GSK-3β/HDAC-IN-2 is a potent inhibitor of GSK-3β (IC50 = 0.04 μM), HDAC2 (IC50 = 1.05 μM, Ki = 0.070 μM), and HDAC6 (IC50 = 1.52 μM, Ki = 0.017 μM). It effectively inhibits HDAC2 and HDAC6 activities and prevents the hyperphosphorylation of tau protein. GSK-3β/HDAC-IN-2 demonstrates neuroprotective properties without significant toxicity and is useful for Alzheimer's disease research.

HDAC6:0.017 μM (Ki)

GSK-3β/HDAC-IN-2 (Compound 19) demonstrates strong inhibitory effects on GSK-3β, HDAC2, and HDAC6 at a concentration of 15 μM, with inhibition rates of 100%, 86.51%, and 88.60%, respectively. It shows no significant cytotoxicity within the range of 0.1-100 μM for 24 hours and can induce cell proliferation at a lower concentration of 0.1 μM. GSK-3β/HDAC-IN-2, at 10 μM after 1 hour of pretreatment followed by 16 hours of co-incubation, completely counteracts tau protein phosphorylation induced by copper sulfate (400 μM, 16 h) in differentiated SH-SY5Y cells. At 0.5 μM for 24 hours, it reduces copper sulfate (600 μM)-induced neurotoxicity in SH-SY5Y cells. The compound (0.1-10 μM, 24 h) also stimulates the phosphorylation of GSK-3β (Ser9) and the acetylation of histone H3 and tubulin. In SH-SY5Y cells, GSK-3β/HDAC-IN-2 selectively induces apoptotic responses through chromatin remodeling, histone regulation, and disruption of cell cycle checkpoints during the mitotic stress response.