Lysosomal storage diseases (LSDs) are a group of inherited metabolic conditions caused by defects in proteins critical for lysosomal function. While individually rare, their collective incidence is just over 1/7,000 live births, which makes the LSDs an important health problem. The majority of these diseases have a progressive neurodegenerative course, resulting in severe debilitation. Symptoms are not manifested unless there is more than 90% reduction in the residual activity of the deficient enzyme. Thus, an enzyme activity level above the critical threshold will prevent lysosomal storage and dysfunction, and can arrest, or reverse the disease process. Current therapies available for LSDs, such as enzyme replacement therapy (ERT) are only available to treatment of a few LSDs, and then only the non-neurological symptoms. Additionally, the approximate cost of over US$250,000/year/patient limits accessibility to ERT. For these reasons, therapies based on small molecules, which could cross the blood-brain barrier and be less expensive;appear to be an attractive approach. Small molecules can function as enzyme enhancement agents by binding, and stabilizing mutant misfolded proteins in the endoplasmic reticulum (ER), allowing them to evade the ER associated degradation (ERAD), and reaches the lysosome. Previously, we screened a small compound library of 1,040-FDA approved compounds and were able to identify and characterize two drugs that function as enzyme enhancement agents for two lysosomal enzymes: hexosaminidase A and glucocerebrosidase. My hypothesis is that small molecules may enhance residual activity found in patients with LSDs by direct interactions with the mutant enzyme, or by an indirect effect on different cellular pathways resulting in increased levels of the mutant and partially functional protein in the lysosomal compartment. I hypothesize that I can identify molecular probes that would restore arylsulfatase A (ASA), lysosomal enzyme deficient in metachromatic leukodystrophy (MLD), a neurodegenerative LSD for which a specific treatment is not available. To test this hypothesis, I aim to: (i) develop a robust and reproducible cell-based high throughput screening (HTS) assay to identify molecular probes that may function as enzyme enhancement agents for ASA;(ii) to configure a quantitative HTS assay for ASA, and carry out validation strategies for the """"""""active"""""""" small molecules generated by the primary screening. I anticipate the submission of the screening project upon completion to the NIH Chemical Genomics Center for implementation. The candidate molecular probes generated by this screening will then be validated using different assays based on my experience previously acquired on the identification and characterization of enzyme enhancement agents for other lysosomal enzymes. The validated small molecules will be potential drug candidates to treat patients with MLD, and may also be used as tools to uncover novel biological pathways involved on the folding, maturation and trafficking of lysosomal proteins.
The development of high throughput screening assay (HTS) for arylsulfatase A (ASA) will allow the identification of molecular probes that, once characterized and validated, may be potential drug candidates to treat patients with metachromatic leukodystrophy, a lysosomal storage disease caused by deficiency of ASA. In addition, these compounds may be used as research tools to examine biological pathways involved on the folding, maturation and trafficking of lysosomal proteins.
| Kano, S; Yuan, M; Cardarelli, R A et al. (2015) Clinical utility of neuronal cells directly converted from fibroblasts of patients for neuropsychiatric disorders: studies of lysosomal storage diseases and channelopathy. Curr Mol Med 15:138-45 |
| Geng, Haifeng; Whiteley, Grace; Ribbens, Jameson et al. (2011) Novel patient cell-based HTS assay for identification of small molecules for a lysosomal storage disease. PLoS One 6:e29504 |
