Glucosylceramide synthase
Ibiglustat (CAS#: 1401090-53-6) targets glucosylceramide synthase (GCS), the rate‑limiting enzyme in glycosphingolipid synthesis.[4]
No IC₅₀, Ki, EC₅₀, or DC₅₀ values are reported in the provided literature.

In untreated WT cells, Ibiglustat (1 μM, 15 days; Fabry disease (FD) cells) produced GL-3 levels that were almost physiological, suggesting that Ibiglustat inhibits further GL-3 accumulation and could enhance GL-3 presence in FD cells. A significant portion of this sphingolipid[4]

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In Fabry disease iPSC‑derived cardiomyocytes (carrying GLA mutations W162X or R220X), treatment with 1 μM Ibiglustat (CAS#: 1401090-53-6) (as SAR402671) for 15 days prevented GL‑3 accumulation that otherwise occurred between day 18 and day 33 of differentiation, as measured by mass spectrometry (prevention study).[4]
In the same cell model, when treatment started on day 28 (after GL‑3 accumulation had plateaued), 1 μM Ibiglustat (CAS#: 1401090-53-6) for 15 days reduced accumulated GL‑3 by approximately 2.3‑ to 2.8‑fold, i.e. >50% reduction (clearance study). Immunocytochemistry showed disappearance of GL‑3 puncta and colocalization with LAMP2; electron microscopy revealed marked reduction of zebra bodies (lamellated lysosomal inclusions).[4]

Mutations in GBA, the gene encoding the lysosomal enzyme glucocerebrosidase (GCase), represent the greatest genetic risk factor for developing synucleinopathies including Parkinson's disease (PD). Additionally, PD patients harboring a mutant GBA allele present with an earlier disease onset and an accelerated disease progression of both motor and non-motor symptoms. Preclinical studies in mouse models of synucleinopathy suggest that modulation of the sphingolipid metabolism pathway via inhibition of glucosylceramide synthase (GCS) using a CNS-penetrant small molecule may be a potential treatment for synucleinopathies. Here, we aim to alleviate the lipid storage burden by inhibiting the de novo synthesis of the primary glycosphingolipid substrate of GCase, glucosylceramide (GlcCer). We have previously shown that systemic GCS inhibition reduced GlcCer and glucosylsphingosine (GlcSph) accumulation, slowed α-synuclein buildup in the hippocampus, and improved cognitive deficits. Here, we studied the efficacy of a brain-penetrant clinical candidate GCS inhibitor, venglustat, in mouse models of GBA-related synucleinopathy, including a heterozygous Gba mouse model which more closely replicates the typical GBA-PD patient genotype. Collectively, these data support the rationale for modulation of GCase-related sphingolipid metabolism as a therapeutic strategy for treating GBA-related synucleinopathies [1].
In GBA-related synucleinopathy mice (GbaD409V/WT), oral administration of Ibiglustat (via tool compound GZ667161, 0.033% wt/wt in diet for 9 months) reduced brain GlcCer to 58±2% of controls (p<0.0001) and numerically decreased proteinase K-resistant α-synuclein (p=0.2), tau (p=0.3), and significantly reduced ubiquitin aggregates (p=0.03) in hippocampus. [1]
In Gaucher-related synucleinopathy mice (GbaD409V/D409V), Ibiglustat (venglustat, 0.03% wt/wt in diet for 8 months) significantly reduced brain GlcCer (73±2% remaining, p<0.0001) and GlcSph (63±2% remaining, p<0.0001); plasma GlcCer (13±1% remaining) and GlcSph (17±1% remaining). It significantly reduced hippocampal protein aggregates: proteinase K-resistant α-synuclein (p=0.009), ubiquitin (p=0.004), and tau (p<0.0001). In the novel object recognition test after 7 months of treatment, Ibiglustat corrected the memory deficit in GbaD409V/D409V mice (preference for novel object, p<0.0001 vs 50% theoretical mean). [1]
In healthy volunteers, repeated once-daily oral doses of Ibiglustat (venglustat L-malate 5, 10, 20 mg for 14 days) caused time- and dose-dependent reductions in plasma GL-1 and GM3. Mean Day 15 GL-1 reductions from baseline: 41.9% (5 mg), 69.6% (10 mg), 74.6% (20 mg). GM3 reductions: 42.7%, 49.4%, 57.8% of baseline, respectively. [2]

