Abstract
Morquio A syndrome, or mucopolysaccharidosis type IVA (MPS IVA), is a lysosomal storage disease due to mutations in the N-acetylgalactosamine-6-sulfatase (GALNS) gene. Systemic skeletal dysplasia and the related clinical features of MPS IVA are due to disruption of cartilage and its extracellular matrix, leading to an imbalance of growth. Enzyme replacement therapy (ERT) with recombinant human GALNS, alpha elosulfase, provides a systemic treatment. However, this therapy has a limited impact on skeletal dysplasia because the infused enzyme cannot penetrate cartilage and bone. Therefore, an alternative therapeutic approach to reach the cartilage is an unmet challenge. We have developed a new drug delivery system based on a nanostructure lipid carrier with the capacity to immobilize enzymes used for ERT and to target the lysosomes. This study aimed to assess the effect of the encapsulated enzyme in this new delivery system, using in vitro proteomic technology. We found a greater internalization of the enzyme carried by nanoparticles inside the cells and an improvement of cellular protein routes previously impaired by the disease, compared with conventional ERT. This is the first qualitative and quantitative proteomic assay that demonstrates the advantages of a new delivery system to improve the MPS IVA ERT.
Highlights
Morquio A syndrome, or mucopolysaccharidosis type IVA (MPS IVA, OMIM #253000), is an autosomal recessive disease, caused by mutations in the N-acetylgalactosamine-6-sulfatase (GALNS), gene which result in deficient activity of N-acetylgalactosamine-6-sulfatase (GALNS, E.C.3.1.6.4), an enzyme that degrades glycosaminoglycan (GAG) keratan sulate (KS) and chondroitin-6-sulfate (C6S) [1,2]
By using dependent acquisition (DDA)-liquid chromatography (LC)-mass spectrometry (MS)/MS, we identified 287 to 1623 proteins in untreated cells and 586 to 1817 proteins in fibroblasts treated with nanostructure lipid carrier (NLC) + enzyme replacement therapy (ERT) or ERT alone (Table 1)
We identified the changes in the cellular component of the expressed proteins related to the mitochondrion and the interaction between the mitochondria and the lysosome of the cells treated with NLC + ERT vs. ERT
Summary
Morquio A syndrome, or mucopolysaccharidosis type IVA (MPS IVA, OMIM #253000), is an autosomal recessive disease, caused by mutations in the N-acetylgalactosamine-6-sulfatase (GALNS), gene which result in deficient activity of N-acetylgalactosamine-6-sulfatase (GALNS, E.C.3.1.6.4), an enzyme that degrades glycosaminoglycan (GAG) keratan sulate (KS) and chondroitin-6-sulfate (C6S) [1,2]. Therapies for resolving bone lesions remain an unmet need, and an alternative therapeutic approach is required to overcome such limitations In this regard, one possible solution is a nanoparticular system that can protect the enzymes against their degradation or inactivation, minimize the associated immunological reactions, and increase cellular internalization to achieve maximum efficacy of the treatment [17]. Other quantitation techniques eliminate labeling (label-free) and rely on advanced software analysis as a new technology named SWATH-MS (sequential window acquisition of all theoretical mass spectra) [30,31] These methods measure the relative concentrations of peptide analytes within two or more samples. We have analyzed the effects of protein expression levels in the MPS IVA fibroblasts by providing the immobilized and free enzyme (GALNS) and by using qualitative and quantitative proteomic techniques
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