Systemic delivery of a glucosylceramide synthase inhibitor reduces CNS substrates and increases lifespan in a mouse model of type 2 Gaucher disease.

Neuropathic Gaucher disease (nGD), also known as type 2 or type 3 Gaucher disease, is caused by a deficiency of the enzyme glucocerebrosidase (GC). This deficiency impairs the degradation of glucosylceramide (GluCer) and glucosylsphingosine (GluSph), leading to their accumulation in the brains of patients and mouse models of the disease. These accumulated substrates have been thought to cause the severe neuropathology and early death observed in patients with nGD and mouse models. Substrate accumulation is evident at birth in both nGD mouse models and humans affected with the most severe type of the disease. Current treatment of non-nGD relies on the intravenous delivery of recombinant human glucocerebrosidase to replace the missing enzyme or the administration of glucosylceramide synthase inhibitors to attenuate GluCer production. However, the currently approved drugs that use these mechanisms do not cross the blood brain barrier, and thus are not expected to provide a benefit for the neurological complications in nGD patients. Here we report the successful reduction of substrate accumulation and CNS pathology together with a significant increase in lifespan after systemic administration of a novel glucosylceramide synthase inhibitor to a mouse model of nGD. To our knowledge this is the first compound shown to cross the blood brain barrier and reduce substrates in this animal model while significantly enhancing its lifespan. These results reinforce the concept that systemically administered glucosylceramide synthase inhibitors could hold enhanced therapeutic promise for patients afflicted with neuropathic lysosomal storage diseases.

Merits of combination cortical, subcortical, and cerebellar injections for the treatment of Niemann-Pick disease type A.

Niemann-Pick disease Type A (NPA) is a neuronopathic lysosomal storage disease (LSD) caused by the loss of acid sphingomyelinase (ASM). The goals of the current study are to ascertain the levels of human ASM that are efficacious in ASM knockout (ASMKO) mice, and determine whether these levels can be attained in non-human primates (NHPs) using a multiple parenchymal injection strategy. Intracranial injections of different doses of AAV1-hASM in ASMKO mice demonstrated that only a small amount of enzyme (<0.5 mg hASM/g tissue) was sufficient to increase survival, and that increasing the amount of hASM did not enhance this survival benefit until a new threshold level of >10 mg hASM/g tissue was reached. In monkeys, injection of 12 tracts of AAV1-hASM resulted in efficacious levels of enzyme in broad regions of the brain that was aided, in part, by axonal transport of adeno-associated virus (AAV) and movement through the perivascular space. This study demonstrates that a combination cortical, subcortical, and cerebellar injection protocol could provide therapeutic levels of hASM to regions of the NHP brain that are highly affected in NPA patients. The information from this study might help design new AAV-mediated enzyme replacement protocols for NPA and other neuronopathic LSDs in future clinical trials.

Iminosugar-based inhibitors of glucosylceramide synthase increase brain glycosphingolipids and survival in a mouse model of Sandhoff disease.

The neuropathic glycosphingolipidoses are a subgroup of lysosomal storage disorders for which there are no effective therapies. A potential approach is substrate reduction therapy using inhibitors of glucosylceramide synthase (GCS) to decrease the synthesis of glucosylceramide and related glycosphingolipids that accumulate in the lysosomes. Genz-529468, a blood-brain barrier-permeant iminosugar-based GCS inhibitor, was used to evaluate this concept in a mouse model of Sandhoff disease, which accumulates the glycosphingolipid GM2 in the visceral organs and CNS. As expected, oral administration of the drug inhibited hepatic GM2 accumulation. Paradoxically, in the brain, treatment resulted in a slight increase in GM2 levels and a 20-fold increase in glucosylceramide levels. The increase in brain glucosylceramide levels might be due to concurrent inhibition of the non-lysosomal glucosylceramidase, Gba2. Similar results were observed with NB-DNJ, another iminosugar-based GCS inhibitor. Despite these unanticipated increases in glycosphingolipids in the CNS, treatment nevertheless delayed the loss of motor function and coordination and extended the lifespan of the Sandhoff mice. These results suggest that the CNS benefits observed in the Sandhoff mice might not necessarily be due to substrate reduction therapy but rather to off-target effects.

