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Lysosomal diseases are a group of over 70 rare genetic conditions in which a protein deficiency (most often an enzyme deficiency) leads to multi-system disease. Current therapies for lysosomal diseases are limited in their ability to treat certain tissues that are major contributors to morbidity and mortality, such as the central nervous system (CNS) and cardiac valves. For this study, the lysosomal disease mucopolysaccharidosis type I (MPS I) was selected as the disease model. In MPS I, mutations in theMPS I mice were hydrodynamically injected through the tail vein with plasmids containing either a codon-optimized cDNA encoding the wild-type IDUA protein or one of four modified IDUAs under the control of the liver-specific human α1-antitrypsin (hAAT) promoter. Two modified IDUAs contained a ligand for the CB1 receptor, which is a highly expressed receptor in the CNS. Iduronidase activity levels were measured in the tissues and plasma using an enzyme activity assay.The modified IDUAs did not appear to have improved activity levels in the brain compared with the unmodified IDUA. However, one modified IDUA exhibited higher activity levels than the unmodified IDUA in the heart (The addition of a six amino acid peptide improved iduronidase's activity levels in the heart and plasma. The short length of this LT peptide facilitates its use as fusion enzymes encoded as gene therapy or administered as enzyme replacement therapy. More broadly, the LT peptide may aid in developing therapies for numerous lysosomal diseases.

Gangliosidoses, including GM1-gangliosidosis and GM2-gangliosidosis (Tay-Sachs disease and Sandhoff disease), are lysosomal disorders resulting from enzyme deficiencies and accumulation of gangliosides. Phenotypes of gangliosidoses range from infantile, late-infantile, juvenile, and to the adult form. The genotype-phenotype correlation is essential for prognosis and clinical care planning for patients with a gangliosidosis condition. Previously, we have developed a method to establish the genotype-phenotype correlation of another lysosomal disease, mucopolysaccharidosis type I, with in silico tools. This same method was applied to analyze the genotype and phenotype of 38 patients diagnosed with a gangliosidosis disease in the United States. Out of 40 mutations identified, 3 were novel, including p.Tyr192His and p.Phe556Ser of the GLB1 gene and p.Gly461Val of the HEXA gene. Furthermore, the mutant protein structure of all missense mutations was constructed by homology modeling. A systemic structural analysis of these models revealed the specific mechanisms of how each mutation may lead to the disease. In summary, the method developed in this study holds promise as a tool that can be broadly applicable to other lysosomal diseases and monogenic diseases. Keywords: Genotype-phenotype correlation, In silico, Gangliosidosis, Disease subtype, Lysosomal disorder

Background The gangliosidoses are rare inherited diseases that result in pathologic accumulation of gangliosides in the central nervous system and other tissues, leading to severe and progressive neurological impairment and early death in the childhood forms. No treatments are currently approved for the gangliosidoses, and development of treatments is impaired by limited understanding of the natural history of these diseases. Objective The objective of this study is to improve understanding of the juvenile gangliosidoses phenotypes and the late-infantile phenotypic subtype. Methods Through a prospective natural history study of subjects with juvenile GM1- and GM2-gangliosidosis, a timeline of clinical changes was developed for the classic juvenile phenotypes and the late-infantile phenotypes and results of serial neurodevelopmental testing was analyzed. Results Several candidate ‘outcome measures’ were identified: changes in ambulation and verbalization skills, the communication domain from neurodevelopmental testing and the caregiver-reported socialization domain. Conclusions The most common symptoms leading caregivers to seek a genetic diagnosis were changes in ambulation and verbalization.