Measurement of glycosphingolipid levels[1]
Quantitative analysis of sphingolipids was performed by liquid chromatography and tandem mass spectrometry (LC–MS/MS)28. Briefly, brain tissue was homogenized in 10 volumes of water (w/v). Ten microliters of homogenate or plasma was extracted with 1 ml of extraction solution (50:50 acetonitrile/methanol) by protein precipitation. Mouse CSF sphingolipids were extracted by liquid–liquid extraction, as previously described37. GlcCer and galactosylceramide were separated using a Waters Acquity UPLC and Cortecs HILIC column (2.1 mm × 100 mm, 2.7 µm particles) and analyzed by an API 5000 triple quadrupole mass spectrometer in MRM mode. GlcSph and psychosine were separated by a Waters Acquity UPLC and BEH HILIC column (2.1 mm × 100 mm, 1.7 µm particles) and analyzed by an API 6500 triple quadrupole mass spectrometer in MRM mode. GlcCer and GlcSph standards were purchased from Matreya, LLC and Avanti Polar Lipids, respectively. All procedures were performed blinded to the genotype or treatment.

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No enzyme assays (e.g. direct GCS inhibition, IC₅₀ determination) are described in the provided literature.

To expose insoluble α-synuclein aggregates, some tissues were pretreated with proteinase K (1:4 dilution) for 7 min at room temperature to digest soluble α-synuclein23. Brain sections were blocked with 10% (vol/vol) normal donkey serum for 1 h at room temperature and incubated with the following antibodies: mouse anti-ubiquitin (1:300; cat# MAB1510), rabbit anti-α-synuclein (1:300), and mouse anti-tau (1:500, Tau-5). Brain sections were then incubated for 1 h with either a donkey anti-mouse Alexa Fluor-488 (1:250 dilution,) or donkey anti-rabbit biotinylated secondary antibody (1:200 dilution). For α-synuclein aggregate quantification, a cyanine 3-tyramide signal amplification kit was used. Cell nuclei were counterstained with 4’, 6-diamino-2-phenylindole (DAPI). Sections were coverslipped with aqua-poly/mount and the stratum radiatum external to the CA1 hippocampal cell body layer was imaged with a SPOT camera (SPOT Imaging) paired with a Nikon Eclipse E800 fluorescence microscope equipped with a 20 × objective lens, as previously described22. Two to three sections were imaged per animal and immunofluorescence was quantitatively measured via threshold fluorescent area on MetaMorph Software. All procedures were performed blinded to the treatment or genotype and the percent threshold area is expressed as the mean ± SE. [1]

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Generation of Fabry iPSC‑derived cardiomyocytes: Skin fibroblasts from two male Fabry patients (GLA mutations W162X and R220X) were reprogrammed using a lentiviral OKSM polycistronic vector. iPSCs were differentiated into beating embryoid bodies (EBs) using a 3D protocol with sequential cytokine cocktails (BMP4, bFGF, activin A, DKK‑1, VEGF) under hypoxic (5% O₂) conditions for the first 12 days, then normoxia. Beating EBs appeared 10‑16 days after differentiation start. One‑month‑old beating EBs were used for experiments.[4]
Prevention study: 18 days after differentiation start, beating EBs were treated with 1 μM Ibiglustat (CAS#: 1401090-53-6) (SAR402671) for 15 days. Culture medium with drug was replaced every 2 days. GL‑3 accumulation was assessed by immunocytochemistry (anti‑CD77/GL‑3, LAMP2), electron microscopy, and mass spectrometry. Untreated WT and Fabry EBs served as controls.[4]
Clearance study: 28 days after differentiation start (when GL‑3 accumulation plateaued), beating EBs were treated with 1 μM Ibiglustat (CAS#: 1401090-53-6) for 15 days (medium changed every 2 days). GL‑3 levels were quantified by LC‑MS/MS. In parallel, agalsidase beta (0.3 μg/ml, 5 days) was used as a positive control.[4]
Immunocytochemistry: Dissociated EB cells were fixed, permeabilized, and stained with anti‑GL‑3 and anti‑LAMP2, followed by Alexa Fluor secondary antibodies. Colocalization was assessed by confocal microscopy.[4]
Electron microscopy: EBs were fixed in glutaraldehyde, post‑fixed in osmium tetroxide, embedded in epon, sectioned, and examined with a transmission electron microscope for lysosomal inclusions (zebra bodies).[4]
Mass spectrometry: EB pellets were extracted with acetonitrile:methanol (1:1). Total GL‑3 (nine isoforms) and lyso‑GL‑3 were quantified by UPLC‑MS/MS in MRM positive‑ion mode. Total GL‑3 was normalized to total phosphatidylcholine (PC) using an external calibration curve.[4]

