Objective: To explore whether inducing brain GCase activity by Ambroxol™ (ABX; a brain-penetrant GCase activator) can reduce a-synuclein (αSyn) serine-129 phosphorylation and oligomer accumulation in LRRK2^R1441G mutant mice of Parkinson’s disease (PD).

Background: Brain accumulation of phosphorylated αSyn aggregates is a pathological hallmark of PD. Ser129-phosphorylated αSyn (pSer129) is a surrogate marker of synucleinopathies. Leucine-rich repeat kinase-2 (LRRK2) mutation is a genetic risk of both familial and sporadic PD. Evidence suggests an interplay among LRRK2 mutation, glucocerebrosidase (GCase) deficiency, and synucleinopathies [1]. Aged mutant LRRK2^R1441G mice showed greater accumulation of αSyn oligomers in striatum [2]. Here, we tested whether activation of GCase can ameliorate synucleinopathies in LRRK2 mutant brains.

Method: LRRK2^R1441G knockin mutant (KI) and wildtype (WT) mice were fed with ABX for 18 weeks before assays of striatal GCase level and activity by ELISA. Brain penetration of ABX was determined by mass spectrometry (LC-MS-MS). Lysosomal activity was assessed in immortalized KI and WT mouse embryonic fibroblasts (MEFs) using real-time DQ™ Red BSA (a lysosomal protease substrate) assay. pSer129 level was determined by Western blotting. αSyn oligomer level was quantified by commercial ELISA.

Results: Treatment of ABX increased GCase activity and reduced pSer129-αSyn levels in KI MEFs. These KI MEFs also exhibited impaired lysosomal activity, which was attenuated by ABX treatment. ABX-containing food pellets enabled ad libitum feeding in mice without physical trauma during long-term treatment. Feeding of ABX (average weekly dose; 300mg/kg) showed significant brain penetration and elevated GCase activity, and reduced αSyn oligomer accumulation in mutant LRRK2 mouse striatum. This ABX regime did not cause significant changes in body weight and abnormal phenotypic changes.

Conclusion: Long-term therapeutic treatment of ABX reduced pSer129 and αSyn oligomer levels in aged LRRK2^R1441G mutant mouse brain. Our findings advocate new therapeutic approach to ameliorate synucleinopathies by increasing GCase activity and lysosomal degradation in PD.

References: [1] Pang SYY, Lo RCN, Ho PWL, Liu HF, Chang EES, Leung CT, Malki Y, Choi ZYK, Wong WY, Kung MHW, Ramsden DB, Ho SL*. LRRK2, GBA and their interaction in the regulation of autophagy: implications on therapeutics in Parkinson’s disease. Translational Neurodegeneration 2022; 11:5.

[2] Ho PWL, Leung GC, Liu HF, Pang SY, Lam CS, Xian JW, Li LF, Kung MHW, Ramsden DB, Ho SL*. Age-dependent accumulation of oligomeric α-synuclein from impaired degradation in mutant LRRK2 knockin mouse model of Parkinson disease: role for therapeutic activation of chaperone-mediated autophagy (CMA). Autophagy 2020; 16(2):347-370.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/glucocerebrosidase-gcase-activator-ambroxol-reduces-alpha-synuclein-serine-129-phosphorylation-and-oligomers-in-mutant-lrrk2-r1441g-mouse-brains/