high doses of AAV2/8, i.e 2×10e12 gc/kg, or weekly ERT. The data collected so far show similar efficacy between low-dose gene therapy combined with rarified ERT and the corresponding single treatments at high doses or frequent regimen. This should increase the safety and reduce the risks and costs associated with both therapeutic approaches.

Fusion of Lysosomes with Secretory Vesicle Leads to Excessive Uncontrolled Exocytosis in Mucolipidosis Type IV

Mutations in TRPML1 cause the lysosomal storage disease (LSD) Mucolipidosis type IV (MLIV), which is characterized by neurodegeneration, muscular hypotonia corneal opacity, achlorhydia, anemia and severe psychomotor deficiency with impaired neuromuscular junctions. The role of the TRP channel TRPML1 in cell function and how mutations in TRPML1 cause MLIV are not well understood. Studies on the causes of MLIV are focused on the potential role of TRPML1 in constitutive membrane trafficking to and from the lysosomes. However, the role of TRPML1 in secretory cells that their cardinal function is regulated exocytosis has not been studied. Here, we analyzed several forms of regulated exocytosis in neurons and secretory epithelia in a knockout mouse model of TRPML1 and, unexpectedly, found excessive exocytosis due to marked enlargement of synaptic vesicles and secretory granules due to fusion with lysosomes. As a result, Trpml1−/− neurons maintain high basal and stimulated exocytosis of the neurotransmitter and neurotoxin glutamate, which may account for the neurodegeneration in ML4. Trpml1−/− pancreatic and salivary glands acini show high Ca2+− and cAMP-stimulated exocytosis, resulting in pancreatitis. These features were not found in another model of LSD, the Niemann-Pick disease type C1 mouse model. These findings indicate that a major role of TRPML1 is to guard against pathological fusion of lysosomes with other intracellular organelles, including secretory vesicles, and suggest a new approach towards developing treatment for MLIV.

Similar Therapeutic Efficacy Between a Single Administration of Gene Therapy and Multiple Administrations of Recombinant Enzyme in a Mouse Model of Lysosomal Storage Disease

Enzyme replacement therapy (ERT) has become the standard of care for several lysosomal storage disorders (LSDs). Despite ERT's undisputed efficacy, the requirement for multiple and costly administrations as well as ERT's limited improvement of some LSD manifestations prompts the search for better therapies. Using a mouse model of mucopolysaccharidosis VI, we compared the efficacy of a single intravascular administration of an adeno-associated viral vector targeting liver to weekly infusions of human recombinant enzyme at the same doses used in mucopolysaccharidosis VI patients. While gene therapy results in increased and stable levels of circulating enzyme up to 1 year after vector administration, ERT has typical peak-and-drop serum kinetics. Both therapies similarly reduced glycosaminoglycan levels in urine and tissues including heart valves and myocardium, with gene therapy improving skeletal skull abnormalities slightly better, although not significantly, than ERT. Both therapies seem to similarly improve animal motor performance, with gene therapy possibly associated with less animal distress. Thus, a single vector administration that converts liver into a factory organ for systemic secretion of therapeutic proteins is at least as effective as ERT in a mouse model of LSD, potentially eliminating problems with compliance and costs. Only testing in humans will prove whether this holds true in a clinical setting.

New Players in the Infertility of a Mouse Model of Lysosomal Storage Disease: The Hypothalamus-Pituitary-Gonadal Axis

Mammalian spermatogenesis is a complex hormone-dependent developmental program where interactions between different cell types are finely regulated. Mouse models in which any of the sperm maturation steps are perturbed provide major insights into the molecular control of spermatogenesis. The Twitcher mouse is a model for the Krabbe disease, characterized by the deficiency of galactosylceramidase (GALC), a lysosomal enzyme that hydrolyzes the terminal galactose from galactosylceramide, a typical component of the myelin membrane. In addition, GALC catalyzes the hydrolysis of the terminal galactose from galactosyl-alkyl-acyl-glycerol, precursor of seminolipids, specifically expressed on the membrane of germ cells. Previous data reported by our group demonstrated that glycolipids play an important role in sperm maturation and differentiation. Moreover, we hypothesized that the severe impairment of the central nervous system that affects the Twitcher mouse could interfere with the hypothalamus-pituitary-gonadal axis function, contributing to infertility. To highlight this hypothesis we have determined, at molecular level, the potential variation in expression pattern of brain hormones involved in spermatogenesis regulation.

Lysosomal storage diseases: Stem cell-based cell- and gene-therapy.

Lysosomal storage diseases (LSDs) are caused by inborn genetic defects and most affected babies show pathology in the CNS. LSDs are caused by a specific inherited enzyme deficiency that results in accumulation of substrates in the lysosomes, distension of the organelles and subsequent cellular malfunction. Currently, no effective treatment is available for most of the LSDs, because the blood?brain barrier bars entry of enzyme preparations into the brain. Treatment for LSDs can be divided into those address symptoms or those address cause. At present, successful treatments for the LSDs are enzyme replacement therapy (ERT) and cell therapy. ERT is most successful in Gaucher disease and has been approved for Fabry disease, and mucopolysaccharidosis I (MPS I). In addition, ERT for Pompe disease, MPS II, MPS VI and MPS VII has been planned and awaiting approval for treatment. Limitations in ERT include need for life-long treatment, development of antibodies, and inability to cross blood brain barrier (BBB) resulting in failure to halt disease progression in the brain. Transplantation of hematopoietic stem cells (HSCs), bone marrow stem cells (BMSCs) and umbilical cord blood-derived stem cells (UCBSCs) offer effective but limited efficacy for patients suffering from Krabbe disease, MPS VII and adrenal leukodystrophy but in other LSDs they are ineffective. Intracranial/intracerebral transplantation of genetically modified stem cells as enzyme delivery system could bypass the BBB effectively and ensure release of therapeutically beneficial amount of enzymes to affected CNS lesion sites. For this reason, stem cell-based gene therapy is the most effective treatment for LSDs. In mouse models of LSDs, genetically modified neural stem cells encoding enzyme genes effectively decreased lysosomal storage, reduced pathology and extended life span of animals. Cell-based gene therapies for LSDs bridge the application of ERT and gene therapy and are important direction to pursue in the future.

Sialic Acid Deposition Impairs the Utility of AAV9, but Not Peptide-modified AAVs for Brain Gene Therapy in a Mouse Model of Lysosomal Storage Disease

Recombinant vector systems have been recently identified that when delivered systemically can transduce neurons, glia, and endothelia in the central nervous system (CNS), providing an opportunity to develop therapies for diseases affecting the brain without performing direct intracranial injections. Vector systems based on adeno-associated virus (AAV) include AAV serotype 9 (AAV9) and AAVs that have been re-engineered at the capsid level for CNS tropism. Here, we performed a head-to-head comparison of AAV9 and a capsid modified AAV for their abilities to rescue CNS and peripheral disease in an animal model of lysosomal storage disease (LSD), the mucopolysacharidoses (MPS) VII mouse. While the peptide-modified AAV reversed cognitive deficits, improved storage burden in the brain, and substantially prolonged survival, we were surprised to find that AAV9 provided no CNS benefit. Additional experiments demonstrated that sialic acid, a known inhibitor of AAV9, is elevated in the CNS of MPS VII mice. These studies highlight how disease manifestations can dramatically impact the known tropism of recombinant vectors, and raise awareness to assuming similar transduction profiles between normal and disease models.