Ambroxol increases glucocerebrosidase (GCase) activity and restores GCase translocation in primary patient-derived macrophages in Gaucher disease and Parkinsonism.

A E Kopytova, G N Rychkov, M A Nikolaev, G V Baydakova, A A Cheblokov, K A Senkevich, D A Bogdanova, O I Bolshakova, I V Miliukhina, V A Bezrukikh, G N Salogub, S V Sarantseva, T C Usenko, E Y Zakharova, A K Emelyanov, S N Pchelina

Journal: Parkinsonism & related disorders 2021;84():112-121

PMID: 33609962

Full Text on Elsevier Science View on PubMed

Abstract

Mutations in the glucocerebrosidase gene (GBA) encoding the lysosomal enzyme glucocerebrosidase (GCase) cause Gaucher disease (GD) and are the most commonly known genetic risk factor for Parkinson disease (PD). Ambroxol is one of the most effective pharmacological chaperones of GCase. Fourteen GD patients, six PD patients with mutations in the GBA gene (GBA-PD), and thirty controls were enrolled. GCase activity and hexosylsphingosine (HexSph) concentration were measured in dried blood and macrophage spots using liquid chromatography coupled with tandem mass spectrometry. The effect of ambroxol on GCase translocation to lysosomes was assessed using confocal microscopy. The results showed that ambroxol treatment significantly increased GCase activity in cultured macrophages derived from patient blood monocytic cell (PBMC) of GD (by 3.3-fold) and GBA-PD patients (by 3.5-fold) compared to untreated cells (p < 0.0001 and p < 0.0001, respectively) four days after cultivation. Ambroxol treatment significantly reduced HexSph concentration in GD (by 2.1-fold) and GBA-PD patients (by 1.6-fold) (p < 0.0001 and p < 0.0001, respectively). GD macrophage treatment resulted in increased GCase level and increased enzyme colocalization with the lysosomal marker LAMP2. The possible binding modes of ambroxol to mutant GCase carrying N370S amino acid substitution at pH 4.7 were examined using molecular docking and molecular dynamics simulations. The ambroxol position characterized by minimal binding free energy was observed in close vicinity to the residue, at position 370. Taken together, these data showed that PBMC-derived macrophages could be used for assessing ambroxol therapy response for GD patients and also for GBA-PD patients.

Address: Petersburg Nuclear Physics Institute Named By B.P. Konstantinov of National Research Center «Kurchatov Institute», Gatchina, Russia. Electronic address: [email protected].; Petersburg Nuclear Physics Institute Named By B.P. Konstantinov of National Research Center «Kurchatov Institute», Gatchina, Russia; Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg, Russia; Kurchatov Genome Center - PNPI, Gatchina, Russia.; Petersburg Nuclear Physics Institute Named By B.P. Konstantinov of National Research Center «Kurchatov Institute», Gatchina, Russia; First Pavlov State Medical University of St. Petersburg, Saint-Petersburg, Russia.; Research Center for Medical Genetics, Moscow, Russia.; Petersburg Nuclear Physics Institute Named By B.P. Konstantinov of National Research Center «Kurchatov Institute», Gatchina, Russia.; Petersburg Nuclear Physics Institute Named By B.P. Konstantinov of National Research Center «Kurchatov Institute», Gatchina, Russia; Institute of Experimental Medicine, Saint-Petersburg, Russia; First Pavlov State Medical University of St. Petersburg, Saint-Petersburg, Russia.; Almazov National Medical Research Centre, Saint-Petersburg, Russia.

DOI: 10.1016/j.parkreldis.2021.02.003

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MeSH Terms: Adult, Aged, Aged, 80 and over, Ambroxol, Cells, Cultured, Enzyme Inhibitors, Female, Gaucher Disease, Glucosylceramidase, Humans, Macrophages, Male, Middle Aged, Molecular Chaperones, Parkinson Disease, Translocation, Genetic