Timing of therapeutic intervention determines functional and survival outcomes in a mouse model of late infantile batten disease.

Classical late infantile neuronal ceroid lipofuscinosis (cLINCL) is a monogenic disorder caused by the loss of tripeptidyl peptidase 1 (TPP1) activity as a result of mutations in CLN2. Absence of TPP1 results in lysosomal storage with an accompanying axonal degeneration throughout the central nervous system (CNS), which leads to progressive neurodegeneration and early death. In this study, we compared the efficacies of pre- and post-symptomatic injections of recombinant adeno-associated virus (AAV) for treating the cellular and functional abnormalities of CLN2 mutant mice. Intracranial injection of AAV1-hCLN2 resulted in widespread human TPP1 (hTPP1) activity in the brain that was 10-100-fold above wild-type levels. Injections before disease onset prevented storage and spared neurons from axonal degeneration, reflected by the preservation of motor function. Furthermore, the majority of CLN2 mutant mice treated pre-symptomatically lived for at least 330 days, compared with a median survival of 151 days in untreated CLN2 mutant controls. In contrast, although injection after disease onset ameliorated lysosomal storage, there was evidence of axonal degeneration, motor function showed limited recovery, and the animals had a median lifespan of 216 days. These data illustrate the importance of early intervention for enhanced therapeutic benefit, which may provide guidance in designing novel treatment strategies for cLINCL patients.

Radiological evidence of early cerebral microvascular disease in young children with Fabry disease.

We report on 2 children with Fabry disease who had radiologic evidence of microvascular central nervous system involvement despite the clinical absence of renal, cardiac, or cerebral manifestations. This suggests that treatment with enzyme replacement therapy may be necessary early in the disease to avoid irreversible complications.

Correlation of surrogate markers of Gaucher disease. Implications for long-term follow up of enzyme replacement therapy.

BACKGROUND: The excessive storage of cellular debris in the lysosomal storage disorders triggers a variety of cellular responses. Some of these responses are maladaptative and result in the pathology of these diseases. To some extent, cellular responses are specific to the stored material, which influences the pathophysiology of the disease and results in some of its characteristic features. METHODS: A large body of data has been collected for three biochemical (surrogate) markers of Gaucher Disease: angiotensin converting enzyme (ACE), tartrate-resistant acid phosphatase (TRAP) and chitotriosidase (CHITO) using currently available enzyme analysis. Follow up data was gathered in a group of 18 patients. RESULTS: The three markers are correlated between each other and are useful indicators of the disease progress and its response to enzyme replacement therapy (ERT). Retrospective analysis of clinical records and comparison of chitotriosidase values with the baseline Severity Score Index (SSI) allowed prediction of the response patterns for this marker when long-term ERT (>24 months) was evaluated. CONCLUSIONS: The less severely affected patients are more likely to normalize their chitotriosidase activities after long term ERT.

Gene therapy for the lysosomal storage disorders.

The lysosomal storage disorders (LSD) are monogenic inborn errors of metabolism with heterogeneous pathophysiology and clinical manifestations. In recent decades, these disorders have been models for the development of molecular and cellular therapies for inherited metabolic diseases. Studies in preclinical in vitro systems and animal models have established proof-of-concept for the development of bone marrow transplantation (BMT) and enzyme-replacement therapy (ERT) as therapeutic options for several LSDs. BMT is limited by poor donor availability and high morbidity and mortality, and although ERT is a good treatment, it is not a life-long cure. Its high cost remains an impediment for developing countries. While substrate synthesis inhibition therapy is an important idea, its clinical use is far from certain. The neuropathology present in many LSDs has responded poorly to BMT or ERT, which makes gene therapy an attractive therapeutic alternative. Oncoretroviral vectors, and more recently adeno-associated and lentiviral vectors have been tested with some success. This review summarizes the main gene therapy strategies which have been employed or are under development for both non-neurological and neuronopathic LSDs. Some of the in vitro and in vivo preclinical studies presented herein have provided the rationale for gene therapy clinical trials for Gaucher disease Type 1.