| In Vitro | In vitro activity: VER-155008 is a potent inhibitor of HSP70 family with IC50 of 0.5 μM, 2.6 μM, and 2.6 μM in cell-free assays for HSP70, HSC70 (Heat shock cognate 70), and GRP78, respectively, it has >100-fold selectivity over HSP90. VER 155008 has been shown to contribute to cancer cell survival via anti-apoptotic functions, that inhibits Hsp70 with IC50 of 0.5 μM as well as heat shock cognate 71 kDa protein (Hsc70) and 78 kDa glucose-regulated protein (Grp78) but to a lesser extent. VER-155008 significantly promoted axonal regrowth in amyloid β-treated neurons in vitro and improved object recognition, location, and episodic-like memory in 5XFAD mice. Furthermore, VER-155008 penetrated into the brain after intraperitoneal administration, suggesting that VER-155008 acts in the brain in situ. Immunohistochemistry revealed that VER-155008 reduced bulb-like axonal swelling in the amyloid plaques in the perirhinal cortex and CA1 in 5XFAD mice, indicating that VER-155008 also reverses axonal degeneration in vivo. Moreover, the two main pathological features of AD, amyloid plaques and paired helical filament tau accumulation, were reduced by VER-155008 administration in 5XFAD mice. This is the first report to show that the inhibition of HSC70 function may be critical for axonal regeneration and AD-like symptom reversal. This study provides evidence that HSC70 can be used as a new therapeutic target for AD treatment. Kinase Assay: VER-155008 is a potent inhibitor of HSP70 family with IC50 of 0.5 μM, 2.6 μM, and 2.6 μM in cell-free assays for HSP70, HSC70 (Heat shock cognate 70), and GRP78, respectively, it has >100-fold selectivity over HSP90. VER 155008 has been shown to contribute to cancer cell survival via anti-apoptotic functions, that inhibits Hsp70 with IC50 of 0.5 μM as well as heat shock cognate 71 kDa protein (Hsc70) and 78 kDa glucose-regulated protein (Grp78) but to a lesser extent.
Cell Assay: Embryos are removed from a pregnant ddY mouse at 14 days of gestation. Cells are treated with or without 10 μM Aβ25-35 for 3 days, followed by the addition of 0.05, 0.5, or 5 μM VER-155008 or vehicle solution (0.1% DMSO) for 4 days. The Aβ25-35 is incubated at 37°C for 4 days prior to treatment to facilitate aggregation. The cells are fixed with 4% paraformaldehyde and immunostained at 4°C for 24 h with antibodies against the axonal marker, mouse phosphorylated neurofilament heavy subunit, and against the neuronal marker, rabbit microtubule-associated protein 2. Alexa Fluor 488-conjugated goat anti-mouse IgG (1:400) and Alexa Fluor 568-conjugated goat anti-rabbit IgG (1:400) are used as secondary antibodies. Fluorescence images (864.98 μm × 645.62 μm) are captured using a fluorescence microscopy system. The lengths of the pNF-H-positive axons are measured using MetaMorph version 7.8 |
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| In Vivo | VER-155008 (25 mg/kg, i.v.) exhibits plasma clearance in naive female BALB/c mice. VER-155008 (40 mg/kg, i.v.) also shows rapid plasma clearance, and reduces the tumor levels in the HCT116 tumor bearing nude BALB/c mice[1]. VER-155008 (10 μmol/kg/day, i.p.) rescues memory deficits, and reduces axonal swelling associated with amyloid plaques in 5XFAD mice. VER-155008 (89.9 μmol/kg/day, i.p.) penetrates into the brain after administration in 5XFAD mice. VER-155008 also decreases amyloid plaques and PHF-tau associated with amyloid plaques in 5XFAD mice. |
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