Administration of the glucosylceramide synthase inhibitors: venglustat and tool compound GZ667161[1]
A subset of animals received glucosylceramide synthase inhibitors, venglustat (aka GZ402671) or GZ667161, via pelleted diet at 0.03%- or 0.033%-wt/wt, respectively. For each experiment, sex and siblings were randomly matched for group assignment. Target engagement and exposure confirmation studies included GbaD409V/D409V or GbaD409V/WT mice administered venglustat for two consecutive weeks beginning at approximately 4 months of age. Mice included in sustained GCS inhibition studies were administered either GZ667161 or venglustat upon weaning at ~ 4 weeks of age. Wild-type, baseline, and control groups were fed vehicle rodent chow. GCS inhibitor and vehicle diets were continuously provided to mice until necropsy and tissue collection.

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CSF collection[1]
Animals were anesthetized via an intraperitoneal injection of a 10:1 Ketamine/Xylazine cocktail prior to being placed into a surgical ear bar rig. After making a midline cut to remove a small patch of skin from the head, the fat and muscle layers were opened using a cautery pen to reveal the base of the skull and occipital crest. The remaining tissue was then removed to expose the cisterna magna membrane. Using a pulled glass pipette, the cisterna magna membrane was punctured to allow CSF to flow freely into the pipette via capillary action. After collecting approximately 10–20 uL, CSF was transferred to a clean protein lo-bind tube . CSF samples with visible blood contamination were excluded from analyses.

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Animal perfusion and tissue and blood collection[1]
Prior to whole blood collection, mice were anesthetized via a 200 uL intraperitoneal injection of sodium pentobarbital. Following the loss of response to a foot-pinch and corneal reflex, approximately 250 uL of whole blood was collected from the retro-orbital sinus using a glass capillary tube into a Microtainer® tube containing K2 EDTA anticoagulant. Whole blood samples were collected retro-orbitally and immediately placed on ice. Plasma was isolated after 5 min centrifugation at 8000 RPM at 4 °C. Immediately following blood collection, animals were transcardially perfused with cold phosphate-buffered saline (PBS) at a rate of 18 mL/minute, for two minutes. After cutting the brains sagittally along the midline, the left hemisphere was microdissected into various regions, snap-frozen in liquid nitrogen, and stored at − 80 °C until use. The right hemisphere was post-fixed in 10% neutral-buffered formalin for 48–72 h. Right hemispheres were then washed three times in 1X PBS and transferred to 30% sucrose for 24–48 h. Right hemispheres were embedded in O.C.T. and sectioned into 20 µm sections using a cryostat, as previously described.