Following allogenic bone marrow transplantation with predominance of donor engraftment, patients with Maroteaux-Lamy syndrome (MPS VI) demonstrate several indications of metabolic correction, such as reduction in liver size, and achieve near-normal levels of urine glycosaminoglycan (GAG). Thus it was somewhat surprising that, a single infusion of galsulfase after 20 years of full donor engraftment resulted in a further decline of GAG into the normal range (Whitley and Jarnes-Utz, Mol Genet Metab 101(4): 346-348, 2010). Urine GAG declined from slightly high pre-treatment levels (7.63 mg GAG/mmol creatinine range 7.0-8.5, N=3) progressively declining below the age-specific normal range (

GM1-gangliosidosis and GM2-gangliosidosis (Tay-Sachs disease and Sandhoff disease) are unrelenting heritable neurodegenerative conditions of lysosomal ganglioside accumulation. Although progressive brain atrophy is characteristic, longitudinal quantification of specific brain structures has not been systematically studied.The goal of this longitudinal study has been to quantify and track brain MRI volume changes, including specific structure volume changes, at different times in disease progression of childhood gangliosidoses, and to explore quantitative brain MRI volumetry (qMRI) as a non-invasive marker of disease progression for future treatment trials.Brain qMRI studies were performed in 14 patients with gangliosidoses (9 infantile, 5 juvenile) yearly. Cerebellar cortex and white matter, caudate, putamen, corpus callosum, ventricles, total brain, and intracranial volumes were measured, as well as total brain volume. Age-matched controls were available for the patients with the juvenile phenotype.The infantile phenotype of all gangliosidoses showed a consistent pattern of macrocephaly and rapidly increasing intracranial MRI volume with both (a) brain tissue volume (cerebral cortex and other smaller structures) and (b) ventricular volume (P0.01 for all). In contrast to apparent enlargement of the total brain volume, and chiefly the enlarged cerebral cortex, a subset of smaller brain substructures generally decreased in size: the corpus callosum, caudate and putamen became smaller with time. The volume of cerebellar cortex also decreased in patients with infantile GM1-gangliosidosis and juvenile GM1- and GM2-gangliosidosis; however, infantile GM2-gangliosidosis cerebellar cortex was the exception, increasing in size. Elevated intracranial pressure (estimated by lumbar spinal pressure) was a common finding in infantile disease and showed continued increases as the disease progressed, yet lacked MRI signs of hydrocephalus except for increasing ventricular size. Notably, in patients with juvenile gangliosidosis, macrocephaly and elevated intracranial pressure were absent and total brain volume decreased with time compared to controls (P=0.004).The disease course of infantile versus juvenile gangliosidoses is clearly distinguished by the rate of brain disease progression as characterized by qMRI. Assessments by qMRI represent a robust non-invasive method for monitoring CNS changes in the clinical course of gangliosidoses and is ideally suited to monitor effects of novel CNS-directed therapies in future clinical trials.

Enzyme replacement and substrate reduction therapies have significantly improved outcomes in patients with Type 1 Gaucher disease (GD1), but do not completely return glucocerebroside levels to the normal range nor prevent continuing tissue injury. Preliminary biomarker analyses indicate a role for oxidative stress and inflammation in GD1 pathophysiology. Thus, additional treatment strategies that target oxidative stress and inflammation have the potential to ameliorate both systemic and brain symptoms and may alter disease progression. N-acetylcysteine (NAC), a FDA-approved medication with antioxidant/anti-inflammatory activity, is also available as an inexpensive dietary supplement with a good safety profile. Our objective is to determine whether oxidative stress and inflammation in the blood and brains of individuals with GD1 can be altered with orally administered NAC. In this open-label study, we have currently enrolled 13 patients who received NAC at 3600mg/day dose. Blood sample collection for biomarker analysis and brain imaging using magnetic resonance spectroscopy (MRS) was done before and following 3 months of NAC. Total glutathione in red blood cells, which is a measure of oxidative stress, and plasma NAC concentrations were measured using a validated liquid chromatography/mass spectrometry method. Intracellular antioxidant enzyme activity and plasma markers for inflammation and lipid and protein modifications were measured using colorimetric and immunoassays. Interim analysis of data from 10 patients revealed that NAC at this dose did not significantly alter any of the neurochemical markers. However, validated MRS markers related to neuronal health such as N-acetyl aspartate (NAA) and glutamate showed trends toward improvement. Similar trends were also observed in blood antioxidant and inflammatory biomarkers. Pharmacokinetic analysis using serial samples collected 90-days after NAC regimen is underway to determine if a dose increase is warranted. If these favorable trends subsequently are shown to be significant, NAC could be a new adjunctive therapeutic option.