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Impairment of glycosaminoglycan synthesis in mucopolysaccharidosis type IIIA cells by using siRNA: a potential therapeutic approach for Sanfilippo disease.

ABSTRACT: Mucopolysaccharidoses (MPS) are severe inherited metabolic disorders from the group of lysosomal storage diseases. They are caused by deficiency in the activity of enzymes involved in the degradation of glycosaminoglycans (GAGs) and resultant accumulation of these compounds in the cells of patients. Although enzyme replacement therapy has become available for some MPS types (MPS I, MPS II and MPS VI), this treatment is not efficient when neurological symptoms occur, especially in MPS III (Sanfilippo disease). Recent studies indicated that substrate reduction therapy (SRT) may be an effective option for the treatment of neurodegenerative lysosomal storage diseases, including MPS III. However, previous attempts to SRT for MPS III focused on the use of non-specific inhibitors of GAG synthesis. Thus, we aimed to use the small interfering RNA (siRNA) procedure to control expression of particular genes, whose products are involved in GAG synthesis. In this report we show that, in MPS IIIA fibroblasts, we were able to reduce mRNA levels of four genes, XYLT1, XYLT2, GALTI and GALTII, whose products are involved in GAG synthesis. This decrease in levels of transcripts corresponded to a decrease in levels of proteins encoded by them. Moreover, efficiency of GAG production in these fibroblasts was considerably reduced after treatment of the cells with siRNA. These results indicate that efficient reduction of GAG synthesis may be achieved by the use of siRNA.

SUBMITTER: Dziedzic D

PROVIDER: S-EPMC2987185 | biostudies-other | 2010 Feb

REPOSITORIES: biostudies-other

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Project description:Mucopolysaccharidosis (MPS) IIIA, also known as Sanfilippo syndrome type A, is a severe, progressive disease that affects the central nervous system (CNS). MPS IIIA is inherited in an autosomal recessive manner and is caused by a deficiency in the lysosomal enzyme sulfamidase, which is required for the degradation of heparan sulfate. The sulfamidase is produced by the N-sulphoglucosamine sulphohydrolase (SGSH) gene. In MPS IIIA patients, the excess of lysosomal storage of heparan sulfate often leads to mental retardation, hyperactive behavior, and connective tissue impairments, which occur due to various known missense mutations in the SGSH, leading to protein dysfunction. In this study, we focused on three mutations (R74C, S66W, and R245H) based on in silico pathogenic, conservation, and stability prediction tool studies. The three mutations were further subjected to molecular dynamic simulation (MDS) analysis using GROMACS simulation software to observe the structural changes they induced, and all the mutants exhibited maximum deviation patterns compared with the native protein. Conformational changes were observed in the mutants based on various geometrical parameters, such as conformational stability, fluctuation, and compactness, followed by hydrogen bonding, physicochemical properties, principal component analysis (PCA), and salt bridge analyses, which further validated the underlying cause of the protein instability. Additionally, secondary structure and surrounding amino acid analyses further confirmed the above results indicating the loss of protein function in the mutants compared with the native protein. The present results reveal the effects of three mutations on the enzymatic activity of sulfamidase, providing a molecular explanation for the cause of the disease. Thus, this study allows for a better understanding of the effect of SGSH mutations through the use of various computational approaches in terms of both structure and functions and provides a platform for the development of therapeutic drugs and potential disease treatments.

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Impairment of glycosaminoglycan synthesis in mucopolysaccharidosis type IIIA cells by using siRNA: a potential therapeutic approach for Sanfilippo disease.

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