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For GZ667161 (tool compound) in GbaD409V/WT mice: starting at 4 weeks of age, mice were fed pelleted diet containing 0.033% wt/wt GZ667161 for 9 months. Control mice received vehicle chow. At sacrifice (10 months), drug exposures were 223±81 ng/mL plasma and 237±72 ng/g brain. [1]
For venglustat in GbaD409V/WT mice (2-week study): 4-month-old mice were fed pelleted diet with 0.03% wt/wt venglustat for 2 weeks. Exposures: plasma 1616±221 ng/mL, brain 901±110 ng/g. CSF not measured for drug. [1]
For venglustat in GbaD409V/D409V mice (2-week study): 4-month-old mice fed 0.03% wt/wt venglustat diet for 2 weeks. Exposures: plasma 1706±51 ng/mL, brain 1184±56 ng/g, CSF 56±3 ng/mL. [1]
For long-term venglustat in GbaD409V/D409V mice: starting at 4 weeks of age, mice fed 0.03% wt/wt venglustat diet for 8 months. Exposures: plasma 944±53 ng/mL, brain 617±41 ng/g. [1]
Ex vivo binding study in mice: whole brains (without cerebellum) collected 1 h after oral drug administration, homogenized, centrifuged, membranes incubated with 0.3 nM [³H]nociceptin to determine NOP receptor inhibition. [1]
Vogel conflict test in mice: Mice water-deprived for 24 h, training session (15 min no shock), then test session (30 min, 0.5 mA shock per 10 licks). Drugs (MCOPPB, not Ibiglustat) given 1 h before test. [1]
Novel object recognition test in mice: Mice acclimated to arena for 10 min, then exposed to two identical objects for 5 min (training). After 24 h retention, one object replaced with novel object, investigations counted for 5 min. Preference calculated as % investigations of novel object. [1]
For phase 1 human studies (healthy volunteers): Single ascending dose (2,5,15,25,50,100,150 mg venglustat L-malate) after ≥10 h fast; blood collected at 0,0.5,1,2,3,4,5,6,8,10,12,16,24,48,72,96 h postdose. Food-effect study: 5 mg with high-fat breakfast (≈815 kcal) or fasting, crossover with 7-day washout. Repeated-dose study: 5,10,20 mg once daily for 14 days; blood collected on Days 1 and 14 at multiple timepoints, and trough on Days 2-5,8,11,13. Urine collected 0-24 h on Day 14. Safety monitored via TEAEs, ECG, labs, vital signs. [2]

In healthy volunteers, Ibiglustat (venglustat) showed linear pharmacokinetics after single oral doses (2-150 mg venglustat L-malate). Median tmax 3.00-5.50 h. Geometric mean t1/2 28.9 h. Mean CL/F 5.18-6.43 L/h. Dose-proportional exposure (75-fold dose increase gave 97.3-, 89.2-, 85.9-fold increases in Cmax, AUClast, AUCinf). Food had no effect on exposure (fed/fasted geometric mean ratios: Cmax 0.92, AUClast 0.91). [2]
After repeated once-daily doses (5,10,20 mg for 14 days), steady state reached within 5 days. Accumulation ratios (Day14/Day1) pooled: Cmax 2.10, AUC0-24 2.22, independent of dose and sex. Mean fraction of dose excreted unchanged in urine (fe0-24) 26.3-33.1%. Renal clearance (CLR(0-24)) 1.49-2.07 L/h. No marked change in 4β-hydroxycholesterol (CYP3A4 induction marker). [2]
In GbaD409V/WT mice, GZ667161 (0.033% diet for 9 months) gave brain exposure 237±72 ng/g. [1]
In GbaD409V/WT mice, venglustat (0.03% diet for 2 weeks) gave plasma 1616±221 ng/mL, brain 901±110 ng/g. [1]
In GbaD409V/D409V mice, venglustat (0.03% diet for 2 weeks) gave plasma 1706±51 ng/mL, brain 1184±56 ng/g, CSF 56±3 ng/mL. After 8 months treatment: plasma 944±53 ng/mL, brain 617±41 ng/g. [1]

In phase 1 healthy volunteer studies (single ascending dose, food-effect, repeated dose), Ibiglustat (venglustat) showed no deaths, no serious adverse events, no severe TEAEs, and no discontinuations due to TEAEs. In single ascending dose (2-150 mg), mild TEAEs reported in 3 subjects (rhinitis, feeling cold, headache); only one headache (100 mg) was drug-related. In food-effect study, one mild contact dermatitis (unrelated). In repeated-dose study (5,10,20 mg for 14 days), 17 subjects reported 32 mild TEAEs: placebo 6/9 (67%), venglustat groups 11/27 (41%). Drug-related TEAEs included constipation, diarrhea, dry mouth, flatulence, pruritus, fatigue. No clinically relevant hematologic, biochemical, ECG, or vital sign abnormalities. [2]
In mouse models, no overt toxicity or adverse effects reported; Ibiglustat did not affect locomotor activity at anxiolytic doses (MCOPPB data, not directly for ibiglustat). [1]

[1]. Preclinical pharmacology of glucosylceramide synthase inhibitor venglustat in a GBA-related synucleinopathy model. Sci Rep. 2021;11(1):20945. Published 2021 Oct 22.

