title: Pharmacological chaperones increase residual β-galactocerebrosidase activity in fibroblasts from Krabbe patients.
abstract: Krabbe disease or globoid cell leukodystrophy is a degenerative, lysosomal storage disease resulting from the deficiency of β-galactocerebrosidase activity. This enzyme catalyzes the lysosomal hydrolysis of galactocerebroside and psychosine. Krabbe disease is inherited as an autosomal recessive trait, and many of the 70 disease-causing mutations identified in the GALC gene are associated with protein misfolding. Recent studies have shown that enzyme inhibitors can sometimes translocate misfolded polypeptides to their appropriate target organelle bypassing the normal cellular quality control machinery and resulting in enhanced activity. In search for pharmacological chaperones that could rescue the β-galactocerebrosidase activity, we investigated the effect of α-Lobeline or 3',4',7-trihydroxyisoflavone on several patient-derived fibroblast cell lines carrying missense mutations, rather than on transduced cell lines. Incubation of these cell lines with α-lobeline or 3',4',7-trihydroxyisoflavone leads to an increase of β-galacocerebrosidase activity in p.G553R + p.G553R, in p.E130K + p.N295T and in p.G57S + p.G57S mutant forms over the critical threshold. The low but sustained expression of β-galactocerebrosidase induced by these compounds is a promising result; in fact, it is known that residual enzyme activity of only 15-20% is sufficient for clinical efficacy. The molecular interaction of the two chaperones with β-galactocerebrosidase is also supported by in silico analysis. Collectively, our combined in silico-in vitro approach indicate α-lobeline and 3',4',7-trihydroxyisoflavone as two potential pharmacological chaperones for the treatment or improvement of quality of life in selected Krabbe disease patients.
authors.affiliation: Department of Molecular and Developmental Medicine, University of Siena, Siena, Italy.
authors.forename: Anna Sara
authors.initials: AS
authors.lastname: Berardi
authors.affiliation: Department of Bio-Medical Sciences, Section of Physiology, University of Catania, Catania, Italy.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Pannuzzo
authors.affiliation: Department of Bio-Medical Sciences, Section of Physiology, University of Catania, Catania, Italy.
authors.forename: Adriana
authors.initials: A
authors.lastname: Graziano
authors.affiliation: Department of Molecular and Developmental Medicine, University of Siena, Siena, Italy.
authors.forename: Elvira
authors.initials: E
authors.lastname: Costantino-Ceccarini
authors.affiliation: Department of Molecular and Developmental Medicine, University of Siena, Siena, Italy.
authors.forename: Paola
authors.initials: P
authors.lastname: Piomboni
authors.affiliation: Department of Molecular and Developmental Medicine, University of Siena, Siena, Italy. Electronic address: alice.luddi@unisi.it.
authors.forename: Alice
authors.initials: A
authors.lastname: Luddi
mesh_terms: D000818:Animals; D019556:COS Cells; D002470:Cell Survival / Q000187:drug effects; D002522:Chlorocebus aethiops; D003198:Computer Simulation; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D005698:Galactosylceramidase / Q000378:metabolism*; D006720:Homozygote; D006801:Humans; D007529:Isoflavones / Q000737:chemistry / Q000494:pharmacology* / Q000627:therapeutic use; D007965:Leukodystrophy, Globoid Cell / Q000188:drug therapy / Q000201:enzymology* / Q000473:pathology; D008120:Lobeline / Q000737:chemistry / Q000494:pharmacology* / Q000627:therapeutic use; D051379:Mice; D008958:Models, Molecular; D020125:Mutation, Missense / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D007529:Isoflavones; D008120:Lobeline; D005698:Galactosylceramidase; C061213:3',4',7-trihydroxyisoflavone
keywords: Krabbe disease; Lysosomal storage disease; Missense mutations; Pharmacological chaperones; β-Galactocerebrosidase