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Intracellular Delivery of β‑Galactosidase Enzyme Using Arginase-Responsive Dextran Sulfate/Poly‑l‑arginine Capsule for Lysosomal Storage Disorder

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journal contribution
posted on 15.12.2017, 11:52 authored by Meenakshi Gupta, Himanshu Pandey, Sri Sivakumar
β-Galactosidase (β-gal) is one of the important lysosomal enzymes that is involved in the breakdown of glycosphingolipids (e.g., GM1 ganglioside), and its deficiency leads to GM1 Gangliosidosis, a lysosomal storage disorder (LSD). Intracellular delivery of β-gal is one of the preferable methods to treat this kind of LSDs. However, it cannot permeate the cell membrane due to its intricate macromolecular nature, low stability, and degradation by endogenous proteases. To this end, we report efficient intracellular delivery of β-gal via arginase-responsive dextran sulfate/poly-l-arginine polymer capsules (DS/PA capsules). The therapeutic activity of β-gal enzyme has been assessed in two gene-deficient diseased cell lines, SV (β-galactosidase gene-deficient mouse fibroblast) and R201C (deficient human β-galactosidase gene-introduced mouse fibroblast), and in wild-type mouse fibroblast immortalized cell lines. The activity of β-gal enzyme has been estimated within cells by using fluorescein isothiocyanate-cholera toxin B as a florescent probe that illustrates the level of GM1 ganglioside, the β-gal substrate. We found 1.8-, 3.4-, and 2.8-fold reduction in the substrate level in R201C, SV, and wild-type mouse fibroblast, respectively, which confirms the release and therapeutic activity of β-gal enzyme inside the cells. Moreover, enzyme delivery in gene-deficient diseased cell lines (SV and R201C) via DS/PA capsules reduced the level of enzyme substrate to a normal endogenous level, which is present in untreated wild-type mouse fibroblast cells. We note that loading of β-gal enzyme within DS/PA capsules was estimated to be 3 mU per hundred capsules and more than 77% of β-gal is released within 12 h. Overall, these results highlight the potential of DS/PA capsules as an efficient delivery carrier for therapeutic enzyme.