[2]. Pharmacokinetics, Pharmacodynamics, Safety, and Tolerability of Oral Venglustat in Healthy Volunteers. Clin Pharmacol Drug Dev. 2021;10(1):86-98.

[3]. Molecular mechanisms of α-synuclein and GBA1 in Parkinson's disease. Cell Tissue Res. 2018;373(1):51-60. doi:10.1007/s00441-017-2704-y.

[4]. Effective clearance of GL-3 in a human iPSC-derived cardiomyocyte model of Fabry disease. J Inherit Metab Dis. 2014 Nov;37(6):1013-22.

Venglustat is being investigated in the clinical trial NCT01674036 (Genz-682452: safety, tolerability, and pharmacokinetics in healthy men).

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Drug Indications
Treatment of autosomal dominant polycystic kidney disease
Treatment of galactosialidosis, GM1 ganglioside deposition, GM2 ganglioside deposition, sialic acid deposition
Treatment of polycystic kidney disease
Treatment of Parkinson's disease

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Ibiglustat (CAS#: 1401090-53-6) (also known as GZ/SAR402671) is a brain‑penetrant glucosylceramide synthase inhibitor that has been evaluated in preclinical models of synucleinopathy and Fabry disease. In a Gaucher‑related synucleinopathy mouse model (GbaD409V/D409V), a closely related tool compound GZ667161 (an analog with similar GCS inhibitory activity) reduced glycosphingolipids, pathological α‑synuclein aggregates, and memory deficits; however, no direct data for ibiglustat itself are provided in the attached literature.[3]
Ibiglustat (CAS#: 1401090-53-6) has been or is being investigated in clinical trials for GBA1‑associated Parkinson’s disease (Phase 2), neuronopathic Gaucher disease type 3 (Phase 2), Fabry disease, GM2 gangliosidosis, and autosomal dominant polycystic kidney disease, as mentioned in the literature.[3][4]
In the Fabry iPSC‑derived cardiomyocyte model, Ibiglustat (CAS#: 1401090-53-6) (1 μM) showed efficacy both in preventing and reversing GL‑3 accumulation, suggesting that substrate reduction therapy may be a viable approach for cardiac Fabry disease.[4]

C20H24FN3O2S

389.4869

389.157

C, 61.68; H, 6.21; F, 4.88; N, 10.79; O, 8.22; S, 8.23

1401090-53-6

Ibiglustat (L-Malic acid);1629063-79-1 (HCl); 1629063-78-0 (malate; Ibiglustat succinate;1629063-80-4; 1401090-53-6; 1629063-80-4 (succinic acid)

60199242

White to off-white solid

1.3±0.1 g/cm3

1.613

4.13

1

6

5

27

533

1

S1C(C2C([H])=C([H])C(=C([H])C=2[H])F)=NC(=C1[H])C(C([H])([H])[H])(C([H])([H])[H])N([H])C(=O)O[C@]1([H])C([H])([H])N2C([H])([H])C([H])([H])C1([H])C([H])([H])C2([H])[H]

YFHRCLAKZBDRHN-MRXNPFEDSA-N

InChI=1S/C20H24FN3O2S/c1-20(2,17-12-27-18(22-17)14-3-5-15(21)6-4-14)23-19(25)26-16-11-24-9-7-13(16)8-10-24/h3-6,12-13,16H,7-11H2,1-2H3,(H,23,25)/t16-/m1/s1

(3S)-1-azabicyclo[2.2.2]octan-3-yl N-{2-[2-(4-fluorophenyl)- 1,3-thiazol-4-yl]propan-2-yl}carbamate

Venglustat; SAR402671; GZ402671; Genz-682452-AA; SAR-402671; GZ402671; Ibiglustat; SAR 402671; GZ-402671; GZ-452; Genz-682452

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~50 mg/mL (~128.37 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (6.42 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (6.42 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (6.42 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)