Abstract

Gaucher disease (GD) is a rare monogenetic disorder leading to dysfunction of acid beta-glucosidase (beta-glucocerebrosidase; GCase) and accumulation of glucosylceramide in lysosomes, especially in macrophages (Gaucher cells). Many of the mutations at the origin of GD do not impair the catalytic activity of GCase, but cause misfolding and subsequent degradation by the quality control system at the endoplasmic reticulum. Pharmacological chaperones (PCs) capable of restoring the correct folding and trafficking of the endogenous mutant enzyme represent promising alternatives to the currently available enzyme replacement and substrate reduction therapies (ERT and SRT, respectively), but unfavorable biodistribution and potential side-effects remain important issues. We have now designed a strategy to enhance the controlled delivery of PCs to macrophages that exploit the formation of ternary complexes between the PC, a trivalent mannosylated beta-cyclodextrin (beta CD) conjugate and the macrophage mannose receptor (MMR). First, PC candidates with appropriate relative avidities towards the beta CD cavity and the GCase active site were selected to ensure efficient transfer of the PC cargo from the host to the GCase active site. Control experiments confirmed that the beta CD carrier was selectively recognized by mannose-specific lectins and that the corresponding PC: mannosylated beta CD supramolecular complex retained both the chaperoning activity, as confirmed in human GD fibroblasts, and the MMR binding ability. Finally, fluorescence microscopy techniques proved targeting and cellular uptake of the PC-loaded system in macrophages. Altogether, the results support that combined cyclodextrin encapsulation and glycotargeting may improve the efficacy of PCs for GD.