Thiamet G is a potent O-GlcNAcase inhibitor (Ki=20 nM) with selectivity. The role of O-GlcNAcase is to remove O-GlcNAc from the modified protein.

O-GlcNAcase (OGA, human):20 nM (Ki), O-GlcNAcase (OGA):21 nM (Ki), O-GlcNAcase (OGA):30 nM (EC50)

METHODS: PC-12 cells were treated with Thiamet G (1 nM-250 μM), and the level of O-GlcNAcase was detected by Western Blot. After PC-12 cells were treated with Thiamet G (100 mM), the level of O-GlcNAcase was detected by Western Blot.
RESULTS: Thiamet G inhibited O-GlcNAcase in PC-12 cells in a dose - and time-dependent manner and reduced the phosphorylation level of tau protein. [1]
METHODS: Mesangial cells were treated with Thiamet G (12.5, 25 nM), and the O-GlcNAcylation level and p38 phosphorylation level were detected by Western Blot.
RESULTS: Thiamet G significantly enhances p38 phosphorylation in glomerular cell lines by increasing O-GlcNAcylation. [2]

METHODS: To study the neuroprotective effect of Thiamet G, Thiamet G (20 mg/kg) was intraperitoneally injected into mice for 3 consecutive days.
RESULTS: Thiamet G exhibited neuroprotective effects by inhibiting O-GlcNAcase. [3]
METHODS: To study the antiepileptic effect of Thiamet G, Thiamet G (10 mg/kg) was intraperitoneally injected into an epileptic rat model once a day for three consecutive days.
RESULTS: Thiamet G significantly reduced the duration of epileptic seizures and the frequency of interictal spikes. [4]

All enzymatic assays are performed in triplicate at 37°C using 4-methylumbelliferyl N-acetyl-β-d-glucosaminide dehydrate as substrate. 1 nM of purified OGA is incubated with the compounds for 5 min, and then 0.2 mM of the substrate is added. The liberation of 4-methylumbellifery is monitored by kinetic reading at excitation/emission 355/460 nm using a Tecan M200 plate in a mode of 60 s/cycle and 15 cycles in total.

Jurkat cells are seeded at 6000 cells/well in a 96-well plate, and 12 h later, cells are treated with compounds for the indicated time. Cell viability is determined by XTT assay.